C/EBPalpha is required to maintain postmitotic growth arrest in adipocytes.

Terminal differentiation is often coupled with irreversible loss of proliferative potential. The CCAAT enhancer binding protein alpha (C/EBPalpha) preferentially accumulates in postmitotic, differentiated 3T3-L1 adipocytes but declines during tumor necrosis factor alpha (TNFalpha)-induced dedifferentiation. We have discovered that this decline in C/EBPalpha correlates with an increased mitotic growth potential. In order to further investigate the antimitotic activity of C/EBPalpha, we introduced antisense C/EBPalpha RNA into 3T3-L1 cells to block endogenous C/EBPalpha expression. When treated according to the standard differentiation protocol, stable cells lines harboring antisense C/EBPalpha RNA did not differentiate into fat-laden adipocytes, consistent with previous findings (Lin F, Lane MD, Genes Dev 1992;6:533-544). We found that these undifferentiated cells expressing antisense-C/EBPalpha can reenter the cell cycle after mitogenic stimulation at a time in development when parental 3T3-L1 cells cannot. Moreover, the expression profiles of the growth-arrest-associated genes gas1 and gas2 revealed that the antisense C/EBPalpha-expressing cells withdrew from the cell cycle after the period of clonal expansion but failed to progress to the state of least proliferative potential characteristic of terminally differentiated adipocytes.