TUG is a calpain-10 substrate involved in the translocation of GLUT4 in adipocytes

The calpain-10 (CAPN10) protease is implicated in the translocation of the glucose transporter 4 (GLUT4), which is retained in the Golgi matrix via the Tether containing a UBX domain for GLUT4 (TUG) protein. Insulin stimulation induces the proteolytic processing of TUG, which leads to the translocation of GLUT4 to the cell membrane. We tested whether TUG is a CAPN10 substrate. Proteolysis of TUG by calpains was assessed using a cell-free system containing calpain-1 and TUG. In situ proteolysis of TUG by calpains was demonstrated in 3T3-L1 adipocytes in the presence of insulin or calpain inhibitors to modulate calpain activity. Proteolysis of TUG by CAPN10 was confirmed using transient or stable silencing of CAPN10 in 3T3-L1 adipocytes. Calpains proteolyzed the C-terminus of TUG in vitro. In adipocytes, insulin-induced cleavage of TUG was correlated with the activation of calpains. Treatment with calpain inhibitors reduced TUG cleavage, resulting in impaired GLUT4 translocation without altering Akt phosphorylation. Furthermore, CAPN10 but not calpain-1 or calpain-2 colocalized with GLUT4 in the absence of insulin, and their colocalization was reduced after stimulation with insulin. Finally, we demonstrated that CAPN10 knockdown reduced the proteolysis of TUG without altering the phosphorylation of Akt or the expression of the Usp25m protease. Thus, our results provide evidence that the TUG protein is cleaved by CAPN10 to regulate GLUT4 translocation.

CAPN10 SNP-19 is Associated with Susceptibility of Type 2 Diabetes Mellitus: A Javanese Case-control Study

BACKGROUND: The health data of Central Java, Indonesia showed that diabetes mellitus (DM) was the second most increasing non-communicable disease in the province. More than 20 genes have been reported to be associated with DM. Calpain-10 (CAPN10) polymorphism has been reported to be associated with Type 2 Diabetes Mellitus (T2DM). However, the association between CAPN10 single nucleotide polymorphism (SNP)-19 and T2DM among Javanese ethnics has never been reported. Therefore, this study was conducted to investigate the association.METHODS: After fasting for 8 hours, blood samples were drawn from veins of 107 T2DM and 107 non diabetic subjects. A half of the drawn blood was collected for identification of CAPN10 SNP-19, and another half for measuring triglycerides and fasting blood glucose (FBG). Identification of CAPN10 SNP-19 was performed with polymerase chain reaction (PCR) method, while measurement of triglycerides and FBG with colorimetric enzymatic method.RESULTS: The number of T2DM Javanese subjects with 2R/3R and 3R/3R CAPN10 SNP-19 genotypes was significantly higher than the number of T2DM Javanese subjects with 2R/2R genotype (p=0.002). When each number of 2R/3R and 3R/3R T2DM subjects was compared with the number of 2R/2R T2DM subjects, the number of 2R/3R T2DM subjects was significantly higher than the number of 2R/2R T2DM subjects (p=0.000).CONCLUSION: Javanese subjects with 2R/3R and 3R/3R CAPN10 SNP19 genotypes might have susceptibility of T2DM.KEYWORDS: Calpain-10, CAPN10, polymorphism, type2 diabetes mellitus, triglycerides, fasting blood glucose