Basic Study
Copyright ©The Author(s) 2024. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Diabetes. Feb 15, 2024; 15(2): 275-286
Published online Feb 15, 2024. doi: 10.4239/wjd.v15.i2.275
Assessment of pathogenicity and functional characterization of APPL1 gene mutations in diabetic patients
Ping Shi, Yang Tian, Feng Xu, Lu-Na Liu, Wan-Hong Wu, Ying-Zhou Shi, An-Qi Dai, Hang-Yu Fang, Kun-Xia Li, Chao Xu
Ping Shi, Yang Tian, Feng Xu, Lu-Na Liu, Wan-Hong Wu, Ying-Zhou Shi, An-Qi Dai, Hang-Yu Fang, Chao Xu, Department of Endocrinology, Shandong Provincial Hospital Affiliated to Shandong First Medical University, Jinan 250021, Shandong Province, China
Kun-Xia Li, Department of Pediatric, Yantai Yuhuangding Hospital Affiliated to Qingdao University, Yantai 264099, Shandong Province, China
Co-corresponding authors: Kun-Xia Li and Chao Xu.
Author contributions: Li KX and Xu C are the co-corresponding authors of this manuscript; Xu C acted as a guarantor for the research design and manuscript revision; Li KX designed the theme of the article; Shi P carried out the experiments and wrote the manuscript; Tian Y contributed to the experiment implementation and data collection; Xu F edited and reviewed the content of the manuscript; Liu LN and Wu WH contributed to the experiment implementation and data analysis; Shi YZ, Dai AQ, and Fang HY performed bioinformation analysis and graph drawing; and all authors read and approved the final manuscript.
Supported by the National Natural Science Foundation, No. 81974124; and Taishan Scholar Project, No. tsqn20161071.
Institutional review board statement: The study was reviewed and approved by the Ethics Committee of Shandong Provincial Hospital.
Conflict-of-interest statement: All the authors report having no relevant conflicts of interest for this article.
Data sharing statement: No additional data are available.
Open-Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: https://creativecommons.org/Licenses/by-nc/4.0/
Corresponding author: Chao Xu, MD, Doctor, Department of Endocrinology, Shandong Provincial Hospital Affiliated to Shandong First Medical University, No. 324 Jingwu Weiqi Road, Jinan 250021, Shandong Province, China. doctorxuchao@163.com
Received: October 2, 2023
Peer-review started: October 2, 2023
First decision: November 9, 2023
Revised: November 22, 2023
Accepted: January 9, 2024
Article in press: January 9, 2024
Published online: February 15, 2024
Abstract
BACKGROUND

Adaptor protein, phosphotyrosine interacting with PH domain and leucine zipper 1 (APPL1) plays a crucial role in regulating insulin signaling and glucose metabolism. Mutations in the APPL1 gene have been associated with the development of maturity-onset diabetes of the young type 14 (MODY14). Currently, only two mutations [c.1655T>A (p.Leu552*) and c.281G>A p.(Asp94Asn)] have been identified in association with this disease. Given the limited understanding of MODY14, it is imperative to identify additional cases and carry out comprehensive research on MODY14 and APPL1 mutations.

AIM

To assess the pathogenicity of APPL1 gene mutations in diabetic patients and to characterize the functional role of the APPL1 domain.

METHODS

Patients exhibiting clinical signs and a medical history suggestive of MODY were screened for the study. Whole exome sequencing was performed on the patients as well as their family members. The pathogenicity of the identified APPL1 variants was predicted on the basis of bioinformatics analysis. In addition, the pathogenicity of the novel APPL1 variant was preliminarily evaluated through in vitro functional experiments. Finally, the impact of these variants on APPL1 protein expression and the insulin pathway were assessed, and the potential mechanism underlying the interaction between the APPL1 protein and the insulin receptor was further explored.

RESULTS

A total of five novel mutations were identified, including four missense mutations (Asp632Tyr, Arg633His, Arg532Gln, and Ile642Met) and one intronic mutation (1153-16A>T). Pathogenicity prediction analysis revealed that the Arg532Gln was pathogenic across all predictions. The Asp632Tyr and Arg633His variants also had pathogenicity based on MutationTaster. In addition, multiple alignment of amino acid sequences showed that the Arg532Gln, Asp632Tyr, and Arg633His variants were conserved across different species. Moreover, in in vitro functional experiments, both the c.1894G>T (at Asp632Tyr) and c.1595G>A (at Arg532Gln) mutations were found to downregulate the expression of APPL1 on both protein and mRNA levels, indicating their pathogenic nature. Therefore, based on the patient’s clinical and family history, combined with the results from bioinformatics analysis and functional experiment, the c.1894G>T (at Asp632Tyr) and c.1595G>A (at Arg532Gln) mutations were classified as pathogenic mutations. Importantly, all these mutations were located within the phosphotyrosine-binding domain of APPL1, which plays a critical role in the insulin sensitization effect.

CONCLUSION

This study provided new insights into the pathogenicity of APPL1 gene mutations in diabetes and revealed a potential target for the diagnosis and treatment of the disease.

Keywords: Adaptor protein, phosphotyrosine interacting with PH domain and leucine zipper 1, Maturity-onset diabetes of the young, Bioinformatics analysis, Gene mutation, Domain

Core Tip: We identified five new mutations in the adaptor protein, phosphotyrosine interacting with PH domain and leucine zipper 1 (APPL1) gene, a critical regulator of insulin signaling and glucose metabolism, in maturity-onset diabetes of the young type 14 patients. We conducted bioinformatics and functional experiments and showed that two mutations were pathogenic, resulting in reduced expression of the APPL1 protein and mRNA. All mutations were in the phosphotyrosine-binding domain of APPL1, which is important for its insulin-sensitizing effect. This study gave new insights into the pathogenicity of APPL1 mutations in diabetes and revealed potential targets for improved diagnosis and treatment strategies.