Parasitic roundworms are remarkable for their ability to manipulate host immune systems and ameliorate inflammatory diseases. Although much is known about the nature of nematode effectors in immune modulation, little is known about the action mode of these molecules. Here, we report that a serine protease inhibitor SPI-I8 in the extracellular vesicles of blood-feeding nematodes like Ancylostoma ceylanicum, Haemonchus contortus and Nippostrongylus brasiliensis, effectively halts excessive inflammatory responses in vitro and in vivo. We demonstrate that H. contortus SPI-I8 promotes the role of a negative regulator of RACK1 and enhances the effects of RACK1 on tumor necrosis factor (TNF)-α-IκB kinases (IKKs)-nuclear factor kappa beta (NF-κB) axis in mammalian cells, by hijacking E3 ubiquitin protein ligase MKRN1-mediated polyubiquitination of RACK1. Administration of recombinant N. brasiliensis SPI-I8 effectively protects mice from dextran sulfate sodium (DSS)-induced colitis and lipopolysaccharide (LPS)-induced sepsis. Considering the structural and functional conservation of SPI-I8s among Strongylida nematodes and the conservation of interactive mediators (i.e., MKRN1 and RACK1) among mammals, our findings provide insights into the host-parasite interface where parasitic roundworms secret molecules to suppress host inflammatory responses. Harnessing these findings should underpin the exploitation of nematode's immunomodulators to relief excessive inflammation associated diseases in animals and humans.

The WD40 repeat proteins are crucial components of eukaryotic genomes and contribute to a wide array of plant developmental processes and environmental interactions. However, the true extent of intraspecific WD40 diversity in plants is unclear.

We defined a nearly complete species-wide pan-WD40ome in maize based on the published genome sequences of 26 nested association mapping (NAM) population founders. The pan-WD40ome largely saturated with inclusion of approximately 20 inbred lines, with about 95% of the pan-WD40ome being present in at least two founders. The architectural diversity of the WD40 domains, additional domains, and consequent spatial protein structures suggested the functional diversity of the maize pan-WD40ome. This finding was supported by significant associations between 87 WD40 genes and 19 agronomic, 3 kernel-quality, and 3 biotic-stress traits, as well as the multiple molecular pathways through which the trait-associated WD40 genes were predicted to function. In addition, WD40 genes exhibited abundant genomic variations among the NAM founders. Sequence analysis indicated that gene duplications and gene translocations caused by Helitron transposons may play important roles in the amplification of WD40 genes during the evolution of the maize WD40 gene family.

In summary, this study provides a comprehensive framework for understanding the structural and functional diversity of the pan-WD40ome in maize and other agronomically important species with complex genomes, as well as excellent candidate genes/alleles for maize genetic improvement.

Sweet wormwood (Artemisia annua), an annual herb belonging to the Compositae family, is the main source of the potent anti-malarial drug artemisinin, which is mainly produced in glandular trichomes of A. annua leaves. The WD40 protein family is one of the largest protein families in eukaryotes and plays crucial roles in regulating plant growth and development, stress responses, and secondary metabolite biosynthesis. However, WD40 proteins have not been comprehensively identified in A. annua. In this study, we identified 236 WD40 proteins in the A. annua genome and examined their conserved domains, motifs, and cis-regulatory elements, gene structures, chromosomal distribution, duplication events of their encoding genes. Furthermore, we isolated and characterized TRANSPARENT TESTA GLABROUS 1 (AaTTG1), a homolog of Arabidopsis TTG1, and confirmed that AaTTG1 was localized to the nucleus and cytoplasm. Indeed, AaTTG1 can rescue the glabrous phenotype of the Arabidopsis ttg1 mutant and enhanced trichome production when heterologously expressed in wild-type Arabidopsis plants. Transgenic A. annua lines overexpressing AaTTG1 displayed a significantly higher density of glandular trichomes and higher artemisinin contents. Transgenic A. annua lines with inhibited AaTTG1 function had fewer glandular trichomes and lower artemisinin levels. Moreover, we demonstrated that AaTTG1 positively regulates glandular trichome development in A. annua through interactions with AaSPL9. This study thus provides fundamental insights into the role of WD40 proteins in A. annua and introduces a promising approach to enhance artemisinin production by manipulating glandular trichome development in this valuable medicinal plant.

COP1 is the essential protein that integrates various environmental and hormonal cues to control plant growth and development at multiple levels. COP1 is a RING-finger-type E3 ubiquitin ligase that acts as a potent repressor of photomorphogenesis and flowering by targeting numerous substrates for ubiquitination and promoting their proteolysis via the 26S proteasome system. The WD40 repeat domain with conserved amino acid residues was shown to be essential for interacting with its targets. However, the role of these amino acids in regulating hypocotyl growth and flowering in response to varying light and temperatures remains unknown. Here, we show that tryptophan amino acid at the position 467 residue (COP1W467) is relevant in mediating the interaction with its targets to regulate the COP1-mediated proteolysis. The COP1W467 plays a critical role in inducing growth responses in shade light by interacting and degrading HY5, a crucial negative regulator of shade-avoidance response (SAR). Moreover, COP1W467 integrates warm ambient temperature signals to promote hypocotyl growth by increasing PIF4 and decreasing HY5 protein stability. Finally, we found that COP1W467 is important in inhibiting flowering under a short-day photoperiod, likely through interacting with CO for degradation. Together, this study highlights that the COP1W467 residue is essential to regulate seedling photomorphogenesis, SAR, thermomorphogenesis and flowering for the fine adjustment of plant growth and development under dynamic light and temperature conditions.

The flagellum in Trypanosoma brucei plays crucial roles in cell locomotion, cell morphogenesis, and cell division, and its inheritance depends on the faithful duplication of multiple flagellum-associated structures. One of such cytoskeletal structures is a hairpin-like structure termed the hook complex composed of a fishhook-like structure and a centrin arm structure, whose cellular functions remain poorly understood. We recently identified KIN-G, an orphan kinesin that promotes hook complex and Golgi biogenesis. Here we report a WD40 repeats-containing protein named WDR2, which interacts with and regulates KIN-G. WDR2 co-localizes with KIN-G at the centrin arm, and knockdown of WDR2 disrupts hook complex integrity and morphology, inhibits flagellum attachment zone elongation and flagellum positioning, and eventually arrests cytokinesis. Knockdown of WDR2 also disrupts the maturation of the centrin arm-associated Golgi, thereby impairing Golgi biogenesis. WDR2 interacts with KIN-G via its N-terminal unknown motifs, the middle domain containing a coiled coil and a PB1 motif, and the C-terminal WD40 domain, and targets KIN-G to its subcellular location. These results uncover a regulatory role for WDR2 in recruiting KIN-G to regulate hook complex and Golgi biogenesis, thereby impacting flagellum inheritance and cell division plane positioning.

Apoptotic protease activating factor 1 (Apaf-1) was traditionally defined as a scaffold protein in mammalian cells for assembling a caspase activation platform known as the 'apoptosome' after its binding to cytochrome c. Although Apaf-1 structurally resembles animal NOD-like receptor (NLR) and plant resistance (R) proteins, whether it is directly involved in innate immunity is still largely unknown. Here, we found that Apaf-1-like molecules from lancelets, fruit flies, mice, and humans have conserved DNA sensing functionality. Mechanistically, mammalian Apaf-1 recruits receptor-interacting protein 2 (RIP2, also known as RIPK2) via its WD40 repeat domain and promotes RIP2 oligomerization to initiate NF-κB-driven inflammation upon cytoplasmic DNA recognition. Furthermore, DNA binding of Apaf-1 determines cell fate by switching the cellular processes between intrinsic stimuli-activated apoptosis and inflammation. These findings suggest that Apaf-1 is an evolutionarily conserved DNA sensor and may serve as a cell fate checkpoint, which determines whether cells initiate inflammation or undergo apoptosis by distinct ligand binding.

Targeting E3 ligases is a challenging area in drug discovery. Despite the human genome encoding for more than 600 E3 ubiquitin ligases, only a handful of E3 ligases have been pharmacologically modulated or exploited for targeted protein degradation (TPD) strategies. The main obstacle for hijacking these E3 ligases is the lack of small-molecule ligands. As research into this field advances, the identification of new small molecules capable of binding to E3 ligases has become an essential pursuit. These ligases not only expand the repertoire of druggable targets but also offer the potential for increased specificity and selectivity in protein degradation. The synergy between academia and industry is key, as it combines academic expertise in fundamental research with the industrial capabilities of translating these findings into novel therapeutics. In this review, we provide an overview of the different strategies employed in academia and industry to the discovery of new E3 ligases ligands, showing them with illustrative cases.

WDR5 is an epigenetic scaffolding protein that has attracted significant interest as an anti-cancer drug target, especially in MLL-rearranged leukemias. The most druggable 'WIN-site' on WDR5, which tethers WDR5 to chromatin, has been successfully targeted with multiple classes of exquisitely potent small-molecule protein-protein interaction inhibitors. Earlier progress has also been made on the development of WDR5 degraders and inhibitors at the 'WBM-site' on the opposite face of WDR5.

Based on an international survey of the patent literature using SciFinder from 2016-2024, herein we provide a comprehensive account of the chemical matter targeting WDR5, with a particular focus on proprietary compounds that are underreported in the existing academic literature. Our survey illuminates challenges for the field to overcome: a broad lack of chemical diversity, confusion about the molecular mechanism of WIN-site inhibitors, a paucity of brain-penetrant scaffolds despite emerging evidence of activity in brain cancers, sparse pharmacokinetic, metabolic, and disposition characterization, and the absence of safety or efficacy data in humans.

It is our opinion that the best-in-class WIN-site inhibitors (from the imidazole class) merit advancement into clinical testing, likely against leukemia, which should provide much-needed clarity about the exciting but unproven potential of WDR5 as a next-generation therapeutic target.

WD repeat domain 12 (WDR12) plays a crucial role in the ribosome biogenesis pathway. However, its biological function in colorectal cancer (CRC) remains poorly understood. Therefore, this study aims to investigate the roles of WDR12 in the occurrence and progression of CRC, as well as its underlying mechanisms.

The expression of WDR12 was assessed through The Cancer Genome Atlas (TCGA) and the Human Protein Atlas (HPA) database. Functional experiments including Celigo assay, MTT assay, and Caspase-3/7 assay were conducted to validate the role of WDR12 in the malignant progression of CRC. Additionally, mRNA chip-sequencing and ingenuity pathway analysis (IPA) were performed to identify the molecular mechanism.

WDR12 expression was significantly upregulated in CRC tissues compared to normal colorectal tissues. Knockdown of WDR12 reduced proliferation and promoted apoptosis of CRC cell lines in vitro and in vivo experiments. Furthermore, WDR12 expression had a significantly inverse association with diseases and functions, including cancer, cell cycle, DNA replication, recombination, cellular growth, proliferation and repair, as revealed by IPA analysis of mRNA chip-sequencing data. Moreover, the activation of cell cycle checkpoint kinases proteins in the cell cycle checkpoint control signaling pathway was enriched in the WDR12 knockdown CRC cell lines. Additionally, downregulation of rac family small GTPase 1 (RAC1) occurred upon WDR12 knockdown, thereby facilitating the proliferation and anti-apoptosis of CRC cells.

Our study demonstrates that the WDR12/RAC1 axis promotes tumor progression in CRC. Therefore, WDR12 may serve as a novel oncogene and a potential target for individualized therapy in CRC. These findings provide an experimental foundation for the clinical development of drugs targeting the WDR12/RAC1 axis.

Ankyrin repeat is a 33-amino acid motif commonly observed in eukaryotes and, to a lesser extent, in prokaryotes and archaea and rarely in viruses. This motif plays a crucial role in regulating various cellular processes like the cell cycle, transcription, cell signaling, and inflammatory responses through interactions between proteins. Poxviruses exhibit a distinctive feature of containing multiple ankyrin repeat proteins within their genomes. All the genera of poxviruses possess these proteins except molluscipox virus, crocodylidpox virus, and red squirrel poxvirus. An intriguing characteristic has generated notable interest in studying the functions of these proteins within poxvirus biology. Within poxviruses, ankyrin repeat proteins exhibit a distinct configuration, featuring ankyrin repeats in the N-terminal region and a cellular F-box homolog in the C-terminal region, which enables interactions with the cellular Skp, Cullin, F-box containing ubiquitin ligase complex. Through the examination of experimental evidences and discussions from current literature, this review elucidates the organization and role of ankyrin repeat proteins in poxviruses. Various research studies have highlighted the significant importance of these proteins in poxviral pathogenesis and, acting as factors that enhance virulence. Consequently, they represent viable targets for developing genetically altered viruses with decreased virulence, thus displaying potential as candidates for vaccines and antiviral therapeutic development contributing to safer and more effective strategies against poxviral infections.

The SnRK1, hexokinase, and TORC1 (TOR, LST8, RAPTOR) are three pivotal kinases at the core of sugar level sensing, significantly impacting plant metabolism and development. We retrieved and analyzed protein sequences of these three kinase pathways from seven sugarcane transcriptome and genome datasets, identifying protein domains, phylogenetic relationships, sequence ancestry, and in silico expression levels. Additionally, we predicted HXK subcellular localization and assessed its enzymatic activity in sugarcane leaves and culms along development in the field. We retrieved 11 TOR, 23 RAPTOR, 55 LST8, 95 SnRK1α, 98 HXK, and 14 HXK-like putative full-length sequences containing all the conserved domains. Most of these transcripts seem to share a common origin with the three ancestral species of sugarcane: Saccharum officinarum, Saccharum spontaneum, and Saccharum barberi. We accessed the expression profile of sequences from one sugarcane transcriptome. We found the highest enzymatic activity of HXK in culms in the first month, which, at this stage, provides carbon (sucrose) and nitrogen (amino acids) for initial plant development. Our approach places novel sugar sensing sequences that work as a guideline for further research into the underlying signaling mechanisms and biotechnology applications in sugarcane.

Flavonoids are secondary metabolites associated with plant seed coat and flower color. These compounds provide health benefits to humans as anti-inflammatory and antioxidant compounds. The expression of the late biosynthetic genes in the flavonoid pathway is controlled by a ternary MBW protein complex consisting of interfacing MYB, beta-helix-loop-helix (bHLH), and WD40 Repeat (WDR) proteins. P, the master regulator gene of the flavonoid expression in common bean (Phaseolus vulgaris L.), was recently determined to encode a bHLH protein. The T and Z genes control the distribution of color in bean seeds and flowers and have historically been considered regulators of the flavonoid gene expression. T and Z candidates were identified using reverse genetics based on genetic mapping, phylogenetic analysis, and mutant analysis. Domain and AlphaFold2 structure analyses determined that T encodes a seven-bladed β-propeller WDR protein, while Z encodes a R2R3 MYB protein. Deletions and SNPs in T and Z mutants, respectively, altered the 3D structure of these proteins. Modeling of the Z MYB/P bHLH/T WDR MBW complex identified interfacing sequence domains and motifs in all three genes that are conserved in dicots. One Z MYB motif is a possible beta-molecular recognition feature (β-MoRF) that only appears in a structured state when Z MYB is modeled as a component of a MBW complex. Complexes containing mutant T and Z proteins changed the interaction of members of the complex in ways that would alter their role in regulating the expression of genes in the flavonoid pathway.

The WD40 domain, one of the most abundant in eukaryotic genomes, is widely involved in plant growth and development, secondary metabolic biosynthesis, and mediating responses to biotic and abiotic stresses. WD40 repeat (WD40) protein has been systematically studied in several model plants but has not been reported in the Capsicum annuum (pepper) genome.

Herein, 269, 237, and 257 CaWD40 genes were identified in the Zunla, CM334, and Zhangshugang genomes, respectively. CaWD40 sequences from the Zunla genome were selected for subsequent analysis, including chromosomal localization, phylogenetic relationships, sequence characteristics, motif compositions, and expression profiling. CaWD40 proteins were unevenly distributed on 12 chromosomes, encompassing 19 tandem duplicate gene pairs. The 269 CaWD40s were divided into six main branches (A to F) with 17 different types of domain distribution. The CaWD40 gene family exhibited diverse expression patterns, and several genes were specifically expressed in flowers and seeds. Yeast two-hybrid (Y2H) and dual-luciferase assay indicated that CaWD40-91 could interact with CaAN1 and CaDYT1, suggesting its involvement in anthocyanin biosynthesis and male sterility in pepper.

In summary, we systematically characterized the phylogeny, classification, structure, and expression of the CaWD40 gene family in pepper. Our findings provide a valuable foundation for further functional investigations on WD40 genes in pepper.

WD repeat domain 43 (WDR43) is a protein component that encodes WD-repeats and is involved in ribosome biogenesis. However, little is known about the role of WDR43 in cancer prognosis and immune modulation.

In this study, we analyzed the expression and prognostic significance of WDR43 in pan-cancer using the Cancer Genome Atlas, the Genotype-Tissue Expression, and the Human Protein Atlas. We also examined the differential expression of WDR43 in liver hepatocellular carcinoma (LIHC) and adjacent tissues of 48 patients using immunohistochemistry. Additionally, we investigated the correlation between WDR43 and clinical characteristics, gene alterations, tumor mutation burden, microsatellite instability, mismatch repair, tumor microenvironment, immune infiltrating cells, and immune-related genes using bioinformatics methods. Gene set enrichment analysis was conducted, and potential biological mechanisms were identified.

Immunohistochemistry staining showed that WDR43 was overexpressed in LIHC among 48 patients. Upregulation of WDR43 was associated with unfavorable prognosis, including overall survival in various types of cancer such as LIHC, uterine corpus endometrial cancer, head and neck squamous cell carcinoma, and pancreatic adenocarcinoma. Differential expression of WDR43 was significantly correlated with microsatellite instability, mismatch repair, and immune cell infiltration. Gene ontology annotation analysis revealed that these genes were significantly enriched in immune-related functions, including immune response, immune regulation, and signaling pathways.

We conducted a thorough investigation of the clinical features, phases of tumor development, immune infiltration, gene mutation, and functional enrichment analysis of WDR43 in various types of cancer. This research offers valuable insight into the significance and function of WDR43 in clinical therapy.

Beta-Propeller Protein-Associated Neurodegeneration (BPAN) is one of the commonest forms of Neurodegeneration with Brain Iron Accumulation, caused by mutations in the gene encoding the autophagy-related protein, WDR45. The mechanisms linking autophagy, iron overload and neurodegeneration in BPAN are poorly understood and, as a result, there are currently no disease-modifying treatments for this progressive disorder. We have developed a patient-derived, induced pluripotent stem cell (iPSC)-based midbrain dopaminergic neuronal cell model of BPAN (3 patient, 2 age-matched controls and 2 isogenic control lines) which shows defective autophagy and aberrant gene expression in key neurodegenerative, neurodevelopmental and collagen pathways. A high content imaging-based medium-throughput blinded drug screen using the FDA-approved Prestwick library identified 5 cardiac glycosides that both corrected disease-related defective autophagosome formation and restored BPAN-specific gene expression profiles. Our findings have clear translational potential and emphasise the utility of iPSC-based modelling in elucidating disease pathophysiology and identifying targeted therapeutics for early-onset monogenic disorders.

The Critical Assessment of Computational Hit-Finding Experiments (CACHE) Challenge series is focused on identifying small molecule inhibitors of protein targets using computational methods. Each challenge contains two phases, hit-finding and follow-up optimization, each of which is followed by experimental validation of the computational predictions. For the CACHE Challenge #1, the Leucine-Rich Repeat Kinase 2 (LRRK2) WD40 Repeat (WDR) domain was selected as the target for in silico hit-finding and optimization. Mutations in LRRK2 are the most common genetic cause of the familial form of Parkinson's disease. The LRRK2 WDR domain is an understudied drug target with no known molecular inhibitors. Herein we detail the first phase of our winning submission to the CACHE Challenge #1. We developed a framework for the high-throughput structure-based virtual screening of a chemically diverse small molecule space. Hit identification was performed using the large-scale Deep Docking (DD) protocol followed by absolute binding free energy (ABFE) simulations. ABFEs were computed using an automated molecular dynamics (MD)-based thermodynamic integration (TI) approach. 4.1 billion ligands from Enamine REAL were screened with DD followed by ABFEs computed by MD TI for 793 ligands. 76 ligands were prioritized for experimental validation, with 59 compounds successfully synthesized and 5 compounds identified as hits, yielding a 8.5% hit rate. Our results demonstrate the efficacy of the combined DD and ABFE approaches for hit identification for a target with no previously known hits. This approach is widely applicable for the efficient screening of ultra-large chemical libraries as well as rigorous protein-ligand binding affinity estimation leveraging modern computational resources.

Chung-Jansen syndrome is a neurodevelopmental disorder characterized by intellectual disability, behavioral problems, obesity and dysmorphic features. It is caused by pathogenic variants in the PHIP gene that encodes for the Pleckstrin homology domain-interacting protein, which is part of an epigenetic modifier protein complex. Therefore, we hypothesized that PHIP haploinsufficiency may impact genome-wide DNA methylation (DNAm). We assessed the DNAm profiles of affected individuals with pathogenic and likely pathogenic PHIP variants with Infinium Methylation EPIC arrays and report a specific and sensitive DNAm episignature biomarker for Chung-Jansen syndrome. In addition, we observed similarities between the methylation profile of Chung-Jansen syndrome and that of functionally related and clinically partially overlapping genetic disorders, White-Kernohan syndrome (caused by variants in DDB1 gene) and Börjeson-Forssman-Lehmann syndrome (caused by variants in PHF6 gene). Based on these observations we also proceeded to develop a common episignature biomarker for these disorders. These newly defined episignatures can be used as part of a multiclass episignature classifier for screening of affected individuals with rare disorders and interpretation of genetic variants of unknown clinical significance, and provide further insights into the common molecular pathophysiology of the clinically-related Chung-Jansen, Börjeson-Forssman-Lehmann and White-Kernohan syndromes.

The WD40 gene family, prevalent in eukaryotes, assumes diverse roles in cellular processes. Spartina alterniflora, a halophyte with exceptional salt tolerance, flood tolerance, reproduction, and diffusion ability, offers great potential for industrial applications and crop breeding analysis. The exploration of growth and development-related genes in this species offers immense potential for enhancing crop yield and environmental adaptability, particularly in industrialized plantations. However, the understanding of their role in regulating plant growth and development remains limited.

In this study, we conducted a comprehensive analysis of WD40 genes in S. alterniflora at the whole-genome level, delving into their characteristics such as physicochemical properties, phylogenetic relationships, gene architecture, and expression patterns. Additionally, we cloned the TTG1 gene, a gene in plant growth and development across diverse species.

We identified a total of 582 WD40 proteins in the S. alterniflora genome, exhibiting an uneven distribution across chromosomes. Through phylogenetic analysis, we categorized the 582 SaWD40 proteins into 12 distinct clades. Examining the duplication patterns of SaWD40 genes, we observed a predominant role of segmental duplication in their expansion. A substantial proportion of SaWD40 gene duplication pairs underwent purifying selection through evolution. To explore the functional aspects, we selected SaTTG1, a homolog of Arabidopsis TTG1, for overexpression in Arabidopsis. Subcellular localization analysis revealed that the SaTTG1 protein localized in the nucleus and plasma membrane, exhibiting transcriptional activation in yeast cells. The overexpression of SaTTG1 in Arabidopsis resulted in early flowering and increased seed size.

These outcomes significantly contribute to our understanding of WD40 gene functions in halophyte species. The findings not only serve as a valuable foundation for further investigations into WD40 genes in halophyte but also offer insights into the molecular mechanisms governing plant development, offering potential avenues in molecular breeding.

Targeted protein degradation (TPD) has recently emerged as an exciting new drug modality. However, the strategy of developing small molecule-based protein degraders has evolved over the past two decades and has now established molecular tags that are already in clinical use, as well as chimeric molecules, PROteolysis TArgeting Chimeras (PROTACs), based mainly on ligand systems developed for the two E3 ligases CRBN and VHL. The large size of the human E3 ligase family suggests that PROTACs can be developed by targeting a large diversity of E3 ligases, some of which have restricted expression patterns with the potential to design disease- or tissue-specific degraders. Indeed, many new E3 ligands have been published recently, confirming the druggability of E3 ligases. This review summarises recent data on E3 ligases and highlights the challenges in developing these molecules into efficient PROTACs rivalling the established degrader systems.

Diversity-generating retroelements (DGRs) are used by bacteria, archaea, and viruses as a targeted mutagenesis tool. Through error-prone reverse transcription, DGRs introduce random mutations at specific genomic loci, enabling rapid evolution of these targeted genes. However, the function and benefits of DGR-diversified proteins in cellular hosts remain elusive. We find that 82% of DGRs from one of the major monophyletic lineages of DGR reverse transcriptases are encoded by multicellular bacteria, which often have two or more DGR loci in their genomes. Using the multicellular purple sulfur bacterium Thiohalocapsa sp. PB-PSB1 as an example, we characterized nine distinct DGR loci capable of generating 10282 different combinations of target proteins. With environmental metagenomes from individual Thiohalocapsa aggregates, we show that most of PB-PSB1's DGR target genes are diversified across its biogeographic range, with spatial heterogeneity in the diversity of each locus. In Thiohalocapsa PB-PSB1 and other bacteria hosting this lineage of cellular DGRs, the diversified target genes are associated with NACHT-domain anti-phage defenses and putative ternary conflict systems previously shown to be enriched in multicellular bacteria. We propose that these DGR-diversified targets act as antigen sensors that confer a form of adaptive immunity to their multicellular consortia, though this remains to be experimentally tested. These findings could have implications for understanding the evolution of multicellularity, as the NACHT-domain anti-phage systems and ternary systems share both domain homology and conceptual similarities with the innate immune and programmed cell death pathways of plants and metazoans.

G protein coupled receptors (GPCRs) are seven-transmembrane domain proteins that modulate cellular processes in response to external stimuli. These receptors represent the largest family of membrane proteins, and in mammals, their signaling regulates important physiological functions, such as vision, taste, and olfaction. Many organisms, including yeast, slime molds, and viruses encode GPCRs. Cytomegaloviruses (CMVs) are large, betaherpesviruses, that encode viral GPCRs (vGPCRs). Human CMV (HCMV) encodes four vGPCRs, including UL33, UL78, US27, and US28. Each of these vGPCRs, as well as their rodent and primate orthologues, have been investigated for their contributions to viral infection and disease. Herein, we discuss how the CMV vGPCRs function during lytic and latent infection, as well as our understanding of how they impact viral pathogenesis.

Dynein, an ancient microtubule-based motor protein, performs diverse cellular functions in nearly all eukaryotic cells, with the exception of land plants. It has evolved into three subfamilies-cytoplasmic dynein-1, cytoplasmic dynein-2, and axonemal dyneins-each differentiated by their cellular functions. These megadalton complexes consist of multiple subunits, with the heavy chain being the largest subunit that generates motion and force along microtubules by converting the chemical energy of ATP hydrolysis into mechanical work. Beyond this catalytic core, the functionality of dynein is significantly enhanced by numerous non-catalytic subunits. These subunits are integral to the complex, contributing to its stability, regulating its enzymatic activities, targeting it to specific cellular locations, and mediating its interactions with other cofactors. The diversity of non-catalytic subunits expands dynein's cellular roles, enabling it to perform critical tasks despite the conservation of its heavy chains. In this review, we discuss recent findings and insights regarding these non-catalytic subunits.

Banded leaf and sheath blight (BLSB) in maize is a soil-borne fungal disease caused by Rhizoctonia solani Kühn, resulting in significant yield losses. Investigating the genes responsible for regulating resistance to BLSB is crucial for yield enhancement. In this study, a multiparent maize population was developed, comprising two recombinant inbred line (RIL) populations totaling 442 F8RILs. The populations were generated by crossing two tropical inbred lines, CML444 and NK40-1, known for their BLSB resistance, as female parents, with the high-yielding but BLSB-susceptible inbred line Ye107 serving as the common male parent. Subsequently, we utilized 562,212 high-quality single nucleotide polymorphisms (SNPs) generated through genotyping-by-sequencing (GBS) for a comprehensive genome-wide association study (GWAS) aimed at identifying genes responsible for BLSB resistance. The objectives of this study were to (1) identify SNPs associated with BLSB resistance through genome-wide association analyses, (2) explore candidate genes regulating BLSB resistance in maize, and (3) investigate pathways involved in BLSB resistance and discover key candidate genes through Gene Ontology (GO) analysis. The GWAS analysis revealed nineteen SNPs significantly associated with BLSB that were consistently identified across four environments in the GWAS, with phenotypic variation explained (PVE) ranging from 2.48% to 11.71%. Screening a 40 kb region upstream and downstream of the significant SNPs revealed several potential candidate genes. By integrating information from maize GDB and the NCBI, we identified five novel candidate genes, namely, Zm00001d009723, Zm00001d009975, Zm00001d009566, Zm00001d009567, located on chromosome 8, and Zm00001d026376, on chromosome 10, related to BLSB resistance. These candidate genes exhibit association with various aspects, including maize cell membrane proteins and cell immune proteins, as well as connections to cell metabolism, transport, transcriptional regulation, and structural proteins. These proteins and biochemical processes play crucial roles in maize defense against BLSB. When Rhizoctonia solani invades maize plants, it induces the expression of genes encoding specific proteins and regulates corresponding metabolic pathways to thwart the invasion of this fungus. The present study significantly contributes to our understanding of the genetic basis of BLSB resistance in maize, offering valuable insights into novel candidate genes that could be instrumental in future breeding efforts to develop maize varieties with enhanced BLSB resistance.

Fertilization is a complex process that depends on the fusion of the cell membrane of sperm with that of oocyte, and it involves sperm-oocyte recognition, binding, and fusion, which are mediated by multiple proteins. Among those proteins, IZUMO1 and its receptor JUNO have been identified as essential factors for sperm-oocyte recognition and fusion. However, the interaction between IZUMO1 and JUNO alone does not lead to cell membrane fusion, suggesting the involvement of additional proteins in sperm-oocyte membrane fusion. In this study, we have discovered that a protein called WDR54, which consists of WD-repeat modules, is located on the cell membrane of sperm, as well as on the cell membrane and in the cytoplasm of the oocyte. We have found that WDR54 is involved in sperm-oocyte fertilization. When sperm and oocyte were treated with anti-WDR54 ascites, the in vitro fertilization (IVF) rate significantly decreased. Furthermore, our research has shown that WDR54 interacts with both IZUMO1 and JUNO, and it colocalizes with IZUMO1 on the surface of the sperm head and with JUNO on the oocyte surface. Through structural analysis of the putative complexes of WDR54-IZUMO1 and WDR54-JUNO, we infer that these three proteins could form a complex of JUNO-WDR54-IZUMO1-JUNO (referred to as the "JWIJ complex") on the oocyte surface. Our findings suggest that WDR54 is an important factor involved in sperm-oocyte adhesion and fusion. This discovery provides new insight into the mechanisms of mammalian sperm-oocyte adhesion and fusion.

Unique patterns of inheritance and selection on Y chromosomes have led to the evolution of specialized gene functions. We report CRISPR mutants in Drosophila of the Y-linked gene, WDY, which is required for male fertility. We demonstrate that the sperm tails of WDY mutants beat approximately half as fast as those of wild-type and that mutant sperm do not propel themselves within the male ejaculatory duct or female reproductive tract. Therefore, although mature sperm are produced by WDY mutant males, and are transferred to females, those sperm fail to enter the female sperm storage organs. We report genotype-dependent and regional differences in sperm motility that appear to break the correlation between sperm tail beating and propulsion. Furthermore, we identify a significant change in hydrophobicity at a residue at a putative calcium-binding site in WDY orthologs at the split between the melanogaster and obscura species groups, when WDY first became Y-linked. This suggests that a major functional change in WDY coincided with its appearance on the Y chromosome. Finally, we show that mutants for another Y-linked gene, PRY, also show a sperm storage defect that may explain their subfertility. Overall, we provide direct evidence for the long-held presumption that protein-coding genes on the Drosophila Y regulate sperm motility.

The chromatin-associated protein WDR5 (WD repeat domain 5) is an essential cofactor for MYC and a conserved regulator of ribosome protein gene transcription. It is also a high-profile target for anti-cancer drug discovery, with proposed utility against both solid and hematological malignancies. We have previously discovered potent dihydroisoquinolinone-based WDR5 WIN-site inhibitors with demonstrated efficacy and safety in animal models. In this study, we sought to optimize the bicyclic core to discover a novel series of WDR5 WIN-site inhibitors with improved potency and physicochemical properties. We identified the 3,4-dihydrobenzo[f][1,4]oxazepin-5(2H)-one core as an alternative scaffold for potent WDR5 inhibitors. Additionally, we used X-ray structural analysis to design partially saturated bicyclic P7 units. These benzoxazepinone-based inhibitors exhibited increased cellular potency and selectivity and favorable physicochemical properties compared to our best-in-class dihydroisoquinolinone-based counterparts. This study opens avenues to discover more advanced WDR5 WIN-site inhibitors and supports their development as novel anti-cancer therapeutics.

Alcohol consumption has been linked to numerous negative health outcomes although it has some beneficial effects on moderate dosages, the most severe of which being alcohol-induced hepatitis. The number of people dying from this liver illness has been shown to climb steadily over time, and its prevalence has been increasing. Researchers have found that alcohol consumption primarily affects the brain, leading to a wide range of neurological and psychological diseases. High-alcohol-consumption addicts not only experienced seizures, but also ataxia, aggression, social anxiety, and variceal hemorrhage that ultimately resulted in death, ascites, and schizophrenia. Drugs treating this liver condition are limited and can cause serious side effects like depression. Serine-threonine kinases, cAMP protein kinases, protein kinase C, ERK, RACK 1, Homer 2, and more have all been observed to have their signaling pathways disrupted by alcohol, and alcohol has also been linked to epigenetic changes. In addition, alcohol consumption induces dysbiosis by changing the composition of the microbiome found in the gastrointestinal tract. Although more studies are needed, those that have been done suggest that probiotics aid in keeping the various microbiota concentrations stable. It has been argued that reducing one's alcohol intake may seem less harmful because excessive drinking is a lifestyle disorder.

The fungal Gβ-like protein has been reported to be involved in a variety of biological processes, such as mycelial growth, differentiation, conidiation, stress responses and infection. However, molecular mechanisms of the Gβ-like protein in regulating fungal development and pathogenicity are largely unknown. Here, we show that the Gβ-like protein gene Bcgbl1 in the gray mold fungus Botrytis cinerea plays a pivotal role in development and pathogenicity by regulating the mitogen-activated protein (MAP) kinases signaling pathways. The Bcgbl1 deletion mutants were defective in mycelial growth, sclerotial formation, conidiation, macroconidial morphogenesis, plant adhesion, and formation of infection cushions and appressorium-like structures, resulting in a complete loss of pathogenicity. Bcgbl1 interacted with BcSte50, the adapter protein of the cascade of MAP kinase (MAPK). Bcgbl1 mutants had reduced phosphorylation levels of two MAPKs, namely Bmp1 and Bmp3, thereby reducing infection. However, deletion of Bcgbl1 did not affect the intracellular cAMP level, and exogenous cAMP could not restore the defects. Moreover, Bcgbl1 mutants exhibited defects in cell wall integrity and oxidative stress tolerance. Transcriptional profiling revealed that Bcgbl1 plays a global role in regulation of gene expression upon hydrophobic surface induction. We further uncovered that three target genes encoding the hydrophobic surface binding proteins (HsbAs) contributed to the adhesion and virulence of B. cinerea. Overall, these findings suggest that Bcgbl1 had multiple functions and provided new insights for deciphering the Bcgbl1-mediated network for regulating development and pathogenicity of B. cinerea.

As a key component of carbon source metabolism in fungi, CreC WD40 repeat protein is regulated by carbon catabolite repression (CCR). However, the understanding of the functions of CreC in entomopathogenic fungi is currently limited. Here, CreC in Metarhizium robertsii (MrCreC) was identified, and its roles in fungal development, conidiation, environmental stress response, and insecticidal virulence were explored. MrCreC is localized to cytoplasm, and MrCreC deletion affects fungal growth on various nutrients. Compared to the wild type, the sporulation of ΔMrCreC strain was significantly decreased by 60.3%. Further qPCR analysis found that deletion of MrCreC resulted in repression of sporulation-related genes such as AbaA, FlbA, Flbc, MedA, FlbD, FluG, and wetA. In addition, MrCreC loss did not alter heat stress tolerance but resulted in enhanced tolerance to UV-B. Interestingly, bioassays showed that the virulence following exposures to topical applications or injection of conidial suspensions of both infection and injection was impaired compared with that of the wild type. Further analysis showed that the adhesion and cuticle penetration genes in ΔMrCreC was down-regulated during infection, and the appressorial formation rate was significantly reduced. A deletion of MrCreC significantly also reduced immune escape and nutrient utilization genes in insect hemocoel. In conclusion, MrCreC is involved in the growth, development and virulence of M. robertsii. These findings advance our understanding of the function of CCR pathway-related genes.

The WD40 superfamily is widely found in eukaryotes and has essential subunits that serve as scaffolds for protein complexes. WD40 proteins play important regulatory roles in plant development and physiological processes, such as transcription regulation and signal transduction; it is also involved in anthocyanin biosynthesis. In rice, only OsTTG1 was found to be associated with anthocyanin biosynthesis, and evolutionary analysis of the WD40 gene family in multiple species is less studied. Here, a genome-wide analysis of the subfamily belonging to WD40-TTG1 was performed in nine AA genome species: Oryza sativa ssp. japonica, Oryza sativa ssp. indica, Oryza rufipogon, Oryza glaberrima, Oryza meridionalis, Oryza barthii, Oryza glumaepatula, Oryza nivara, and Oryza longistaminata. In this study, 383 WD40 genes in the Oryza genus were identified, and they were classified into four groups by phylogenetic analysis, with most members in group C and group D. They were found to be unevenly distributed across 12 chromosomes. A total of 39 collinear gene pairs were identified in the Oryza genus, and all were segmental duplications. WD40s had similar expansion patterns in the Oryza genus. Ka/Ks analyses indicated that they had undergone mainly purifying selection during evolution. Furthermore, WD40s in the Oryza genus have similar evolutionary patterns, so Oryza sativa ssp. indica was used as a model species for further analysis. The cis-acting elements analysis showed that many genes were related to jasmonic acid and light response. Among them, OsiWD40-26/37/42 contained elements of flavonoid synthesis, and OsiWD40-15 had MYB binding sites, indicating that they might be related to anthocyanin synthesis. The expression profile analysis at different stages revealed that most OsiWD40s were expressed in leaves, roots, and panicles. The expression of OsiWD40s was further analyzed by qRT-PCR in 9311 (indica) under various hormone treatments and abiotic stresses. OsiWD40-24 was found to be responsive to both phytohormones and abiotic stresses, suggesting that it might play an important role in plant stress resistance. And many OsiWD40s might be more involved in cold stress tolerance. These findings contribute to a better understanding of the evolution of the WD40 subfamily. The analyzed candidate genes can be used for the exploration of practical applications in rice, such as cultivar culture for colored rice, stress tolerance varieties, and morphological marker development.

Rice (Oryza sativa L.) is a cold-sensitive species that often faces cold stress, which adversely affects yield productivity and quality. However, the genetic basis for low-temperature adaptation in rice remains unclear. Here, we demonstrate that 2 functional polymorphisms in O. sativa SEC13 Homolog 1 (OsSEH1), encoding a WD40-repeat nucleoporin, between the 2 subspecies O. sativa japonica and O. sativa indica rice, may have facilitated cold adaptation in japonica rice. We show that OsSEH1 of the japonica variety expressed in OsSEH1MSD plants (transgenic line overexpressing the OsSEH1 allele from Mangshuidao [MSD], cold-tolerant landrace) has a higher affinity for O. sativa metallothionein 2b (OsMT2b) than that of OsSEH1 of indica. This high affinity of OsSEH1MSD for OsMT2b results in inhibition of OsMT2b degradation, with decreased accumulation of reactive oxygen species and increased cold tolerance. Transcriptome analysis indicates that OsSEH1 positively regulates the expression of the genes encoding dehydration-responsive element-binding transcription factors, i.e. OsDREB1 genes, and induces the expression of multiple cold-regulated genes to enhance cold tolerance. Our findings highlight a breeding resource for improving cold tolerance in rice.

Lymph node (LN) metastasis is one of the key prognostic factors in bladder cancer, but its underlying mechanisms remain unclear. Here, we found that elevated expression of WD repeat domain 4 (WDR4) in bladder cancer correlated with worse prognosis. WDR4 can promote the LN metastasis and proliferation of bladder cancer cells. Mechanistic studies showed that WDR4 can promote the nuclear localization of DEAD-box helicase 20 (DDX20) and act as an adaptor to bind DDX20 and Early growth response 1 (Egr1), thereby inhibiting Egr1-promoted transcriptional expression of arrestin beta 2 (ARRB2) and ultimately contributing to the progression of bladder cancer. Immunohistochemical analysis confirmed that WDR4 expression is also an independent predictor of LN metastasis in bladder cancer. Our results reveal a novel mechanism of LN metastasis and progression in bladder cancer and identify WDR4 as a potential therapeutic target for metastatic bladder cancer.

V-ATPase, which comprises 13-14 subunits, is essential for pH homeostasis in all eukaryotes, but its proper function requires a regulator to assemble its subunits. While RAVE (regulator of H+-ATPase of vacuolar and endosomal membranes) and Raboconnectin-3 complexes assemble V-ATPase subunits in Saccharomyces cerevisiae and humans, respectively, the function of the RAVE complex in fungal pathogens remains largely unknown. In this study, we identified two RAVE complex components, Rav1 and Wdr1, in the fungal meningitis pathogen Cryptococcus neoformans, and analyzed their roles. Rav1 and Wdr1 are orthologous to yeast RAVE and human Rabconnectin-3 counterparts, respectively, forming the hybrid RAVE (hRAVE) complex. Deletion of RAV1 caused severe defects in growth, cell cycle control, morphogenesis, sexual development, stress responses, and virulence factor production, while the deletion of WDR1 resulted in similar but modest changes, suggesting that Rav1 and Wdr1 play central and accessary roles, respectively. Proteomics analysis confirmed that Wdr1 was one of the Rav1-interacting proteins. Although the hRAVE complex generally has V-ATPase-dependent functions, it also has some V-ATPase-independent roles, suggesting a unique role beyond conventional intracellular pH regulation in C. neoformans. The hRAVE complex played a critical role in the pathogenicity of C. neoformans, and RAV1 deletion attenuated virulence and impaired blood-brain barrier crossing ability. This study provides comprehensive insights into the pathobiological roles of the fungal RAVE complex and suggests a novel therapeutic strategy for controlling cryptococcosis.

Rice is a crucial food crop worldwide, but it is highly susceptible to Hirschmanniella mucronata, a migratory parasitic nematode. No rice variety has been identified that could resist H. mucronata infection. Therefore, it is very important to study the interaction between rice and H. mucronata to breed resistant rice varieties. Here, we demonstrated that protein OsWD40-193 interacted with the extension factor OseEF1A1 and both were negative regulators inhibiting rice resistance to H. mucronata infection. Overexpression of either OsWD40-193 or OseEF1A1 led to enhance susceptibility to H. mucronata, whereas the absence of OsWD40-193 or OseEF1A1 led to resistance. Further transcriptomic analysis showed that OseEF1A1 deletion altered the expression of genes association with salicylic acid, jasmonic acid and abolic acid signaling pathways and increased the accumulation of secondary metabolites to enhance resistance in rice. Our study showed that H. mucronata infection affected the expression of negative regulators in rice and inhibited rice resistance, which was conducive to the infection of nematode. Together, our data showed that H. mucronata affected the expression of negative regulators to facilitate its infection and provided potential target genes to engineering resistance germplasm via gene editing of the negative regulators.

WD40 transcription factors (TFs) constitute a large gene family in eukaryotes, playing diverse roles in cellular processes. However, their functions in the major ornamental plant, Rhododendron simsii, remain poorly understood. In this study, we identified 258 WD40 proteins in the R. simsii genome, which exhibited an uneven distribution across chromosomes. Based on domain compositions and phylogenetic analysis, we classified these 258 RsWD40 proteins into 42 subfamilies and 47 clusters. Comparative genomic analysis suggested that the expansion of the WD40 gene family predates the divergence of green algae and higher plants, indicating an ancient origin. Furthermore, by analyzing the duplication patterns of RsWD40 genes, we found that transposed duplication played a major role in their expansion. Notably, the majority of RsWD40 gene duplication pairs underwent purifying selection during evolution. Synteny analysis identified significant orthologous gene pairs between R. simsii and Arabidopsis thaliana, Oryza sativa, Vitis vinifera, and Malus domestica. We also investigated potential candidate genes involved in anthocyanin biosynthesis during different flower development stages in R. simsii using RNA-seq data. Specifically, we identified 10 candidate genes during the bud stage and 7 candidate genes during the full bloom stage. GO enrichment analysis of these candidate genes revealed the potential involvement of the ubiquitination process in anthocyanin biosynthesis. Overall, our findings provide a valuable foundation for further investigation and functional analysis of WD40 genes, as well as research on the molecular mechanisms underlying anthocyanin biosynthesis in Rhododendron species.

Tilapia is one of the most essential farmed fishes in the world. It is a tropical and subtropical freshwater fish well adapted to warm water but sensitive to cold weather. Extreme cold weather could cause severe stress and mass mortalities in tilapia. The present study was carried out to investigate the effects of cold stress on the up-regulation of antifreeze protein (AFP) genes in Nile tilapia (Oreochromis niloticus). Two treatment groups of fish were investigated (5 replicates of 15 fish for each group in fibreglass tanks/70 L each): 1) a control group; the fish were acclimated to lab conditions for two weeks and the water temperature was maintained at 25 °C during the whole experimental period with feeding on a commercial diet (30% crude protein). 2) Cold stress group; the same conditions as the control group except for the temperature. Initially, the temperature was decreased by one degree every 12 h. The fish started showing death symptoms when the water temperature reached 6-8 °C. In this stage the tissue (muscle) samples were taken from both groups. The immune response of fish exposed to cold stress was detected and characterized using Differential Display-PCR (DD-PCR).

The results indicated that nine different up-regulation genes were detected in the cold-stressed fish compared to the control group. These genes are Integrin-alpha-2 (ITGA-2), Gap junction gamma-1 protein-like (GJC1), WD repeat-containing protein 59 isoform X2 (WDRP59), NUAK family SNF1-like kinase, G-protein coupled receptor-176 (GPR-176), Actin cytoskeleton-regulatory complex protein pan1-like (PAN-1), Whirlin protein (WHRN), Suppressor of tumorigenicity 7 protein isoform X2 (ST7P) and ATP-binding cassette sub-family A member 1-like isoform X2 (ABCA1). The antifreeze gene type-II amplification using a specific PCR product of 600 bp, followed by cloning and sequencing analysis revealed that the identified gene is antifreeze type-II, with similarity ranging from 70 to 95%. The in-vitro transcribed gene induced an antifreeze protein with a molecular size of 22 kDa. The antifreeze gene, ITGA-2 and the WD repeat protein belong to the lectin family (sugar-protein).

In conclusion, under cold stress, Nile tilapia express many defence genes, an antifreeze gene consisting of one open reading frame of approximately 0.6 kbp.