Regulated expression of three C/EBP isoforms during adipose conversion of 3T3-L1 cells.

TLDR

C/EBP β and δ share DNA‑binding properties with C/EBP α, readily heterodimerize, and their temporally distinct expression pattern suggests they orchestrate a regulatory cascade during adipocyte differentiation. The study aimed to identify protein factors regulating adipocyte differentiation by isolating genes encoding C/EBP α‑related polypeptides. The authors demonstrated transcriptional activation by C/EBP β and δ via transient transfection of serum albumin promoter constructs in hepatoma cells and characterized their expression during 3T3‑L1 adipogenesis using Western and Northern blotting. C/EBP β and δ mRNAs were expressed in tissues overlapping C/EBP α sites; they were induced early in 3T3‑L1 differentiation, directly by adipogenic hormones, peaked within two days, and declined before C/EBP α accumulation.

Abstract

In an effort to identify protein factors that play a regulatory role in the differentiation of adipocytes, we have isolated two genes that encode polypeptides related to CCAAT/enhancer-binding protein (C/EBP; hereafter termed C/EBP alpha). The proteins encoded by these C/EBP-related genes, termed C/EBP beta and C/EBP delta, exhibit similar DNA-binding specificities and affinities compared with C/EBP alpha. Furthermore, C/EBP beta and C/EBP delta readily form heterodimers with one another as well as with C/EBP alpha. The transcriptional activating capacity of these two newly identified C/EBP isoforms was demonstrated by transient transfection experiments in which expression vectors encoding C/EBP beta and C/EBP delta were observed to induce transcription from the promoter of the serum albumin gene in cultured hepatoma cells. The mRNAs encoding C/EBP beta and C/EBP delta were detected in a number of tissues, most of which corresponded to sites of expression of C/EBP alpha. The expression pattern of C/EBP beta and C/EBP delta during adipose conversion of 3T3-L1 cells was examined by Western and Northern blotting assays. In contrast to the expression profile of the gene encoding C/EBP alpha, whose product is not detectable until the late phase of adipocyte differentiation, the c/ebp beta and c/ebp delta genes were actively expressed very early during adipocyte differentiation. Moreover, transcription of the c/ebp beta and c/ebp delta genes was observed to be induced directly by adipogenic hormones. The accumulation of C/EBP beta and C/EBP delta reached a maximal level during the first 2 days of differentiation and declined sharply before the onset of C/EBP alpha accumulation. The temporal pattern of expression of these three C/EBP isoforms during adipocyte differentiation may reflect the underpinnings of a regulatory cascade that controls the process of terminal cell differentiation.