890-P: CDK5 Regulatory Subunit-Associated Protein 1-Like 1 Promotes Adipocyte Proliferation through Transcriptional Activation of NEK8 in T2DM and Obesity

Abstract

Background: Adipogenesis is a complex biological process and the main cause of obesity. CDK5 Regulatory Subunit-Associated Protein 1-like 1 (CDKAL1) serves as a susceptibility gene for obesity and type 2 diabetes (T2DM). However, whether CDKAL1 plays any functional role in adipocyte proliferation had not been well investigated. Methods: CDKAL1 expression was manipulated using a specific shRNA or the CDKAL1-full length expressing recombinant plasmids. MTT assay was performed to determine 3T3-L1 cell growth with either overexpression or downregulation of CDKAL1. Interaction between CDKAL1 and never-in-mitosis A-related kinase 8 (NEK8) in HEK293T and 3T3-L1 cells was examined using Co-immunoprecipitation (Co-IP) and Confocal immunofluorescence staining. Western blot assay was performed to determine the protein expression of NEK8 and CDKAL1. Dual-luciferase reporter assay was used to detect the transcriptional regulation of CDKAL1 on NEK8. The in vivo role of CDKAL1 and NEK8 was explored in an in vivo mouse model of T2DM. Results: In the present study, we found that overexpression of CDKAL1 promoted adipocyte proliferation of 3T3-L1 cells, whereas knockdown of CDKAL1 delayed 3T3-L1 proliferation. Mechanically, CDKAL1 interacted with NEK8 and transcriptionally activated its expression in adipocytes. Moreover, elevated levels of CDKAL1 and NEK8 were observed in patients with T2DM compared to the healthy subjects. Implantation of 3T3-L1 cells with CDKAL1 overexpression increased blood glucose in dietetic mice. Conclusions: Collectively, these findings suggest that CDKAL1 functions as a positive regulator of adipocyte proliferation, providing novel experimental evidences into the link between CDKAL1 and T2DM and obesity. Disclosure Y. Chen: None. Funding Chinese Postdoctoral Science Foundation (2019M651218); Natural Science Foundation of Jilin Province (20190201031JC)