REGULATION AND EXPRESSION OF RETINOBLASTOMA PROTEINS P107 AND P130 DURING 3T3-L1 ADIPOCYTE DIFFERENTIATION

During 3T3-L1 adipocyte differentiation, growth-arrested, postconfluen t preadipocytes are required to reenter the cell cycle and proceed thr ough a mitotic clonal expansion phase prior to terminal differentiatio n. The retinoblastoma proteins (pRB, p107, and p130) are thought to be critical in controlling cell cycle progression by binding to and regu lating the activity of the E2F transcription factors.We show here that p130/p107 protein levels, p107 mRNA levels, and E2F DNA binding compl exes are regulated during 3T3-L1 adipogenesis. The predominant E2F bin ding complex in day 0 preadipocytes was p130 E2F with no detectable fr ee E2F or p107. On Day 1, during mitotic clonal expansion, there was a distinct switch to free E2F and p107-E2F complexes associated with in creased p107 mRNA and protein along with decreased p130 protein levels . Following differentiation, the day 0 pattern is reestablished. The s witch is not just a consequence of reentry into the cell cycle, in tha t p107 protein levels are both detectable and unchanged in dividing, s erum-restricted, or serum restimulated preconfluent cells. Interesting ly, hormonal stimulation of 3T3-C2 cells, a related nondifferentiating cell line, also induces a mitotic clonal expansion phase that is asso ciated with the p130:p107 switch in a pattern very similar to 3T3-L1 c ells, suggesting the block in differentiation observed in 3T3-C2 cells occurs after clonal expansion. Combined, these findings suggest that the regulatory mechanisms of the p130:p107 switch are not specific to differentiation but may play a key role in regulating the mitotic clon al expansion necessary for adipocyte differentiation in 3T3-L1 cells.