|
1
|
Kim SH, Yasunaga AB, Zhang H, Whitley KD, Li ITS. Quantitative Super-Resolution Imaging of Molecular Tension. ADVANCED SCIENCE (WEINHEIM, BADEN-WURTTEMBERG, GERMANY) 2025:e2408280. [PMID: 40245301 DOI: 10.1002/advs.202408280] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/29/2024] [Revised: 02/08/2025] [Indexed: 04/19/2025]
Abstract
DNA-based molecular tension probes have revolutionized the localization of mechanical events in live cells with super-resolution. However, imaging the magnitude of these forces at super-resolution has been challenging. Here, qtPAINT (quantitative tension points accumulation for imaging in nanoscale topography) is introduced as a strategy to image the magnitude of molecular tension with super-resolution accuracy. By leveraging the force-dependent dissociation kinetics of short DNA oligonucleotides on their complementary strands, tension is encoded on individual molecules through their binding kinetics. This method allowed for a quantitative analysis of these kinetics, providing a detailed reconstruction of the force magnitudes acting on each tension probe. The technique integrates a molecular-beacon PAINT imager with a hairpin molecular tension probe, achieving a force quantification range of 9-30 pN and maintaining a spatial resolution of 30-120 nm in low and high-density regions. Additionally, qtPAINT offers a temporal resolution on the order of a minute, enhancing its applicability for studying dynamic cellular processes.
Collapse
Affiliation(s)
- Seong Ho Kim
- Department of Chemistry, University of British Columbia, Kelowna, BC, V1V 1V7, Canada
- Department of Chemistry and Advanced Materials, College of Natural Sciences, Gangneung-Wonju National University, Gangneung, 25457, Republic of Korea
| | - Adam B Yasunaga
- Department of Chemistry, University of British Columbia, Kelowna, BC, V1V 1V7, Canada
| | - Hongyuan Zhang
- Department of Chemistry, University of British Columbia, Kelowna, BC, V1V 1V7, Canada
| | - Kevin D Whitley
- Centre for Bacterial Cell Biology, Biosciences Institute, Newcastle University, Newcastle upon Tyne, NE1 7RU, UK
| | - Isaac T S Li
- Department of Chemistry, University of British Columbia, Kelowna, BC, V1V 1V7, Canada
| |
Collapse
|
|
2
|
Liu X, Yu L, Xiao A, Sun W, Wang H, Wang X, Zhou Y, Li C, Li J, Wang Y, Wang G. Analytical methods in studying cell force sensing: principles, current technologies and perspectives. Regen Biomater 2025; 12:rbaf007. [PMID: 40337625 PMCID: PMC12057814 DOI: 10.1093/rb/rbaf007] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2024] [Revised: 12/16/2024] [Accepted: 02/10/2025] [Indexed: 05/09/2025] Open
Abstract
Mechanical stimulation plays a crucial role in numerous biological activities, including tissue development, regeneration and remodeling. Understanding how cells respond to their mechanical microenvironment is vital for investigating mechanotransduction with adequate spatial and temporal resolution. Cell force sensing-also known as mechanosensation or mechanotransduction-involves force transmission through the cytoskeleton and mechanochemical signaling. Insights into cell-extracellular matrix interactions and mechanotransduction are particularly relevant for guiding biomaterial design in tissue engineering. To establish a foundation for mechanical biomedicine, this review will provide a comprehensive overview of cell mechanotransduction mechanisms, including the structural components essential for effective mechanical responses, such as cytoskeletal elements, force-sensitive ion channels, membrane receptors and key signaling pathways. It will also discuss the clutch model in force transmission, the role of mechanotransduction in both physiology and pathological contexts, and biomechanics and biomaterial design. Additionally, we outline analytical approaches for characterizing forces at cellular and subcellular levels, discussing the advantages and limitations of each method to aid researchers in selecting appropriate techniques. Finally, we summarize recent advancements in cell force sensing and identify key challenges for future research. Overall, this review should contribute to biomedical engineering by supporting the design of biomaterials that integrate precise mechanical information.
Collapse
Affiliation(s)
- Xiaojun Liu
- College of Life Sciences and Health, University of Health and Rehabilitation Sciences, Qingdao 266113, China
- Qingdao Municipal Hospital, University of Health and Rehabilitation Sciences, Qingdao 266024, China
| | - Lei Yu
- Department of Traditional Chinese Medicine, Qingdao Special Service Sanatorium of PLA Navy, Qingdao 266071, China
| | - Adam Xiao
- Department of Chemistry, University of British Columbia, Vancouver, BC V6T 1Z1, Canada
| | - Wenxu Sun
- School of Sciences, Nantong University, Nantong 226019, China
| | - Han Wang
- State Key Laboratory of Precision Measuring Technology and Instruments, School of Precision Instrument and Optoelectronics Engineering, Tianjin University, Tianjin 300072, China
| | - Xiangxiu Wang
- Key Laboratory for Biorheological Science and Technology of Ministry of Education, State and Local Joint Engineering Laboratory for Vascular Implants, Bioengineering College of Chongqing University, Chongqing 400030, China
| | - Yanghao Zhou
- Key Laboratory for Biorheological Science and Technology of Ministry of Education, State and Local Joint Engineering Laboratory for Vascular Implants, Bioengineering College of Chongqing University, Chongqing 400030, China
| | - Chao Li
- College of Life Sciences and Health, University of Health and Rehabilitation Sciences, Qingdao 266113, China
- Qingdao Municipal Hospital, University of Health and Rehabilitation Sciences, Qingdao 266024, China
| | - Jiangtao Li
- College of Life Sciences and Health, University of Health and Rehabilitation Sciences, Qingdao 266113, China
| | - Yongliang Wang
- College of Life Sciences and Health, University of Health and Rehabilitation Sciences, Qingdao 266113, China
- Qingdao Municipal Hospital, University of Health and Rehabilitation Sciences, Qingdao 266024, China
- Key Laboratory for Biorheological Science and Technology of Ministry of Education, State and Local Joint Engineering Laboratory for Vascular Implants, Bioengineering College of Chongqing University, Chongqing 400030, China
- Qindao Central Hospital, University of Health and Rehabilitation Sciences, Qingdao 266044, China
| | - Guixue Wang
- Key Laboratory for Biorheological Science and Technology of Ministry of Education, State and Local Joint Engineering Laboratory for Vascular Implants, Bioengineering College of Chongqing University, Chongqing 400030, China
- JinFeng Laboratory, Chongqing 401329, China
| |
Collapse
|
|
3
|
Ren Y, Yang J, Saito T, Glomb O, Mousavi SI, Naughton B, de Fontnouvelle C, Fujita B, Schlieker C, Yogev S, Zhang Y, Berro J. Genetically encoded mechano-sensors with versatile readouts and compact size. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2025:2025.01.16.633409. [PMID: 39868242 PMCID: PMC11760715 DOI: 10.1101/2025.01.16.633409] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 01/28/2025]
Abstract
Mechanical forces are critical for virtually all fundamental biological processes, yet quantification of mechanical forces at the molecular scale in vivo remains challenging. Here, we present a new strategy using calibrated coiled-coils as genetically encoded, compact, tunable, and modular mechano-sensors to substantially simplify force measurement in vivo, via diverse readouts (luminescence, fluorescence and analytical biochemistry) and instrumentation readily available in biology labs. We demonstrate the broad applicability and ease-of-use of these coiled-coil mechano-sensors by measuring forces during cytokinesis (formin Cdc12) and endocytosis (epsin Ent1) in yeast, force distributions in nematode axons (β-spectrin UNC-70), and forces transmitted to the nucleus (mini-nesprin-2G) and within focal adhesions (vinculin) in mammalian cells. We report discoveries in intracellular force transmission that have been elusive to existing tools.
Collapse
Affiliation(s)
- Yuan Ren
- Department of Molecular Biophysics and Biochemistry; Yale University, New Haven, CT, USA
- Nanobiology Institute, Yale University; West Haven, CT 06516, USA
| | - Jie Yang
- Department of Cell Biology, Yale University School of Medicine; New Haven, CT 06520, USA
| | - Takumi Saito
- Department of Molecular Biophysics and Biochemistry; Yale University, New Haven, CT, USA
- Nanobiology Institute, Yale University; West Haven, CT 06516, USA
- Graduate School of Biomedical Engineering, Tohoku University, Sendai 980-8579, Japan
| | - Oliver Glomb
- Institut für Klinische Anatomie und Zellanalytik, Medizinische Fakultät, Eberhard Karls Universität Tübingen; Österbergstraße 3, 72074 Tübingen, Germany
| | - Sayed Iman Mousavi
- Department of Molecular Biophysics and Biochemistry; Yale University, New Haven, CT, USA
- Nanobiology Institute, Yale University; West Haven, CT 06516, USA
| | - Brigitte Naughton
- Department of Molecular Biophysics and Biochemistry; Yale University, New Haven, CT, USA
- Department of Cell Biology, Yale University School of Medicine; New Haven, CT 06520, USA
| | - Christina de Fontnouvelle
- Department of Molecular Biophysics and Biochemistry; Yale University, New Haven, CT, USA
- Department of Cell Biology, Yale University School of Medicine; New Haven, CT 06520, USA
| | - Barbara Fujita
- Department of Molecular Biophysics and Biochemistry; Yale University, New Haven, CT, USA
- Nanobiology Institute, Yale University; West Haven, CT 06516, USA
| | - Christian Schlieker
- Department of Molecular Biophysics and Biochemistry; Yale University, New Haven, CT, USA
- Department of Cell Biology, Yale University School of Medicine; New Haven, CT 06520, USA
| | - Shaul Yogev
- Department of Neuroscience, Yale University School of Medicine; New Haven, CT 06520, USA
| | - Yongli Zhang
- Department of Molecular Biophysics and Biochemistry; Yale University, New Haven, CT, USA
- Department of Cell Biology, Yale University School of Medicine; New Haven, CT 06520, USA
| | - Julien Berro
- Department of Molecular Biophysics and Biochemistry; Yale University, New Haven, CT, USA
- Nanobiology Institute, Yale University; West Haven, CT 06516, USA
- Department of Cell Biology, Yale University School of Medicine; New Haven, CT 06520, USA
| |
Collapse
|
|
4
|
Mittal S, Wang RE, Ros R, Ondrus AE, Singharoy A. Molecular dynamics model of mechanophore sensors for biological force measurement. Heliyon 2025; 11:e41178. [PMID: 39807516 PMCID: PMC11728885 DOI: 10.1016/j.heliyon.2024.e41178] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2024] [Revised: 11/12/2024] [Accepted: 12/11/2024] [Indexed: 01/16/2025] Open
Abstract
Cellular forces regulate an untold spectrum of living processes, such as cell migration, gene expression, and ion conduction. However, a quantitative description of mechanical control remains elusive due to the lack of general, live-cell tools to measure discrete forces between biomolecules. Here we introduce a computational pipeline for force measurement that leverages well-defined, tunable release of a mechanically activated small molecule fluorophore. These sensors are characterized using a multiscale approach combining equilibrium and steered QM/MM molecular dynamics models to capture the chemical, mechanical, and conformational transitions underlying force activation thresholds on a nano Newton scale. We find that chemical modification of the mechanophore and variation of its biomolecular tethers can tune the rate-determining step for fluorophore release and adjust the mechanochemical activation barrier. The models offer a new molecular framework for calibrated, programmable biomolecular force reporting within the live-cell regime, opening new opportunities to study mechanical phenomena in biological systems.
Collapse
Affiliation(s)
- Sumit Mittal
- School of Advanced Sciences and Languages, VIT Bhopal University, Kothrikalan, Sehore, Madhya Pradesh, 466114, India
| | - Rongsheng E. Wang
- Department of Chemistry, Temple University, 1901 N. 13th Street, Philadelphia, PA, 19122, USA
| | - Robert Ros
- Department of Physics, Arizona State University, Tempe, AZ, 85287, USA
- Center for Single Molecule Biophysics, The Biodesign Institute, Arizona State University, Tempe, AZ, 85287, USA
- Center for Biological Physics, Arizona State University, Tempe, AZ, 85287, USA
| | - Alison E. Ondrus
- Department of Chemistry and Pharmaceutical Sciences, University of Illinois Chicago, 900 W Taylor St, Science & Engineering Laboratories West South Building #608 Room 2230, Chicago, IL, 60607, USA
| | - Abhishek Singharoy
- School of Molecular Sciences, Arizona State University, Tempe, AZ, 85287, USA
| |
Collapse
|
|
5
|
Bighi B, Ragazzini G, Gallerani A, Mescola A, Scagliarini C, Zannini C, Marcuzzi M, Olivi E, Cavallini C, Tassinari R, Bianchi M, Corsi L, Ventura C, Alessandrini A. Cell stretching devices integrated with live cell imaging: a powerful approach to study how cells react to mechanical cues. PROGRESS IN BIOMEDICAL ENGINEERING (BRISTOL, ENGLAND) 2024; 7:012005. [PMID: 39655854 DOI: 10.1088/2516-1091/ad9699] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/29/2024] [Accepted: 11/20/2024] [Indexed: 12/18/2024]
Abstract
Mechanical stimuli have multiple effects on cell behavior, affecting a number of cellular processes including orientation, proliferation or apoptosis, migration and invasion, the production of extracellular matrix proteins, the activation and translocation of transcription factors, the expression of different genes such as those involved in inflammation and the reprogramming of cell fate. The recent development of cell stretching devices has paved the way for the study of cell reactions to stretching stimuliin-vitro, reproducing physiological situations that are experienced by cells in many tissues and related to functions such as breathing, heart beating and digestion. In this work, we review the highly-relevant contributions cell stretching devices can provide in the field of mechanobiology. We then provide the details for the in-house construction and operation of these devices, starting from the systems that we already developed and tested. We also review some examples where cell stretchers can supply meaningful insights into mechanobiology topics and we introduce new results from our exploitation of these devices.
Collapse
Affiliation(s)
- Beatrice Bighi
- Department of Physics, Informatics and Mathematics, University of Modena and Reggio Emilia, via Campi 213/A, 41125 Modena, Italy
- CNR-Nanoscience Institute-S3, via Campi 213/A, 41125 Modena, Italy
| | | | - Alessia Gallerani
- Department of Physics, Informatics and Mathematics, University of Modena and Reggio Emilia, via Campi 213/A, 41125 Modena, Italy
| | - Andrea Mescola
- CNR-Nanoscience Institute-S3, via Campi 213/A, 41125 Modena, Italy
| | - Chiara Scagliarini
- Department of Physics, Informatics and Mathematics, University of Modena and Reggio Emilia, via Campi 213/A, 41125 Modena, Italy
| | - Chiara Zannini
- Eldor Lab, via di Corticella 183, 40128 Bologna, Italy
- National Laboratory of Molecular Biology and Stem Cell Engineering, National Institute of Biostructures and Biosystems (I.N.B.B.), via di Corticella 183, 40128 Bologna, Italy
| | - Martina Marcuzzi
- Department of Medical and Surgical Sciences, University of Bologna, via G. Massarenti 9, Bologna 40138, Italy
| | - Elena Olivi
- Eldor Lab, via di Corticella 183, 40128 Bologna, Italy
| | - Claudia Cavallini
- Eldor Lab, via di Corticella 183, 40128 Bologna, Italy
- National Laboratory of Molecular Biology and Stem Cell Engineering, National Institute of Biostructures and Biosystems (I.N.B.B.), via di Corticella 183, 40128 Bologna, Italy
| | | | - Michele Bianchi
- Department of Life Sciences, University of Modena and Reggio Emilia, Via G. Campi 287, 41125 Modena, Italy
| | - Lorenzo Corsi
- Department of Life Sciences, University of Modena and Reggio Emilia, Via G. Campi 287, 41125 Modena, Italy
| | - Carlo Ventura
- Eldor Lab, via di Corticella 183, 40128 Bologna, Italy
- National Laboratory of Molecular Biology and Stem Cell Engineering, National Institute of Biostructures and Biosystems (I.N.B.B.), via di Corticella 183, 40128 Bologna, Italy
| | - Andrea Alessandrini
- Department of Physics, Informatics and Mathematics, University of Modena and Reggio Emilia, via Campi 213/A, 41125 Modena, Italy
- CNR-Nanoscience Institute-S3, via Campi 213/A, 41125 Modena, Italy
| |
Collapse
|
|
6
|
Fenelon KD, Krause J, Koromila T. Opticool: Cutting-edge transgenic optical tools. PLoS Genet 2024; 20:e1011208. [PMID: 38517915 PMCID: PMC10959397 DOI: 10.1371/journal.pgen.1011208] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/24/2024] Open
Abstract
Only a few short decades have passed since the sequencing of GFP, yet the modern repertoire of transgenically encoded optical tools implies an exponential proliferation of ever improving constructions to interrogate the subcellular environment. A myriad of tags for labeling proteins, RNA, or DNA have arisen in the last few decades, facilitating unprecedented visualization of subcellular components and processes. Development of a broad array of modern genetically encoded sensors allows real-time, in vivo detection of molecule levels, pH, forces, enzyme activity, and other subcellular and extracellular phenomena in ever expanding contexts. Optogenetic, genetically encoded optically controlled manipulation systems have gained traction in the biological research community and facilitate single-cell, real-time modulation of protein function in vivo in ever broadening, novel applications. While this field continues to explosively expand, references are needed to assist scientists seeking to use and improve these transgenic devices in new and exciting ways to interrogate development and disease. In this review, we endeavor to highlight the state and trajectory of the field of in vivo transgenic optical tools.
Collapse
Affiliation(s)
- Kelli D. Fenelon
- Department of Biology, University of Texas at Arlington, Arlington, Texas, United States of America
| | - Julia Krause
- Department of Biology, University of Texas at Arlington, Arlington, Texas, United States of America
| | - Theodora Koromila
- Department of Biology, University of Texas at Arlington, Arlington, Texas, United States of America
- School of Biology, Aristotle University of Thessaloniki, Thessaloniki, Greece
| |
Collapse
|
|
7
|
Halma MTJ, Xu L. Life under tension: the relevance of force on biological polymers. BIOPHYSICS REPORTS 2024; 10:48-56. [PMID: 38737478 PMCID: PMC11079598 DOI: 10.52601/bpr.2023.230019] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2023] [Accepted: 12/06/2023] [Indexed: 05/14/2024] Open
Abstract
Optical tweezers have elucidated numerous biological processes, particularly by enabling the precise manipulation and measurement of tension. One question concerns the biological relevance of these experiments and the generalizability of these experiments to wider biological systems. Here, we categorize the applicability of the information garnered from optical tweezers in two distinct categories: the direct relevance of tension in biological systems, and what experiments under tension can tell us about biological systems, while these systems do not reach the same tension as the experiment, still, these artificial experimental systems reveal insights into the operations of biological machines and life processes.
Collapse
Affiliation(s)
- Matthew T. J. Halma
- Department of Physics and Astronomy and LaserLab, Vrije Universiteit Amsterdam, De Boelelaan 1081, 1081 HV, Amsterdam, the Netherlands
- LUMICKS B. V., 1081 HV, Amsterdam, the Netherlands
| | - Longfu Xu
- Department of Physics and Astronomy and LaserLab, Vrije Universiteit Amsterdam, De Boelelaan 1081, 1081 HV, Amsterdam, the Netherlands
| |
Collapse
|
|
8
|
Singh Y, Hocky GM. Improved Prediction of Molecular Response to Pulling by Combining Force Tempering with Replica Exchange Methods. J Phys Chem B 2024; 128:706-715. [PMID: 38230998 PMCID: PMC10823473 DOI: 10.1021/acs.jpcb.3c07081] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2023] [Revised: 01/02/2024] [Accepted: 01/03/2024] [Indexed: 01/18/2024]
Abstract
Small mechanical forces play important functional roles in many crucial cellular processes, including in the dynamic behavior of the cytoskeleton and in the regulation of osmotic pressure through membrane-bound proteins. Molecular simulations offer the promise of being able to design the behavior of proteins that sense and respond to these forces. However, it is difficult to predict and identify the effect of the relevant piconewton (pN) scale forces due to their small magnitude. Previously, we introduced the Infinite Switch Simulated Tempering in Force (FISST) method, which allows one to estimate the effect of a range of applied forces from a single molecular dynamics simulation, and also demonstrated that FISST additionally accelerates sampling of a molecule's conformational landscape. For some problems, we find that this acceleration is not sufficient to capture all relevant conformational fluctuations, and hence, here we demonstrate that FISST can be combined with either temperature replica exchange or solute tempering approaches to produce a hybrid method that enables more robust prediction of the effect of small forces on molecular systems.
Collapse
Affiliation(s)
- Yuvraj Singh
- Department
of Chemistry, New York University, New York, New York 10003, United States
| | - Glen M. Hocky
- Department
of Chemistry, New York University, New York, New York 10003, United States
- Simons
Center for Computational Physical Chemistry, New York University, New York, New York 10003, United States
| |
Collapse
|
|
9
|
Wang R, Bialas AL, Goel T, Collins EMS. Mechano-Chemical Coupling in Hydra Regeneration and Patterning. Integr Comp Biol 2023; 63:1422-1441. [PMID: 37339912 DOI: 10.1093/icb/icad070] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2023] [Revised: 06/07/2023] [Accepted: 06/12/2023] [Indexed: 06/22/2023] Open
Abstract
The freshwater cnidarian Hydra can regenerate from wounds, small tissue fragments and even from aggregated cells. This process requires the de novo development of a body axis and oral-aboral polarity, a fundamental developmental process that involves chemical patterning and mechanical shape changes. Gierer and Meinhardt recognized that Hydra's simple body plan and amenability to in vivo experiments make it an experimentally and mathematically tractable model to study developmental patterning and symmetry breaking. They developed a reaction-diffusion model, involving a short-range activator and a long-range inhibitor, which successfully explained patterning in the adult animal. In 2011, HyWnt3 was identified as a candidate for the activator. However, despite the continued efforts of both physicists and biologists, the predicted inhibitor remains elusive. Furthermore, the Gierer-Meinhardt model cannot explain de novo axis formation in cellular aggregates that lack inherited tissue polarity. The aim of this review is to synthesize the current knowledge on Hydra symmetry breaking and patterning. We summarize the history of patterning studies and insights from recent biomechanical and molecular studies, and highlight the need for continued validation of theoretical assumptions and collaboration across disciplinary boundaries. We conclude by proposing new experiments to test current mechano-chemical coupling models and suggest ideas for expanding the Gierer-Meinhardt model to explain de novo patterning, as observed in Hydra aggregates. The availability of a fully sequenced genome, transgenic fluorescent reporter strains, and modern imaging techniques, that enable unprecedented observation of cellular events in vivo, promise to allow the community to crack Hydra's secret to patterning.
Collapse
Affiliation(s)
- Rui Wang
- Department of Bioengineering, University of California San Diego, 9500 Gilman Drive, La Jolla, 92093 CA, USA
| | - April L Bialas
- Department of Biology, Swarthmore College, 500 College Ave, Swarthmore, 19081 PA, USA
| | - Tapan Goel
- School of Biological Sciences, Georgia Institute of Technology, Atlanta, 30332 GA, USA
- Department of Physics, University of California San Diego, 9500 Gilman Drive, La Jolla, 92093 CA, USA
| | - Eva-Maria S Collins
- Department of Biology, Swarthmore College, 500 College Ave, Swarthmore, 19081 PA, USA
- Department of Physics, University of California San Diego, 9500 Gilman Drive, La Jolla, 92093 CA, USA
- Department of Neuroscience, Perelman School of Medicine, University of Pennsylvania, Philadelphia, 19104 PA, USA
| |
Collapse
|
|
10
|
Amiri S, Muresan C, Shang X, Huet-Calderwood C, Schwartz MA, Calderwood DA, Murrell M. Intracellular tension sensor reveals mechanical anisotropy of the actin cytoskeleton. Nat Commun 2023; 14:8011. [PMID: 38049429 PMCID: PMC10695988 DOI: 10.1038/s41467-023-43612-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/12/2023] [Accepted: 11/15/2023] [Indexed: 12/06/2023] Open
Abstract
The filamentous actin (F-actin) cytoskeleton is a composite material consisting of cortical actin and bundled F-actin stress fibers, which together mediate the mechanical behaviors of the cell, from cell division to cell migration. However, as mechanical forces are typically measured upon transmission to the extracellular matrix, the internal distribution of forces within the cytoskeleton is unknown. Likewise, how distinct F-actin architectures contribute to the generation and transmission of mechanical forces is unclear. Therefore, we have developed a molecular tension sensor that embeds into the F-actin cytoskeleton. Using this sensor, we measure tension within stress fibers and cortical actin, as the cell is subject to uniaxial stretch. We find that the mechanical response, as measured by FRET, depends on the direction of applied stretch relative to the cell's axis of alignment. When the cell is aligned parallel to the direction of the stretch, stress fibers and cortical actin both accumulate tension. By contrast, when aligned perpendicular to the direction of stretch, stress fibers relax tension while the cortex accumulates tension, indicating mechanical anisotropy within the cytoskeleton. We further show that myosin inhibition regulates this anisotropy. Thus, the mechanical anisotropy of the cell and the coordination between distinct F-actin architectures vary and depend upon applied load.
Collapse
Affiliation(s)
- Sorosh Amiri
- Systems Biology Institute, 850 West Campus Drive, Yale University, West Haven, CT, 06516, USA
- Department of Mechanical Engineering and Material Science, 17 Hillhouse Ave, Yale University, New Haven, CT, 06511, USA
| | - Camelia Muresan
- Systems Biology Institute, 850 West Campus Drive, Yale University, West Haven, CT, 06516, USA
- Department of Biomedical Engineering, 17 Hillhouse Ave, Yale University, New Haven, CT, 06511, USA
| | - Xingbo Shang
- Systems Biology Institute, 850 West Campus Drive, Yale University, West Haven, CT, 06516, USA
- Department of Biomedical Engineering, 17 Hillhouse Ave, Yale University, New Haven, CT, 06511, USA
| | | | - Martin A Schwartz
- Department of Biomedical Engineering, 17 Hillhouse Ave, Yale University, New Haven, CT, 06511, USA
- Department of Cell Biology, 333 Cedar St, Yale University, New Haven, CT, 06510, USA
- Yale Cardiovascular Research Center, 300 George St, New Haven, CT, 06511, USA
| | - David A Calderwood
- Department of Pharmacology, 333 Cedar St, Yale University, New Haven, CT, 06510, USA
- Department of Cell Biology, 333 Cedar St, Yale University, New Haven, CT, 06510, USA
| | - Michael Murrell
- Systems Biology Institute, 850 West Campus Drive, Yale University, West Haven, CT, 06516, USA.
- Department of Biomedical Engineering, 17 Hillhouse Ave, Yale University, New Haven, CT, 06511, USA.
- Department of Physics, 217 Prospect Street, Yale University, New Haven, CT, 06511, USA.
| |
Collapse
|
|
11
|
Cheikh MI, Tchoufag J, Osterfield M, Dean K, Bhaduri S, Zhang C, Mandadapu KK, Doubrovinski K. A comprehensive model of Drosophila epithelium reveals the role of embryo geometry and cell topology in mechanical responses. eLife 2023; 12:e85569. [PMID: 37782009 PMCID: PMC10584372 DOI: 10.7554/elife.85569] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/14/2022] [Accepted: 09/29/2023] [Indexed: 10/03/2023] Open
Abstract
In order to understand morphogenesis, it is necessary to know the material properties or forces shaping the living tissue. In spite of this need, very few in vivo measurements are currently available. Here, using the early Drosophila embryo as a model, we describe a novel cantilever-based technique which allows for the simultaneous quantification of applied force and tissue displacement in a living embryo. By analyzing data from a series of experiments in which embryonic epithelium is subjected to developmentally relevant perturbations, we conclude that the response to applied force is adiabatic and is dominated by elastic forces and geometric constraints, or system size effects. Crucially, computational modeling of the experimental data indicated that the apical surface of the epithelium must be softer than the basal surface, a result which we confirmed experimentally. Further, we used the combination of experimental data and comprehensive computational model to estimate the elastic modulus of the apical surface and set a lower bound on the elastic modulus of the basal surface. More generally, our investigations revealed important general features that we believe should be more widely addressed when quantitatively modeling tissue mechanics in any system. Specifically, different compartments of the same cell can have very different mechanical properties; when they do, they can contribute differently to different mechanical stimuli and cannot be merely averaged together. Additionally, tissue geometry can play a substantial role in mechanical response, and cannot be neglected.
Collapse
Affiliation(s)
- Mohamad Ibrahim Cheikh
- Department of Biophysics, University of Texas Southwestern Medical CenterDallasUnited States
| | - Joel Tchoufag
- Department of Chemical and Biomolecular Engineering, University of California, BerkeleyBerkeleyUnited States
- Chemical Sciences Division, Lawrence Berkeley National LaboratoryBerkeleyUnited States
| | - Miriam Osterfield
- Department of Biophysics, University of Texas Southwestern Medical CenterDallasUnited States
| | - Kevin Dean
- Department of Bioinformatics, University of Texas Southwestern Medical CenterDallasUnited States
| | - Swayamdipta Bhaduri
- Department of Biophysics, University of Texas Southwestern Medical CenterDallasUnited States
| | - Chuzhong Zhang
- Department of Material Science and Engineering, University of Texas at ArlingtonArlingtonUnited States
| | - Kranthi Kiran Mandadapu
- Department of Chemical and Biomolecular Engineering, University of California, BerkeleyBerkeleyUnited States
- Chemical Sciences Division, Lawrence Berkeley National LaboratoryBerkeleyUnited States
| | - Konstantin Doubrovinski
- Department of Biophysics, University of Texas Southwestern Medical CenterDallasUnited States
| |
Collapse
|
|
12
|
Yousafzai MS, Hammer JA. Using Biosensors to Study Organoids, Spheroids and Organs-on-a-Chip: A Mechanobiology Perspective. BIOSENSORS 2023; 13:905. [PMID: 37887098 PMCID: PMC10605946 DOI: 10.3390/bios13100905] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/26/2023] [Revised: 09/13/2023] [Accepted: 09/19/2023] [Indexed: 10/28/2023]
Abstract
The increasing popularity of 3D cell culture models is being driven by the demand for more in vivo-like conditions with which to study the biochemistry and biomechanics of numerous biological processes in health and disease. Spheroids and organoids are 3D culture platforms that self-assemble and regenerate from stem cells, tissue progenitor cells or cell lines, and that show great potential for studying tissue development and regeneration. Organ-on-a-chip approaches can be used to achieve spatiotemporal control over the biochemical and biomechanical signals that promote tissue growth and differentiation. These 3D model systems can be engineered to serve as disease models and used for drug screens. While culture methods have been developed to support these 3D structures, challenges remain to completely recapitulate the cell-cell and cell-matrix biomechanical interactions occurring in vivo. Understanding how forces influence the functions of cells in these 3D systems will require precise tools to measure such forces, as well as a better understanding of the mechanobiology of cell-cell and cell-matrix interactions. Biosensors will prove powerful for measuring forces in both of these contexts, thereby leading to a better understanding of how mechanical forces influence biological systems at the cellular and tissue levels. Here, we discussed how biosensors and mechanobiological research can be coupled to develop accurate, physiologically relevant 3D tissue models to study tissue development, function, malfunction in disease, and avenues for disease intervention.
Collapse
Affiliation(s)
- Muhammad Sulaiman Yousafzai
- Cell and Developmental Biology Center, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA
| | - John A. Hammer
- Cell and Developmental Biology Center, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA
| |
Collapse
|
|
13
|
Metze F, Sant S, Meng Z, Klok HA, Kaur K. Swelling-Activated, Soft Mechanochemistry in Polymer Materials. LANGMUIR : THE ACS JOURNAL OF SURFACES AND COLLOIDS 2023; 39:3546-3557. [PMID: 36848262 PMCID: PMC10018775 DOI: 10.1021/acs.langmuir.2c02801] [Citation(s) in RCA: 27] [Impact Index Per Article: 13.5] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/13/2022] [Revised: 01/26/2023] [Indexed: 06/12/2023]
Abstract
Swelling in polymer materials is a ubiquitous phenomenon. At a molecular level, swelling is dictated by solvent-polymer interactions, and has been thoroughly studied both theoretically and experimentally. Favorable solvent-polymer interactions result in the solvation of polymer chains. For polymers in confined geometries, such as those that are tethered to surfaces, or for polymer networks, solvation can lead to swelling-induced tensions. These tensions act on polymer chains and can lead to stretching, bending, or deformation of the material both at the micro- and macroscopic scale. This Invited Feature Article sheds light on such swelling-induced mechanochemical phenomena in polymer materials across dimensions, and discusses approaches to visualize and characterize these effects.
Collapse
|
|
14
|
Abstract
Plasma membrane tension functions as a global physical organizer of cellular activities. Technical limitations of current membrane tension measurement techniques have hampered in-depth investigation of cellular membrane biophysics and the role of plasma membrane tension in regulating cellular processes. Here, we develop an optical membrane tension reporter by repurposing an E. coli mechanosensitive channel via insertion of circularly permuted GFP (cpGFP), which undergoes a large conformational rearrangement associated with channel activation and thus fluorescence intensity changes under increased membrane tension.
Collapse
Affiliation(s)
- Yen-Yu Hsu
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States
| | - Agnes M Resto Irizarry
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States
| | - Jianping Fu
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States
- Department of Cell & Developmental Biology, University of Michigan, Ann Arbor, Michigan 48109, United States
| | - Allen P Liu
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, Michigan 48109, United States
- Department of Biophysics, University of Michigan, Ann Arbor, Michigan 48109, United States
- Cellular and Molecular Biology Program, University of Michigan, Ann Arbor, Michigan 48109, United States
| |
Collapse
|
|
15
|
Windgasse L, Grashoff C. Multiplexed Molecular Tension Sensor Measurements Using PIE-FLIM. Methods Mol Biol 2023; 2600:221-237. [PMID: 36587101 DOI: 10.1007/978-1-0716-2851-5_15] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023]
Abstract
Genetically encoded Förster Resonance Energy Transfer (FRET)-based tension sensors were developed to enable the quantification of piconewton (pN)-scale forces that act across distinct proteins in living cells and organisms. An important extension of this technology is the multiplexing of tension sensors to monitor several independent FRET probes in parallel. Here we describe how pulsed interleaved excitation (PIE)-fluorescence lifetime imaging microscopy (FLIM) can be implemented to enable the analysis of two co-expressed tension sensor constructs. Our protocol covers all essential steps from biosensor expression and live cell PIE image acquisition to lifetime calculations.
Collapse
Affiliation(s)
- Lukas Windgasse
- Department of Quantitative Cell Biology, Institute of Integrative Cell Biology and Physiology, University of Münster, Münster, Germany
| | - Carsten Grashoff
- Department of Quantitative Cell Biology, Institute of Integrative Cell Biology and Physiology, University of Münster, Münster, Germany.
| |
Collapse
|
|
16
|
Sanfeliu-Cerdán N, Lin LC, Dunn AR, Goodman MB, Krieg M. Visualizing Neurons Under Tension In Vivo with Optogenetic Molecular Force Sensors. Methods Mol Biol 2023; 2600:239-266. [PMID: 36587102 PMCID: PMC11874908 DOI: 10.1007/978-1-0716-2851-5_16] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023]
Abstract
The visualization of mechanical stress distribution in specific molecular networks within a living and physiologically active cell or animal remains a formidable challenge in mechanobiology. The advent of fluorescence-resonance energy transfer (FRET)-based molecular tension sensors overcame a significant hurdle that now enables us to address previously technically limited questions. Here, we describe a method that uses genetically encoded FRET tension sensors to visualize the mechanics of cytoskeletal networks in neurons of living animals with sensitized emission FRET and confocal scanning light microscopy. This method uses noninvasive immobilization of living animals to image neuronal β-spectrin cytoskeleton at the diffraction limit, and leverages multiple imaging controls to verify and underline the quality of the measurements. In combination with a semiautomated machine-vision algorithm to identify and trace individual neurites, our analysis performs simultaneous calculation of FRET efficiencies and visualizes statistical uncertainty on a pixel by pixel basis. Our approach is not limited to genetically encoded spectrin tension sensors, but can also be used for any kind of ratiometric imaging in neuronal cells both in vivo and in vitro.
Collapse
Affiliation(s)
- Neus Sanfeliu-Cerdán
- Neurophotonics and Mechanical Systems Biology, ICFO, Institut de Ciències Fotòniques, ICFO, Castelldefels, Spain
| | - Li-Chun Lin
- Neurophotonics and Mechanical Systems Biology, ICFO, Institut de Ciències Fotòniques, ICFO, Castelldefels, Spain
| | - Alexander R Dunn
- Department of Chemical Engineering, Stanford University, Stanford, CA, USA
| | - Miriam B Goodman
- Molecular and Cellular Physiology, Stanford University, Stanford, CA, USA
| | - Michael Krieg
- Neurophotonics and Mechanical Systems Biology, ICFO, Institut de Ciències Fotòniques, ICFO, Castelldefels, Spain.
| |
Collapse
|
|
17
|
Beedle AEM, Garcia-Manyes S. The role of single protein elasticity in mechanobiology. NATURE REVIEWS. MATERIALS 2023; 8:10-24. [PMID: 37469679 PMCID: PMC7614781 DOI: 10.1038/s41578-022-00488-z] [Citation(s) in RCA: 10] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Accepted: 09/07/2022] [Indexed: 07/21/2023]
Abstract
In addition to biochemical signals and genetic considerations, mechanical forces are rapidly emerging as a master regulator of human physiology. Yet the molecular mechanisms that regulate force-induced functionalities across a wide range of scales, encompassing the cell, tissue or organ levels, are comparatively not so well understood. With the advent, development and refining of single molecule nanomechanical techniques, enabling to exquisitely probe the conformational dynamics of individual proteins under the effect of a calibrated force, we have begun to acquire a comprehensive knowledge on the rich plethora of physicochemical principles that regulate the elasticity of single proteins. Here we review the major advances underpinning our current understanding of how the elasticity of single proteins regulates mechanosensing and mechanotransduction. We discuss the present limitations and future challenges of such a prolific and burgeoning field.
Collapse
Affiliation(s)
- Amy EM Beedle
- Department of Physics, Randall Centre for Cell and Molecular Biophysics, Centre for the Physical Science of Life and London Centre for Nanotechnology, King’s College London, Strand, WC2R 2LS London, United Kingdom
- Institute for Bioengineering of Catalonia (IBEC), the Barcelona Institute of Technology (BIST), 08028 Barcelona, Spain
| | - Sergi Garcia-Manyes
- Department of Physics, Randall Centre for Cell and Molecular Biophysics, Centre for the Physical Science of Life and London Centre for Nanotechnology, King’s College London, Strand, WC2R 2LS London, United Kingdom
- Single Molecule Mechanobiology Laboratory, The Francis Crick Institute, 1 Midland Road, London NW1 1AT, London, UK
| |
Collapse
|
|
18
|
Cheng N, Zhang Y, Wu Y, Li B, Wang H, Chen S, Zhao P, Cui J, Shen X, Zhu X, Zheng Y. Hydrogel platform capable of molecularly resolved pulling on cells for mechanotransduction. Mater Today Bio 2022; 17:100476. [DOI: 10.1016/j.mtbio.2022.100476] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2022] [Revised: 10/26/2022] [Accepted: 10/26/2022] [Indexed: 11/06/2022]
|
|
19
|
Magnetomechanical Stress-Induced Colon Cancer Cell Growth Inhibition. JOURNAL OF NANOTHERANOSTICS 2022. [DOI: 10.3390/jnt3030010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Abstract
The application of magnetomechanical stress in cells using internalized magnetic nanoparticles (MNPs) actuated by low-frequency magnetic fields has been attracting considerable interest in the field of cancer research. Recent developments prove that magnetomechanical stress can inhibit cancer cells’ growth. However, the MNPs’ type and the magnetic field’s characteristics are crucial parameters. Their variability allows multiple combinations, which induce specific biological effects. We previously reported the antiproliferative effects induced in HT29 colon cancer cells by static-magnetic-field (200 mT)-actuated spherical MNPs (100 nm). Herein, we show that similar growth inhibitory effects are induced in other colon cancer cell lines. The effect of magnetomechanical stress was also examined in the growth rate of tumor spheroids. Moreover, we examined the biological mechanisms involved in the observed cell growth inhibition. Under the experimental conditions employed, no cell death was detected by PI (propidium iodide) staining analysis. Flow cytometry and Western blotting revealed that G2/M cell cycle arrest might mediate the antiproliferative effects. Furthermore, MNPs were found to locate in the lysosomes, and a decreased number of lysosomes was detected in cells that had undergone magnetomechanical stress, implying that the mechanical activation of the internalized MNPs could induce lysosome membrane disruption. Of note, the lysosomal acidic conditions were proven to affect the MNPs’ magnetic properties, evidenced by vibrating sample magnetometry (VSM) analysis. Further research on the combination of the described magnetomechanical stress with lysosome-targeting chemotherapeutic drugs could lay the groundwork for the development of novel anticancer combination treatment schemes.
Collapse
|
|
20
|
Vesga-Castro C, Aldazabal J, Vallejo-Illarramendi A, Paredes J. Contractile force assessment methods for in vitro skeletal muscle tissues. eLife 2022; 11:e77204. [PMID: 35604384 PMCID: PMC9126583 DOI: 10.7554/elife.77204] [Citation(s) in RCA: 14] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2022] [Accepted: 04/27/2022] [Indexed: 02/06/2023] Open
Abstract
Over the last few years, there has been growing interest in measuring the contractile force (CF) of engineered muscle tissues to evaluate their functionality. However, there are still no standards available for selecting the most suitable experimental platform, measuring system, culture protocol, or stimulation patterns. Consequently, the high variability of published data hinders any comparison between different studies. We have identified that cantilever deflection, post deflection, and force transducers are the most commonly used configurations for CF assessment in 2D and 3D models. Additionally, we have discussed the most relevant emerging technologies that would greatly complement CF evaluation with intracellular and localized analysis. This review provides a comprehensive analysis of the most significant advances in CF evaluation and its critical parameters. In order to compare contractile performance across experimental platforms, we have used the specific force (sF, kN/m2), CF normalized to the calculated cross-sectional area (CSA). However, this parameter presents a high variability throughout the different studies, which indicates the need to identify additional parameters and complementary analysis suitable for proper comparison. We propose that future contractility studies in skeletal muscle constructs report detailed information about construct size, contractile area, maturity level, sarcomere length, and, ideally, the tetanus-to-twitch ratio. These studies will hopefully shed light on the relative impact of these variables on muscle force performance of engineered muscle constructs. Prospective advances in muscle tissue engineering, particularly in muscle disease models, will require a joint effort to develop standardized methodologies for assessing CF of engineered muscle tissues.
Collapse
Affiliation(s)
- Camila Vesga-Castro
- University of Navarra, Tecnun School of Engineering, Manuel de LardizábalSan SebastianSpain
- University of Navarra, Biomedical Engineering Center, Campus UniversitarioPamplonaSpain
- Group of Neurosciences, Department of Pediatrics, UPV/EHU, Hospital Donostia - IIS BiodonostiaSan SebastianSpain
| | - Javier Aldazabal
- University of Navarra, Tecnun School of Engineering, Manuel de LardizábalSan SebastianSpain
- University of Navarra, Biomedical Engineering Center, Campus UniversitarioPamplonaSpain
| | - Ainara Vallejo-Illarramendi
- Group of Neurosciences, Department of Pediatrics, UPV/EHU, Hospital Donostia - IIS BiodonostiaSan SebastianSpain
- CIBERNED, Instituto de Salud Carlos III, Ministry of Science, Innovation, and UniversitiesMadridSpain
| | - Jacobo Paredes
- University of Navarra, Tecnun School of Engineering, Manuel de LardizábalSan SebastianSpain
- University of Navarra, Biomedical Engineering Center, Campus UniversitarioPamplonaSpain
| |
Collapse
|
|
21
|
Molecular sensors for detection of tumor-stroma crosstalk. Adv Cancer Res 2022; 154:47-91. [PMID: 35459472 DOI: 10.1016/bs.acr.2022.01.001] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
In most solid tumors, malignant cells coexist with non-cancerous host tissue comprised of a variety of extracellular matrix components and cell types, notably fibroblasts, immune cells, and endothelial cells. It is becoming increasingly evident that the non-cancerous host tissue, often referred to as the tumor stroma or the tumor microenvironment, wields tremendous influence in the proliferation, survival, and metastatic ability of cancer cells. The tumor stroma has an active biological role in the transmission of signals, such as growth factors and chemokines that activate oncogenic signaling pathways by autocrine and paracrine mechanisms. Moreover, the constituents of the stroma define the mechanical properties and the physical features of solid tumors, which influence cancer progression and response to therapy. Inspired by the emerging importance of tumor-stroma crosstalk and oncogenic physical forces, numerous biosensors, or advanced imaging and analysis techniques have been developed and applied to investigate complex and challenging questions in cancer research. These techniques facilitate measurements and biological readouts at scales ranging from subcellular to tissue-level with unprecedented level of spatial and temporal precision. Here we examine the application of biosensor technology for studying the complex and dynamic multiscale interactions of the tumor-host system.
Collapse
|
|
22
|
Sun W, Gao X, Lei H, Wang W, Cao Y. Biophysical Approaches for Applying and Measuring Biological Forces. ADVANCED SCIENCE (WEINHEIM, BADEN-WURTTEMBERG, GERMANY) 2022; 9:e2105254. [PMID: 34923777 PMCID: PMC8844594 DOI: 10.1002/advs.202105254] [Citation(s) in RCA: 13] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/15/2021] [Indexed: 05/13/2023]
Abstract
Over the past decades, increasing evidence has indicated that mechanical loads can regulate the morphogenesis, proliferation, migration, and apoptosis of living cells. Investigations of how cells sense mechanical stimuli or the mechanotransduction mechanism is an active field of biomaterials and biophysics. Gaining a further understanding of mechanical regulation and depicting the mechanotransduction network inside cells require advanced experimental techniques and new theories. In this review, the fundamental principles of various experimental approaches that have been developed to characterize various types and magnitudes of forces experienced at the cellular and subcellular levels are summarized. The broad applications of these techniques are introduced with an emphasis on the difficulties in implementing these techniques in special biological systems. The advantages and disadvantages of each technique are discussed, which can guide readers to choose the most suitable technique for their questions. A perspective on future directions in this field is also provided. It is anticipated that technical advancement can be a driving force for the development of mechanobiology.
Collapse
Affiliation(s)
- Wenxu Sun
- School of SciencesNantong UniversityNantong226019P. R. China
| | - Xiang Gao
- Key Laboratory of Intelligent Optical Sensing and IntegrationNational Laboratory of Solid State Microstructureand Department of PhysicsCollaborative Innovation Center of Advanced MicrostructuresNanjing UniversityNanjing210023P. R. China
- Institute of Brain ScienceNanjing UniversityNanjing210023P. R. China
| | - Hai Lei
- Key Laboratory of Intelligent Optical Sensing and IntegrationNational Laboratory of Solid State Microstructureand Department of PhysicsCollaborative Innovation Center of Advanced MicrostructuresNanjing UniversityNanjing210023P. R. China
- Institute of Brain ScienceNanjing UniversityNanjing210023P. R. China
- Chemistry and Biomedicine Innovation CenterNanjing UniversityNanjing210023P. R. China
| | - Wei Wang
- Key Laboratory of Intelligent Optical Sensing and IntegrationNational Laboratory of Solid State Microstructureand Department of PhysicsCollaborative Innovation Center of Advanced MicrostructuresNanjing UniversityNanjing210023P. R. China
- Institute of Brain ScienceNanjing UniversityNanjing210023P. R. China
| | - Yi Cao
- Key Laboratory of Intelligent Optical Sensing and IntegrationNational Laboratory of Solid State Microstructureand Department of PhysicsCollaborative Innovation Center of Advanced MicrostructuresNanjing UniversityNanjing210023P. R. China
- Institute of Brain ScienceNanjing UniversityNanjing210023P. R. China
- MOE Key Laboratory of High Performance Polymer Materials and TechnologyDepartment of Polymer Science & EngineeringCollege of Chemistry & Chemical EngineeringNanjing UniversityNanjing210023P. R. China
- Chemistry and Biomedicine Innovation CenterNanjing UniversityNanjing210023P. R. China
| |
Collapse
|
|
23
|
Pan J, Kmieciak T, Liu YT, Wildenradt M, Chen YS, Zhao Y. Quantifying molecular- to cellular-level forces in living cells. JOURNAL OF PHYSICS D: APPLIED PHYSICS 2021; 54:483001. [PMID: 34866655 PMCID: PMC8635116 DOI: 10.1088/1361-6463/ac2170] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/13/2023]
Abstract
Mechanical cues have been suggested to play an important role in cell functions and cell fate determination, however, such physical quantities are challenging to directly measure in living cells with single molecule sensitivity and resolution. In this review, we focus on two main technologies that are promising in probing forces at the single molecule level. We review their theoretical fundamentals, recent technical advancements, and future directions, tailored specifically for interrogating mechanosensitive molecules in live cells.
Collapse
Affiliation(s)
- Jason Pan
- Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801, United States of America
| | - Tommy Kmieciak
- Department of Engineering Physics, University of Illinois at Urbana-Champaign, Urbana, IL 61801, United States of America
| | - Yen-Ting Liu
- Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801, United States of America
| | - Matthew Wildenradt
- Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801, United States of America
| | - Yun-Sheng Chen
- Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801, United States of America
- Beckman Institute for Advanced Science and Technology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, United States of America
| | - Yang Zhao
- Department of Electrical and Computer Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801, United States of America
- Holonyak Micro and Nanotechnology Laboratory, University of Illinois at Urbana-Champaign, 208 N. Wright Street, Urbana, IL 61801, United States of America
| |
Collapse
|
|
24
|
Wang Q, Duan M, Liao J, Xie J, Zhou C. Are Osteoclasts Mechanosensitive Cells? J Biomed Nanotechnol 2021; 17:1917-1938. [PMID: 34706793 DOI: 10.1166/jbn.2021.3171] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
Skeleton metabolism is a process in which osteoclasts constantly remove old bone and osteoblasts form new osteoid and induce mineralization; disruption of this balance may cause diseases. Osteoclasts play a key role in bone metabolism, as osteoclastogenesis marks the beginning of each bone remodeling cycle. As the only cell capable of bone resorption, osteoclasts are derived from the monocyte/macrophage hematopoietic precursors that terminally adhere to mineralized extracellular matrix, and they subsequently break down the extracellular compartment. Bone is generally considered the load-burdening tissue, bone homeostasis is critically affected by mechanical conductions, and the bone cells are mechanosensitive. The functions of various bone cells under mechanical forces such as chondrocytes and osteoblasts have been reported; however, the unique bone-resorbing osteoclasts are less studied. The oversuppression of osteoclasts in mechanical studies may be because of its complicated differentiation progress and flexible structure, which increases difficulty in targeting mechanical structures. This paper will focus on recent findings regarding osteoclasts and attempt to uncover proposed candidate mechanosensing structures in osteoclasts including podosome-associated complexes, gap junctions and transient receptor potential family (ion channels). We will additionally describe possible mechanotransduction signaling pathways including GTPase ras homologue family member A (RhoA), Yes-associated protein/transcriptional co-activator with PDZ-binding motif (TAZ), Ca2+ signaling and non-canonical Wnt signaling. According to numerous studies, evaluating the possible influence of various physical environments on osteoclastogenesis is conducive to the study of bone homeostasis.
Collapse
Affiliation(s)
- Qingxuan Wang
- State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610064, China
| | - Mengmeng Duan
- State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610064, China
| | - Jingfeng Liao
- State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610064, China
| | - Jing Xie
- State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610064, China
| | - Chenchen Zhou
- State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, 610064, China
| |
Collapse
|
|
25
|
Hobson CM, Aaron JS, Heddleston JM, Chew TL. Visualizing the Invisible: Advanced Optical Microscopy as a Tool to Measure Biomechanical Forces. Front Cell Dev Biol 2021; 9:706126. [PMID: 34552926 PMCID: PMC8450411 DOI: 10.3389/fcell.2021.706126] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/06/2021] [Accepted: 08/09/2021] [Indexed: 01/28/2023] Open
Abstract
The importance of mechanical force in biology is evident across diverse length scales, ranging from tissue morphogenesis during embryo development to mechanotransduction across single adhesion proteins at the cell surface. Consequently, many force measurement techniques rely on optical microscopy to measure forces being applied by cells on their environment, to visualize specimen deformations due to external forces, or even to directly apply a physical perturbation to the sample via photoablation or optogenetic tools. Recent developments in advanced microscopy offer improved approaches to enhance spatiotemporal resolution, imaging depth, and sample viability. These advances can be coupled with already existing force measurement methods to improve sensitivity, duration and speed, amongst other parameters. However, gaining access to advanced microscopy instrumentation and the expertise necessary to extract meaningful insights from these techniques is an unavoidable hurdle. In this Live Cell Imaging special issue Review, we survey common microscopy-based force measurement techniques and examine how they can be bolstered by emerging microscopy methods. We further explore challenges related to the accompanying data analysis in biomechanical studies and discuss the various resources available to tackle the global issue of technology dissemination, an important avenue for biologists to gain access to pre-commercial instruments that can be leveraged for biomechanical studies.
Collapse
Affiliation(s)
- Chad M. Hobson
- Advanced Imaging Center, Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, United States
| | - Jesse S. Aaron
- Advanced Imaging Center, Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, United States
| | - John M. Heddleston
- Cleveland Clinic Florida Research and Innovation Center, Port St. Lucie, FL, United States
| | - Teng-Leong Chew
- Advanced Imaging Center, Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, United States
| |
Collapse
|
|
26
|
Arif N, Zinnhardt M, Nyamay’Antu A, Teber D, Brückner R, Schaefer K, Li Y, Trappmann B, Grashoff C, Vestweber D. PECAM-1 supports leukocyte diapedesis by tension-dependent dephosphorylation of VE-cadherin. EMBO J 2021; 40:e106113. [PMID: 33604918 PMCID: PMC8090850 DOI: 10.15252/embj.2020106113] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/01/2020] [Revised: 01/15/2021] [Accepted: 01/27/2021] [Indexed: 01/21/2023] Open
Abstract
Leukocyte extravasation is an essential step during the immune response and requires the destabilization of endothelial junctions. We have shown previously that this process depends in vivo on the dephosphorylation of VE-cadherin-Y731. Here, we reveal the underlying mechanism. Leukocyte-induced stimulation of PECAM-1 triggers dissociation of the phosphatase SHP2 which then directly targets VE-cadherin-Y731. The binding site of PECAM-1 for SHP2 is needed for VE-cadherin dephosphorylation and subsequent endocytosis. Importantly, the contribution of PECAM-1 to leukocyte diapedesis in vitro and in vivo was strictly dependent on the presence of Y731 of VE-cadherin. In addition to SHP2, dephosphorylation of Y731 required Ca2+ -signaling, non-muscle myosin II activation, and endothelial cell tension. Since we found that β-catenin/plakoglobin mask VE-cadherin-Y731 and leukocyte docking to endothelial cells exert force on the VE-cadherin-catenin complex, we propose that leukocytes destabilize junctions by PECAM-1-SHP2-triggered dephosphorylation of VE-cadherin-Y731 which becomes accessible by actomyosin-mediated mechanical force exerted on the VE-cadherin-catenin complex.
Collapse
Affiliation(s)
- Nida Arif
- Max Planck Institute for Molecular BiomedicineMünsterGermany
| | - Maren Zinnhardt
- Max Planck Institute for Molecular BiomedicineMünsterGermany
| | | | - Denise Teber
- Max Planck Institute for Molecular BiomedicineMünsterGermany
| | - Randy Brückner
- Max Planck Institute for Molecular BiomedicineMünsterGermany
| | | | - Yu‐Tung Li
- Max Planck Institute for Molecular BiomedicineMünsterGermany
| | | | - Carsten Grashoff
- Institute for Molecular Cell BiologyUniversity of MünsterMünsterGermany
| | | |
Collapse
|
|
27
|
Hang X, He S, Dong Z, Minnick G, Rosenbohm J, Chen Z, Yang R, Chang L. Nanosensors for single cell mechanical interrogation. Biosens Bioelectron 2021; 179:113086. [DOI: 10.1016/j.bios.2021.113086] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2020] [Revised: 02/07/2021] [Accepted: 02/09/2021] [Indexed: 02/08/2023]
|
|
28
|
Fischer LS, Rangarajan S, Sadhanasatish T, Grashoff C. Molecular Force Measurement with Tension Sensors. Annu Rev Biophys 2021; 50:595-616. [PMID: 33710908 DOI: 10.1146/annurev-biophys-101920-064756] [Citation(s) in RCA: 47] [Impact Index Per Article: 11.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
The ability of cells to generate mechanical forces, but also to sense, adapt to, and respond to mechanical signals, is crucial for many developmental, postnatal homeostatic, and pathophysiological processes. However, the molecular mechanisms underlying cellular mechanotransduction have remained elusive for many decades, as techniques to visualize and quantify molecular forces across individual proteins in cells were missing. The development of genetically encoded molecular tension sensors now allows the quantification of piconewton-scale forces that act upon distinct molecules in living cells and even whole organisms. In this review, we discuss the physical principles, advantages, and limitations of this increasingly popular method. By highlighting current examples from the literature, we demonstrate how molecular tension sensors can be utilized to obtain access to previously unappreciated biophysical parameters that define the propagation of mechanical forces on molecular scales. We discuss how the methodology can be further developed and provide a perspective on how the technique could be applied to uncover entirely novel aspects of mechanobiology in the future.
Collapse
Affiliation(s)
- Lisa S Fischer
- Department of Quantitative Cell Biology, Institute of Molecular Cell Biology, University of Münster, Münster D-48149, Germany;
| | - Srishti Rangarajan
- Department of Quantitative Cell Biology, Institute of Molecular Cell Biology, University of Münster, Münster D-48149, Germany;
| | - Tanmay Sadhanasatish
- Department of Quantitative Cell Biology, Institute of Molecular Cell Biology, University of Münster, Münster D-48149, Germany;
| | - Carsten Grashoff
- Department of Quantitative Cell Biology, Institute of Molecular Cell Biology, University of Münster, Münster D-48149, Germany;
| |
Collapse
|
|
29
|
A differentiated Ca 2+ signalling phenotype has minimal impact on myocardin expression in an automated differentiation assay using A7r5 cells. Cell Calcium 2021; 96:102369. [PMID: 33677175 DOI: 10.1016/j.ceca.2021.102369] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2020] [Revised: 02/06/2021] [Accepted: 02/08/2021] [Indexed: 01/12/2023]
Abstract
Vascular smooth muscle cells are unusual in that differentiated, contractile cells possess the capacity to "de-differentiate" into a synthetic phenotype that is characterized by being replicative, secretory, and migratory. One aspect of this phenotypic modulation is a shift from voltage-gated Ca2+ signalling in electrically coupled, differentiated cells to increased dependence on store-operated Ca2+ entry and sarcoplasmic reticulum Ca2+ release in synthetic cells. Conversely, an increased voltage-gated Ca2+ entry is seen when proliferating A7r5 smooth muscle cells quiesce. We asked whether this change in Ca2+ signalling was linked to changes in the expression of the phenotype-regulating transcriptional co-activator myocardin or α-smooth muscle actin, using correlative epifluorescence Ca2+ imaging and immunocytochemistry. Cells were cultured in growth media (DMEM, 10% serum, 25 mM glucose) or differentiation media (DMEM, 1% serum, 5 mM glucose). Coinciding with growth arrest, A7r5 cells became electrically coupled, and spontaneous Ca2+ signalling showed increasing dependence on L-type voltage-gated Ca2+ channels that were blocked with nifedipine (5 μM). These synchronized oscillations were modulated by ryanodine receptors, based on their sensitivity to dantrolene (5 μM). Actively growing cultures had spontaneous Ca2+ transients that were insensitive to nifedipine and dantrolene but were blocked by inhibition of the sarco-endoplasmic reticulum ATPase with cyclopiazonic acid (10 μM). In cells treated with differentiation media, myocardin and αSMA immunoreactivity increased prior to changes in the Ca2+ signalling phenotype, while chronic inhibition of voltage-gated Ca2+ entry modestly increased immunoreactivity of myocardin. Stepwise regression analyses suggested that changes in myocardin expression had a weak relationship with Ca2+ signalling synchronicity, but not frequency or amplitude. In conclusion, we report a 96-well assay and analytical pipeline to study the link between Ca2+ signalling and smooth muscle differentiation. This assay showed that changes in the expression of two molecular differentiation markers (myocardin and αSMA) tended to precede changes in the Ca2+ signalling phenotype.
Collapse
|
|
30
|
Abstract
Bioimage analysis (BIA) has historically helped study how and why cells move; biological experiments evolved in intimate feedback with the most classical image processing techniques because they contribute objectivity and reproducibility to an eminently qualitative science. Cell segmentation, tracking, and morphology descriptors are all discussed here. Using ameboid motility as a case study, these methods help us illustrate how proper quantification can augment biological data, for example, by choosing mathematical representations that amplify initially subtle differences, by statistically uncovering general laws or by integrating physical insight. More recently, the non-invasive nature of quantitative imaging is fertilizing two blooming fields: mechanobiology, where many biophysical measurements remain inaccessible, and microenvironments, where the quest for physiological relevance has exploded data size. From relief to remedy, this trend indicates that BIA is to become a main vector of biological discovery as human visual analysis struggles against ever more complex data.
Collapse
Affiliation(s)
- Aleix Boquet-Pujadas
- Institut Pasteur, Bioimage Analysis Unit, 25 rue du Dr. Roux, Paris Cedex 15 75724, France
- Centre National de la Recherche Scientifique, CNRS UMR3691, Paris, France
- Sorbonne Université, Paris 75005, France
| | - Jean-Christophe Olivo-Marin
- Institut Pasteur, Bioimage Analysis Unit, 25 rue du Dr. Roux, Paris Cedex 15 75724, France
- Centre National de la Recherche Scientifique, CNRS UMR3691, Paris, France
| | - Nancy Guillén
- Institut Pasteur, Bioimage Analysis Unit, 25 rue du Dr. Roux, Paris Cedex 15 75724, France
- Centre National de la Recherche Scientifique, CNRS ERL9195, Paris, France
| |
Collapse
|
|
31
|
De La Pena A, Mukhtar M, Yokosawa R, Carrasquilla S, Simmons CS. Quantifying cellular forces: Practical considerations of traction force microscopy for dermal fibroblasts. Exp Dermatol 2021; 30:74-83. [PMID: 32767472 PMCID: PMC7769991 DOI: 10.1111/exd.14166] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2020] [Revised: 07/12/2020] [Accepted: 07/30/2020] [Indexed: 12/28/2022]
Abstract
Traction force microscopy (TFM) is a well-established technique traditionally used by biophysicists to quantify the forces adherent biological cells exert on their microenvironment. As image processing software becomes increasingly user-friendly, TFM is being adopted by broader audiences to quantify contractility of (myo)fibroblasts. While many technical reviews of TFM's computational mechanics are available, this review focuses on practical experimental considerations for dermatology researchers new to cell mechanics and TFM who may wish to implement a higher throughput and less expensive alternative to collagen compaction assays. Here, we describe implementation of experimental methods, analysis using open-source software and troubleshooting of common issues to enable researchers to leverage TFM for their investigations into skin fibroblasts.
Collapse
Affiliation(s)
| | | | | | | | - Chelsey S. Simmons
- Department of Mechanical and Aerospace Engineering
- J. Crayton Pruitt Department of Biomedical Engineering
- Division of Cardiovascular Medicine, University of Florida
| |
Collapse
|
|
32
|
Xenopus Deep Cell Aggregates: A 3D Tissue Model for Mesenchymal-to-Epithelial Transition. Methods Mol Biol 2021; 2179:275-287. [PMID: 32939727 PMCID: PMC9972462 DOI: 10.1007/978-1-0716-0779-4_21] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/02/2023]
Abstract
Mesenchymal-to-epithelial transition (MET) describes the ability of loosely associated migratory cells to form a more adherent sheet-like assembly of cells. MET is a conserved motif occurring throughout organogenesis and plays a key role in regeneration and cancer metastasis, and is the first step in producing induced pluripotent stem cells (iPSCs). To resolve fundamental biological questions about MET, its relation to epithelial-to-mesenchymal transition, and to explore MET's role in tissue assembly and remodeling requires live models for MET that are amenable to experimentation. Many cases of clinically important MET are inferred since they occur deep with the body of the embryo or adult. We have developed a tractable model for MET, where cellular transitions can be directly observed under conditions where molecular, mechanical, and cellular contexts can be controlled experimentally. In this chapter, we introduce a 3-dimensional (3D) tissue model to study MET using Xenopus laevis embryonic mesenchymal cell aggregates.
Collapse
|
|
33
|
Bajpai A, Li R, Chen W. The cellular mechanobiology of aging: from biology to mechanics. Ann N Y Acad Sci 2020; 1491:3-24. [PMID: 33231326 DOI: 10.1111/nyas.14529] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/26/2020] [Revised: 10/10/2020] [Accepted: 10/21/2020] [Indexed: 12/14/2022]
Abstract
Aging is a chronic, complicated process that leads to degenerative physical and biological changes in living organisms. Aging is associated with permanent, gradual physiological cellular decay that affects all aspects of cellular mechanobiological features, including cellular cytoskeleton structures, mechanosensitive signaling pathways, and forces in the cell, as well as the cell's ability to sense and adapt to extracellular biomechanical signals in the tissue environment through mechanotransduction. These mechanobiological changes in cells are directly or indirectly responsible for dysfunctions and diseases in various organ systems, including the cardiovascular, musculoskeletal, skin, and immune systems. This review critically examines the role of aging in the progressive decline of the mechanobiology occurring in cells, and establishes mechanistic frameworks to understand the mechanobiological effects of aging on disease progression and to develop new strategies for halting and reversing the aging process. Our review also highlights the recent development of novel bioengineering approaches for studying the key mechanobiological mechanisms in aging.
Collapse
Affiliation(s)
- Apratim Bajpai
- Department of Mechanical and Aerospace Engineering, Tandon School of Engineering, New York University, Brooklyn, New York
| | - Rui Li
- Department of Mechanical and Aerospace Engineering, Tandon School of Engineering, New York University, Brooklyn, New York.,Department of Biomedical Engineering, Tandon School of Engineering, New York University, Brooklyn, New York
| | - Weiqiang Chen
- Department of Mechanical and Aerospace Engineering, Tandon School of Engineering, New York University, Brooklyn, New York.,Department of Biomedical Engineering, Tandon School of Engineering, New York University, Brooklyn, New York.,Laura and Isaac Perlmutter Cancer Center, NYU Langone Health, New York, New York
| |
Collapse
|
|
34
|
Traeger H, Kiebala DJ, Weder C, Schrettl S. From Molecules to Polymers-Harnessing Inter- and Intramolecular Interactions to Create Mechanochromic Materials. Macromol Rapid Commun 2020; 42:e2000573. [PMID: 33191595 DOI: 10.1002/marc.202000573] [Citation(s) in RCA: 62] [Impact Index Per Article: 12.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2020] [Revised: 10/16/2020] [Indexed: 12/30/2022]
Abstract
The development of mechanophores as building blocks that serve as predefined weak linkages has enabled the creation of mechanoresponsive and mechanochromic polymer materials, which are interesting for a range of applications including the study of biological specimens or advanced security features. In typical mechanophores, covalent bonds are broken when polymers that contain these chemical motifs are exposed to mechanical forces, and changes of the optical properties upon bond scission can be harnessed as a signal that enables the detection of applied mechanical stresses and strains. Similar chromic effects upon mechanical deformation of polymers can also be achieved without relying on the scission of covalent bonds. The dissociation of motifs that feature directional noncovalent interactions, the disruption of aggregated molecules, and conformational changes in molecules or polymers constitute an attractive element for the design of mechanoresponsive and mechanochromic materials. In this article, it is reviewed how such alterations of molecules and polymers can be exploited for the development of mechanochromic materials that signal deformation without breaking covalent bonds. Recent illustrative examples are highlighted that showcase how the use of such mechanoresponsive motifs enables the visual mapping of stresses and damage in a reversible and highly sensitive manner.
Collapse
Affiliation(s)
- Hanna Traeger
- Adolphe Merkle Institute, University of Fribourg, Chemin des Verdiers 4, Fribourg, CH-1700, Switzerland
| | - Derek J Kiebala
- Adolphe Merkle Institute, University of Fribourg, Chemin des Verdiers 4, Fribourg, CH-1700, Switzerland
| | - Christoph Weder
- Adolphe Merkle Institute, University of Fribourg, Chemin des Verdiers 4, Fribourg, CH-1700, Switzerland
| | - Stephen Schrettl
- Adolphe Merkle Institute, University of Fribourg, Chemin des Verdiers 4, Fribourg, CH-1700, Switzerland
| |
Collapse
|
|
35
|
Engel MC, Romano F, Louis AA, Doye JPK. Measuring Internal Forces in Single-Stranded DNA: Application to a DNA Force Clamp. J Chem Theory Comput 2020; 16:7764-7775. [PMID: 33147408 DOI: 10.1021/acs.jctc.0c00286] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
We present a new method for calculating internal forces in DNA structures using coarse-grained models and demonstrate its utility with the oxDNA model. The instantaneous forces on individual nucleotides are explored and related to model potentials, and using our framework, internal forces are calculated for two simple DNA systems and for a recently published nanoscopic force clamp. Our results highlight some pitfalls associated with conventional methods for estimating internal forces, which are based on elastic polymer models, and emphasize the importance of carefully considering secondary structure and ionic conditions when modeling the elastic behavior of single-stranded DNA. Beyond its relevance to the DNA nanotechnological community, we expect our approach to be broadly applicable to calculations of internal force in a variety of structures-from DNA to protein-and across other coarse-grained simulation models.
Collapse
Affiliation(s)
- Megan C Engel
- School of Engineering and Applied Sciences, Harvard University, 29 Oxford Street, Cambridge, Massachusetts 02138, United States.,Rudolf Peierls Centre for Theoretical Physics, University of Oxford, 1 Keble Road, Oxford, OX1 3NP, U.K
| | - Flavio Romano
- Dipartimento di Scienze Molecolari e Nanosistemi, Universitá Ca Foscari di Venezia, Via Torino 155, 30172, Venezia Mestre, Italy
| | - Ard A Louis
- Rudolf Peierls Centre for Theoretical Physics, University of Oxford, 1 Keble Road, Oxford, OX1 3NP, U.K
| | - Jonathan P K Doye
- Department of Chemistry, University of Oxford, South Parks Road, Oxford, OX1 3QZ, U.K
| |
Collapse
|
|
36
|
Kolawole EM, Lamb TJ, Evavold BD. Relationship of 2D Affinity to T Cell Functional Outcomes. Int J Mol Sci 2020; 21:E7969. [PMID: 33120989 PMCID: PMC7662510 DOI: 10.3390/ijms21217969] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2020] [Revised: 10/14/2020] [Accepted: 10/23/2020] [Indexed: 12/11/2022] Open
Abstract
T cells are critical for a functioning adaptive immune response and a strong correlation exists between T cell responses and T cell receptor (TCR): peptide-loaded MHC (pMHC) binding. Studies that utilize pMHC tetramer, multimers, and assays of three-dimensional (3D) affinity have provided advancements in our understanding of T cell responses across different diseases. However, these technologies focus on higher affinity and avidity T cells while missing the lower affinity responders. Lower affinity TCRs in expanded polyclonal populations almost always constitute a significant proportion of the response with cells mediating different effector functions associated with variation in the proportion of high and low affinity T cells. Since lower affinity T cells expand and are functional, a fully inclusive view of T cell responses is required to accurately interpret the role of affinity for adaptive T cell immunity. For example, low affinity T cells are capable of inducing autoimmune disease and T cells with an intermediate affinity have been shown to exhibit an optimal anti-tumor response. Here, we focus on how affinity of the TCR may relate to T cell phenotype and provide examples where 2D affinity influences functional outcomes.
Collapse
Affiliation(s)
| | | | - Brian D. Evavold
- Department of Pathology, University of Utah, 15 N Medical Drive, Salt Lake City, UT 84112, USA; (E.M.K.); (T.J.L.)
| |
Collapse
|
|
37
|
|
|
38
|
Abstract
At the nanoscale, pushing, pulling, and shearing forces drive biochemical processes in development and remodeling as well as in wound healing and disease progression. Research in the field of mechanobiology investigates not only how these loads affect biochemical signaling pathways but also how signaling pathways respond to local loading by triggering mechanical changes such as regional stiffening of a tissue. This feedback between mechanical and biochemical signaling is increasingly recognized as fundamental in embryonic development, tissue morphogenesis, cell signaling, and disease pathogenesis. Historically, the interdisciplinary field of mechanobiology has been driven by the development of technologies for measuring and manipulating cellular and molecular forces, with each new tool enabling vast new lines of inquiry. In this review, we discuss recent advances in the manufacturing and capabilities of molecular-scale force and strain sensors. We also demonstrate how DNA nanotechnology has been critical to the enhancement of existing techniques and to the development of unique capabilities for future mechanosensor assembly. DNA is a responsive and programmable building material for sensor fabrication. It enables the systematic interrogation of molecular biomechanics with forces at the 1- to 200-pN scale that are needed to elucidate the fundamental means by which cells and proteins transduce mechanical signals.
Collapse
Affiliation(s)
- Susana M. Beltrán
- Department of Mechanical Engineering, Carnegie Mellon
University, Pittsburgh, Pennsylvania
| | - Marvin J. Slepian
- Department of Medicine and Sarver Heart Center, University
of Arizona, Tucson
- Department of Biomedical Engineering, University of
Arizona, Tucson
- Department of Materials Science and Engineering, University
of Arizona, Tucson
| | - Rebecca E. Taylor
- Department of Mechanical Engineering, Carnegie Mellon
University, Pittsburgh, Pennsylvania
- Department of Biomedical Engineering, Carnegie Mellon
University, Pittsburgh, Pennsylvania
- Department of Electrical and Computer Engineering, Carnegie
Mellon University, Pittsburgh, Pennsylvania
| |
Collapse
|
|
39
|
Noll N, Streichan SJ, Shraiman BI. A variational method for image-based inference of internal stress in epithelial tissues. PHYSICAL REVIEW. X 2020; 10:011072. [PMID: 33767909 PMCID: PMC7989596 DOI: 10.1103/physrevx.10.011072] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 06/12/2023]
Abstract
Cellular mechanics drives epithelial morphogenesis, the process wherein cells collectively rearrange to produce tissue-scale deformations that determine organismal shape. However, quantitative understanding of tissue mechanics is impaired by the difficulty of direct measurement of stress in-vivo. This difficulty has spurred the development of image-based inference algorithms that estimate stress from snapshots of epithelial geometry. Such methods are challenged by sensitivity to measurement error and thus require accurate geometric segmentation for practical use. We overcome this difficulty by introducing a novel approach - the Variational Method of Stress Inference (VMSI) - which exploits the fundamental duality between stress and geometry at equilibrium of discrete mechanical networks that model confluent cellular layers. We approximate the apical geometry of an epithelial tissue by a 2D tiling with Circular Arc Polygons (CAP) in which arcs represent intercellular interfaces defined by the balance of local line tension and pressure differentials between adjacent cells. The mechanical equilibrium of such networks imposes extensive local constraints on CAP geometry. These constraints provide the foundation of VMSI which, starting with images of epithelial monolayers, simultaneously approximates both tissue geometry and internal forces, subject to the constraint of equilibrium. We find VMSI to be more robust than previous methods. Specifically, the VMSI performance is validated by the comparison of the predicted cellular and mesoscopic scale stress with the measured myosin II patterns during early Drosophila embryogenesis. VMSI prediction of mesoscopic stress tensor correlates at the 80% level with the measured myosin distribution and reveals that most of the myosin activity in that case is involved in a static internal force balance within the epithelial layer. In addition to insight into cell mechanics, this study provides a practical method for non-destructive estimation of stress in live epithelial tissue.
Collapse
Affiliation(s)
- Nicholas Noll
- Department of Physics, University of California Santa Barbara
- Biozentrum, University of Basel
- Swiss Institute of Bioinformatics
| | - Sebastian J Streichan
- Department of Physics, University of California Santa Barbara
- Kavli Institute for Theoretical Physics
| | - Boris I Shraiman
- Department of Physics, University of California Santa Barbara
- Kavli Institute for Theoretical Physics
| |
Collapse
|
|
40
|
Zhang J, Chada NC, Reinhart-King CA. Microscale Interrogation of 3D Tissue Mechanics. Front Bioeng Biotechnol 2019; 7:412. [PMID: 31921816 PMCID: PMC6927918 DOI: 10.3389/fbioe.2019.00412] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2019] [Accepted: 11/28/2019] [Indexed: 01/02/2023] Open
Abstract
Cells in vivo live in a complex microenvironment composed of the extracellular matrix (ECM) and other cells. Growing evidence suggests that the mechanical interaction between the cells and their microenvironment is of critical importance to their behaviors under both normal and diseased conditions, such as migration, differentiation, and proliferation. The study of tissue mechanics in the past two decades, including the assessment of both mechanical properties and mechanical stresses of the extracellular microenvironment, has greatly enriched our knowledge about how cells interact with their mechanical environment. Tissue mechanical properties are often heterogeneous and sometimes anisotropic, which makes them difficult to obtain from macroscale bulk measurements. Mechanical stresses were first measured for cells cultured on two-dimensional (2D) surfaces with well-defined mechanical properties. While 2D measurements are relatively straightforward and efficient, and they have provided us with valuable knowledge on cell-ECM interactions, that knowledge may not be directly applicable to in vivo systems. Hence, the measurement of tissue stresses in a more physiologically relevant three-dimensional (3D) environment is required. In this mini review, we will summarize and discuss recent developments in using optical, magnetic, genetic, and mechanical approaches to interrogate 3D tissue stresses and mechanical properties at the microscale.
Collapse
|
|
41
|
Yasunaga A, Murad Y, Li ITS. Quantifying molecular tension-classifications, interpretations and limitations of force sensors. Phys Biol 2019; 17:011001. [PMID: 31387091 DOI: 10.1088/1478-3975/ab38ff] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
Molecular force sensors (MFSs) have grown to become an important tool to study the mechanobiology of cells and tissues. They provide a minimally invasive means to optically report mechanical interactions at the molecular level. One of the challenges in molecular force sensor studies is the interpretation of the fluorescence readout. In this review, we divide existing MFSs into three classes based on the force-sensing mechanism (reversibility) and the signal output (analog/digital). From single-molecule force spectroscopy (SMFS) perspectives, we provided a critical discussion on how the sensors respond to force and how the different sensor designs affect the interpretation of their fluorescence readout. Lastly, the review focuses on the limitations and attention one must pay in designing MFSs and biological experiments using them; in terms of their tunability, signal-to-noise ratio (SNR), and perturbation of the biological system under investigation.
Collapse
Affiliation(s)
- Adam Yasunaga
- These authors contributed equally to the manuscript (co-first author)
| | | | | |
Collapse
|
|
42
|
Barbic M. Possible magneto-mechanical and magneto-thermal mechanisms of ion channel activation in magnetogenetics. eLife 2019; 8:45807. [PMID: 31373554 PMCID: PMC6693891 DOI: 10.7554/elife.45807] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2019] [Accepted: 07/28/2019] [Indexed: 01/11/2023] Open
Abstract
The palette of tools for perturbation of neural activity is continually expanding. On the forefront of this expansion is magnetogenetics, where ion channels are genetically engineered to be closely coupled to the iron-storage protein ferritin. Initial reports on magnetogenetics have sparked a vigorous debate on the plausibility of physical mechanisms of ion channel activation by means of external magnetic fields. The criticism leveled against magnetogenetics as being physically implausible is based on the specific assumptions about the magnetic spin configurations of iron in ferritin. I consider here a wider range of possible spin configurations of iron in ferritin and the consequences these might have in magnetogenetics. I propose several new magneto-mechanical and magneto-thermal mechanisms of ion channel activation that may clarify some of the mysteries that presently challenge our understanding of the reported biological experiments. Finally, I present some additional puzzles that will require further theoretical and experimental investigation.
Collapse
Affiliation(s)
- Mladen Barbic
- Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, United States
| |
Collapse
|
|
43
|
Shams H, Hoffman BD, Mofrad MRK. The "Stressful" Life of Cell Adhesion Molecules: On the Mechanosensitivity of Integrin Adhesome. J Biomech Eng 2019; 140:2667887. [PMID: 29272321 DOI: 10.1115/1.4038812] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2017] [Indexed: 02/06/2023]
Abstract
Cells have evolved into complex sensory machines that communicate with their microenvironment via mechanochemical signaling. Extracellular mechanical cues trigger complex biochemical pathways in the cell, which regulate various cellular processes. Integrin-mediated focal adhesions (FAs) are large multiprotein complexes, also known as the integrin adhesome, that link the extracellular matrix (ECM) to the actin cytoskeleton, and are part of powerful intracellular machinery orchestrating mechanotransduction pathways. As forces are transmitted across FAs, individual proteins undergo structural and functional changes that involve a conversion of chemical to mechanical energy. The local composition of early adhesions likely defines the regional stress levels and determines the type of newly recruited proteins, which in turn modify the local stress distribution. Various approaches have been used for detecting and exploring molecular mechanisms through which FAs are spatiotemporally regulated, however, many aspects are yet to be understood. Current knowledge on the molecular mechanisms of mechanosensitivity in adhesion proteins is discussed herein along with important questions yet to be addressed, are discussed.
Collapse
Affiliation(s)
- Hengameh Shams
- Molecular Cell Biomechanics Laboratory, Departments of Bioengineering and Mechanical Engineering, University of California, Berkeley, CA 94720-1762
| | - Brenton D Hoffman
- Department of Biomedical Engineering, Duke University, Durham, NC 27708
| | - Mohammad R K Mofrad
- Molecular Cell Biomechanics Laboratory, Departments of Bioengineering and Mechanical Engineering, University of California, 208A Stanley Hall #1762, Berkeley, CA 94720-1762.,Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Lab, Berkeley, CA 94720 e-mail:
| |
Collapse
|
|
44
|
Ungai-Salánki R, Peter B, Gerecsei T, Orgovan N, Horvath R, Szabó B. A practical review on the measurement tools for cellular adhesion force. Adv Colloid Interface Sci 2019; 269:309-333. [PMID: 31128462 DOI: 10.1016/j.cis.2019.05.005] [Citation(s) in RCA: 50] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2019] [Revised: 05/05/2019] [Accepted: 05/06/2019] [Indexed: 01/03/2023]
Abstract
Cell-cell and cell-matrix adhesions are fundamental in all multicellular organisms. They play a key role in cellular growth, differentiation, pattern formation and migration. Cell-cell adhesion is substantial in the immune response, pathogen-host interactions, and tumor development. The success of tissue engineering and stem cell implantations strongly depends on the fine control of live cell adhesion on the surface of natural or biomimetic scaffolds. Therefore, the quantitative and precise measurement of the adhesion strength of living cells is critical, not only in basic research but in modern technologies, too. Several techniques have been developed or are under development to quantify cell adhesion. All of them have their pros and cons, which has to be carefully considered before the experiments and interpretation of the recorded data. Current review provides a guide to choose the appropriate technique to answer a specific biological question or to complete a biomedical test by measuring cell adhesion.
Collapse
|
|
45
|
Bene L, Damjanovich L. Förster Resonance Energy Transfer Pioneers Biomechanics: Molecular-Scale Force Meters for Visualizing Intracellular Stress Fields. Cytometry A 2019; 95:819-822. [PMID: 31034713 DOI: 10.1002/cyto.a.23780] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2019] [Revised: 04/08/2019] [Accepted: 04/13/2019] [Indexed: 12/29/2022]
Affiliation(s)
- László Bene
- Department of Surgery, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
| | - László Damjanovich
- Department of Surgery, Faculty of Medicine, University of Debrecen, Debrecen, Hungary
| |
Collapse
|
|
46
|
Chen T, Guo Y, Shan J, Zhang J, Shen X, Guo J, Liu XM. Vector Analysis of Cytoskeletal Structural Tension and the Mechanisms that Underpin Spectrin-Related Forces in Pyroptosis. Antioxid Redox Signal 2019; 30:1503-1520. [PMID: 29669427 DOI: 10.1089/ars.2017.7366] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/22/2023]
Abstract
Aims: Pyroptotic cells are characterized by plasma swelling, membrane blebbing, and disintegration of the cell membrane mediated by spectrin-based membrane skeleton and intercellular competitive tension activities. The spectrin-based membrane skeleton is involved in membrane organization through the regulation of intercellular tension. Using genetically encoded tension sensors to attain noninvasive force measurements in structural proteins, we investigated how cytoskeletal structural tension influences changes in plasma morphology during pyroptosis and the regulatory mechanism of cytoskeletal structural tension that underpins pyroptosis. Results: The results indicate that increasing spectrin tension is caused by osmotic swelling. Hightened tension of spectrin was closely associated with the shrink tension transmitted synergistically by microfilaments (MFs) and microtubules (MTs). However, the increment of spectrin tension in pyroptotic cells was controlled antagonistically by MF and MT forces. Different from MF tension, outward MT forces participated in the formation of membrane blebs. Spectrin tension caused by inward MF forces resisted pyroptosis swelling. Stabilization of MF and MT structure had little influence on intracellular tension and pyroptosis deformation. Pyroptosis-induced cytoskeletal structural tension was highly dependent on calcium signaling and reactive oxygen species generation. Blocking of membrane pores, nonselective ion flux, or elimination of caspase-1 cleavage resulted in the remission of structural forces associated with pyroptosis failure. Innovation and Conclusions: The data suggest that subcellular tension, in terms of magnitude and vector, is integral to pyroptosis through the mediation of swelling and blebbing and the elimination of structural tension, especially MT forces, may result in pyroptosis inhibition.
Collapse
Affiliation(s)
- Tingting Chen
- Department of Biochemistry and Molecular Biology, Nanjing Medical University, Nanjing, People's Republic of China.,State Key Laboratory Cultivation Base for TCM Quality and Efficacy, School of Medicine and Life Science, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China
| | - Yichen Guo
- Key Laboratory of Drug Target and Drug for Degenerative Disease, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China.,Department of Biomedical Engineering, University of Alabama at Birmingham (UAB), Birmingham, Alabama
| | - Jinjun Shan
- Jiangsu Key Laboratory of Pediatric Respiratory Disease, Institute of Pediatrics, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China
| | - Jiarui Zhang
- State Key Laboratory Cultivation Base for TCM Quality and Efficacy, School of Medicine and Life Science, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China.,Key Laboratory of Drug Target and Drug for Degenerative Disease, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China
| | - Xu Shen
- State Key Laboratory Cultivation Base for TCM Quality and Efficacy, School of Medicine and Life Science, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China.,Key Laboratory of Drug Target and Drug for Degenerative Disease, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China
| | - Jun Guo
- Department of Biochemistry and Molecular Biology, Nanjing Medical University, Nanjing, People's Republic of China.,State Key Laboratory Cultivation Base for TCM Quality and Efficacy, School of Medicine and Life Science, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China.,Key Laboratory of Drug Target and Drug for Degenerative Disease, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China.,Jiangsu Key Laboratory of Pediatric Respiratory Disease, Institute of Pediatrics, Nanjing University of Chinese Medicine, Nanjing, People's Republic of China
| | - Xiaoguang Margaret Liu
- Department of Biomedical Engineering, University of Alabama at Birmingham (UAB), Birmingham, Alabama
| |
Collapse
|
|
47
|
Cost A, Khalaji S, Grashoff C. Genetically Encoded FRET‐Based Tension Sensors. ACTA ACUST UNITED AC 2019; 83:e85. [DOI: 10.1002/cpcb.85] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Affiliation(s)
- Anna‐Lena Cost
- Group of Molecular Mechanotransduction, Max Planck Institute of Biochemistry Martinsried Germany
- Department of Quantitative Cell Biology, Institute of Molecular Cell BiologyUniversity of Münster Münster Germany
| | - Samira Khalaji
- Group of Molecular Mechanotransduction, Max Planck Institute of Biochemistry Martinsried Germany
- Department of Quantitative Cell Biology, Institute of Molecular Cell BiologyUniversity of Münster Münster Germany
| | - Carsten Grashoff
- Group of Molecular Mechanotransduction, Max Planck Institute of Biochemistry Martinsried Germany
- Department of Quantitative Cell Biology, Institute of Molecular Cell BiologyUniversity of Münster Münster Germany
| |
Collapse
|
|
48
|
Mechanical loading of desmosomes depends on the magnitude and orientation of external stress. Nat Commun 2018; 9:5284. [PMID: 30538252 PMCID: PMC6290003 DOI: 10.1038/s41467-018-07523-0] [Citation(s) in RCA: 63] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/16/2018] [Accepted: 11/08/2018] [Indexed: 01/19/2023] Open
Abstract
Desmosomes are intercellular adhesion complexes that connect the intermediate filament cytoskeletons of neighboring cells, and are essential for the mechanical integrity of mammalian tissues. Mutations in desmosomal proteins cause severe human pathologies including epithelial blistering and heart muscle dysfunction. However, direct evidence for their load-bearing nature is lacking. Here we develop Förster resonance energy transfer (FRET)-based tension sensors to measure the forces experienced by desmoplakin, an obligate desmosomal protein that links the desmosomal plaque to intermediate filaments. Our experiments reveal that desmoplakin does not experience significant tension under most conditions, but instead becomes mechanically loaded when cells are exposed to external mechanical stresses. Stress-induced loading of desmoplakin is transient and sensitive to the magnitude and orientation of the applied tissue deformation, consistent with a stress absorbing function for desmosomes that is distinct from previously analyzed cell adhesion complexes. Desmosomes are intercellular adhesion complexes that connect the intermediate filament cytoskeletons of neighboring cells but direct evidence for their load-bearing nature is lacking. Here the authors develop FRET-based tension sensors to measure the forces experienced by desmoplakin and infer that desmosomes become mechanically loaded when cells are exposed to external mechanical stresses.
Collapse
|
|
49
|
Kilinc D. The Emerging Role of Mechanics in Synapse Formation and Plasticity. Front Cell Neurosci 2018; 12:483. [PMID: 30574071 PMCID: PMC6291423 DOI: 10.3389/fncel.2018.00483] [Citation(s) in RCA: 42] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2018] [Accepted: 11/27/2018] [Indexed: 12/26/2022] Open
Abstract
The regulation of synaptic strength forms the basis of learning and memory, and is a key factor in understanding neuropathological processes that lead to cognitive decline and dementia. While the mechanical aspects of neuronal development, particularly during axon growth and guidance, have been extensively studied, relatively little is known about the mechanical aspects of synapse formation and plasticity. It is established that a filamentous actin network with complex spatiotemporal behavior controls the dendritic spine shape and size, which is thought to be crucial for activity-dependent synapse plasticity. Accordingly, a number of actin binding proteins have been identified as regulators of synapse plasticity. On the other hand, a number of cell adhesion molecules (CAMs) are found in synapses, some of which form transsynaptic bonds to align the presynaptic active zone (PAZ) with the postsynaptic density (PSD). Considering that these CAMs are key components of cellular mechanotransduction, two critical questions emerge: (i) are synapses mechanically regulated? and (ii) does disrupting the transsynaptic force balance lead to (or exacerbate) synaptic failure? In this mini review article, I will highlight the mechanical aspects of synaptic structures-focusing mainly on cytoskeletal dynamics and CAMs-and discuss potential mechanoregulation of synapses and its relevance to neurodegenerative diseases.
Collapse
Affiliation(s)
- Devrim Kilinc
- INSERM U1167, Institut Pasteur de Lille, Lille, France
| |
Collapse
|
|
50
|
Teunissen AJP, Pérez-Medina C, Meijerink A, Mulder WJM. Investigating supramolecular systems using Förster resonance energy transfer. Chem Soc Rev 2018; 47:7027-7044. [PMID: 30091770 PMCID: PMC6441672 DOI: 10.1039/c8cs00278a] [Citation(s) in RCA: 88] [Impact Index Per Article: 12.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/24/2023]
Abstract
Supramolecular systems have applications in areas as diverse as materials science, biochemistry, analytical chemistry, and nanomedicine. However, analyzing such systems can be challenging due to the wide range of time scales, binding strengths, distances, and concentrations at which non-covalent phenomena take place. Due to their versatility and sensitivity, Förster resonance energy transfer (FRET)-based techniques are excellently suited to meet such challenges. Here, we detail the ways in which FRET has been used to study non-covalent interactions in both synthetic and biological supramolecular systems. Among other topics, we examine methods to measure molecular forces, determine protein conformations, monitor assembly kinetics, and visualize in vivo drug release from nanoparticles. Furthermore, we highlight multiplex FRET techniques, discuss the field's limitations, and provide a perspective on new developments.
Collapse
Affiliation(s)
- Abraham J. P. Teunissen
- Translational and Molecular Imaging Institute, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY 10029, USA
| | - Carlos Pérez-Medina
- Translational and Molecular Imaging Institute, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY 10029, USA
| | - Andries Meijerink
- Department of Chemistry, Utrecht University, Princetonplein 1, 3584 CC Utrecht, The Netherlands
| | - Willem J. M. Mulder
- Translational and Molecular Imaging Institute, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY 10029, USA
- Department of Medical Biochemistry, Academic Medical Center, Meibergdreef 9, 1105 AZ Amsterdam, The Netherlands
- Laboratory of Chemical biology, Department of Biomedical Engineering and Institute for Complex Molecular systems, Eindhoven University of Technology, P.O. Box 513, 5600 MB, The Netherlands
| |
Collapse
|