Transcription factor activating enhancer-binding protein-2beta. A negative regulator of adiponectin gene expression.

We previously reported the association between the activating enhancer-binding protein-2beta (AP-2beta) transcription factor gene and type 2 diabetes. This gene is preferentially expressed in adipose tissue, and subjects with the disease-susceptible allele of AP-2beta showed stronger expression in adipose tissue than those without the susceptible allele. Furthermore, overexpression of AP-2beta leads to lipid accumulation by enhancing glucose transport and inducing insulin resistance in 3T3-L1 adipocytes. In this study we demonstrated that overexpression of AP-2beta in 3T3-L1 adipocytes decreased the expression and secretion of adiponectin and increased those of interleukin-6 (IL-6). Interestingly, the effects of AP-2beta on the expressions of adiponectin and IL-6 and the mechanisms by which AP-2beta modulated their expressions were different. We found that the promoter activity of adiponectin gene was inhibited by AP-2beta overexpression and enhanced by knockdown of endogenous AP-2beta, whereas IL-6 was unaffected. Electrophoretic mobility shift assays revealed the existence of putative responsive elements for AP-2beta and NF-YA in human and mouse adiponectin promoter regions, and mutation of this AP-2beta binding site abolished the inhibitory effect of AP-2beta. Furthermore, chromatin immunoprecipitation assays demonstrated that AP-2beta and NF-YA competitively bind to the same region of the adiponectin promoter. Our results clearly demonstrated that AP-2beta directly inhibits adiponectin gene expression by displacing NF-YA and binding to its promoter. We conclude that AP-2beta might modulate the expression of adiponectin by directly inhibiting its transcriptional activity.

The transcription factor AP-2beta causes cell enlargement and insulin resistance in 3T3-L1 adipocytes.

We have reported the association of variations in the activating protein-2beta (AP-2beta) transcription factor gene with type 2 diabetes. This gene was preferentially expressed in 3T3-L1 adipocytes in a differentiation stage-dependent manner, and preliminary experiments showed that subjects with the disease-susceptible allele showed stronger expression in adipose tissue than those without the susceptible allele. Thus, we overexpressed the AP-2beta gene in 3T3-L1 adipocytes to clarify whether AP-2beta might play a crucial role in the pathogenesis of type 2 diabetes through dysregulation of adipocyte function. In cells overexpressing AP-2beta, cells increased in size by accumulation of triglycerides accompanied by enhanced glucose uptake. On the contrary, suppression of AP-2beta expression by small interfering RNA inhibited glucose uptake. Enhancement of glucose uptake by AP-2beta overexpression was attenuated by inhibitors of phospholipase C (PLC) and atypical protein kinase Czeta/lambda (PKCzeta/lambda), but not by a phosphatidylinositol 3-kinase (PI3-K) inhibitor. Consistently, we found activation of PLC and atypical PKC, but not PI3-K, by AP-2beta expression. Furthermore, overexpression of PLCgamma enhanced glucose uptake, and this activation was inhibited by an atypical PKC inhibitor, suggesting that the enhanced glucose uptake may be mediated through PLC and atypical PKCzeta/lambda, but not PI3-K. Moreover, we observed the increased tyrosine phosphorylation of Grb2-associated binder-1 (Gab1) and its association with PLCgamma, indicating that Gab1 may be involved in AP-2beta-induced PLCgamma activation. Finally, AP-2beta overexpression was found to relate to the impaired insulin signaling. We propose that AP-2beta is a candidate gene for producing adipocyte hypertrophy and may relate to the abnormal characteristics of adipocytes observed in obesity.