ABSTRACT
The events at the beginning of adipocyte differentiation are not well known. We previously cloned the genes expressed early in the differentiation of mouse 3T3-L1 preadipocyte cells. One of them, similar in sequence to human TC10, was identified as TC10-like/TC10betaLong (TCL/TC10betaL), a new Rho GTPase by the cloning of full-length cDNA. The expression of TCL/TC10betaL increased rapidly right after the addition of inducers for differentiation, whereas the levels of other Rho family genes were unchanged at this stage. The antisense TCL/TC10betaL-expressing experiment revealed that the differentiation of 3T3-L1 cells into adipocytes was inhibited. Moreover, the sense TCL/TC10betaL-expressing experiment using NIH-3T3 cells, which do not usually differentiate into adipocytes, clearly showed the accumulation of oil droplets as well as the elevated expression of various adipogenic marker genes in the presence of the ligand for peroxisome proliferator-activated receptor gamma (PPARgamma). These results strongly indicated that TCL/TC10betaL has a crucial role in the early stage of adipocyte differentiation, probably linked to the PPARgamma pathway. Using a subtraction protocol, the genes specifically regulated by TCL/TC10betaL were also isolated. The expression pattern of some of them was similar to TCL/TC10betaL expression in adipogenesis, suggesting that the expression of these genes would be regulated by TCL/TC10betaL.
Subject(s)
Adipocytes/cytology , GTP Phosphohydrolases/physiology , 3T3 Cells , Animals , Cell Differentiation , Cloning, Molecular , DNA, Complementary/isolation & purification , GTP Phosphohydrolases/antagonists & inhibitors , GTP Phosphohydrolases/genetics , Gene Expression Profiling , Mice , Oligonucleotides, Antisense/pharmacology , RNA, Messenger/analysis , Receptors, Cytoplasmic and Nuclear/physiology , Transcription Factors/physiology , Transfection , Up-Regulation , rho GTP-Binding Proteins/genetics , rho GTP-Binding Proteins/physiologyABSTRACT
Adipocyte differentiation is a complex process in which the expression of many transcription factors and adipocyte-specific genes is regulated under a strict program. The peroxisome proliferator-activated receptor gamma (PPARgamma), a member of the steroid/thyroid nuclear hormone receptor superfamily of ligand-activated transcription factors, is an important regulator of adipocyte gene expression and differentiation. In this study, we tried to identify downstream target genes of PPARgamma, by using PPARgamma-expressing cells and a subtractive cloning strategy, and isolated cDNA clones which were up-regulated or down-regulated by PPARgamma. Northern blot analyses revealed that the expression levels of the aldehyde dehydrogenase-2-like, type VI collagen alpha 3 subunit, cellular retinoic acid binding protein 1 and thrombospondin 1 are changed during the differentiation of mouse 3T3-L1 preadipocyte cells, indicating that these genes might be downstream targets of PPARgamma in adipocyte differentiation.
