Identification and Characterization of a Transcript for a Novel Rac GTPase-Activating Protein in Terminally Differentiating 3T3-L1 Adipocytes

Abstract

Using differential display, we sought to identify novel genes expressed in the early stages of 3T3-L1 adipocyte differentiation. A gene which we have named "band25" was identified, and a full-length cDNA sequence was assembled. Sequence analysis revealed that the 2842-bp cDNA encodes a putative 628-amino acid protein product, which is a member of the GTPase-activating protein (GAP) family. This gene may be the murine homolog of the human M gcRacGAP protein, which was identified in male germ cells. Other closely related proteins include the Drosophila protein Rotund, several chimerins, and the human breakpoint cluster region (Bcr) protein. These GAP proteins all specifically inactivate Rac, a member of the Ras-like family of proteins. A consensus sequence for a diacyl glycerol/phorbol ester-binding domain was also found in the Band25 sequence. The expression of band25 mRNA is regulated during the differentiation of both adipocytes and myoblasts. Its mRNA was shown to be expressed at a low level in confluent 3T3-L1 preadipocytes and in differentiated 3T3-L1 adipocytes. Expression of band25 was increased 15.5 fold by 24 h after the induction of differentiation, when 3T3-L1 cells undergo several rounds of postconfluent cell division. Expression was also high in growing 3T3-L1 and C2C12 cells but decreased progressively as C2C12 cells underwent differentiation. These observations suggest that the expression of band25 is growth regulated and that the protein could play a role in the regulation of growth-related processes.