Complexity of the early genetic response to growth factors in mouse fibroblasts.

Abstract

Genes whose expression is growth factor regulated are likely to be important components in the mechanisms controlling cell proliferation and differentiation. With the aim of identifying some of those genes, a A cDNA library was prepared with poly(A)+ RNA from quiescent NIH 3T3 cells stimulated with serum for 4 h in the presence of cycloheximide. Differential screening of approximately 200,000 recombinant phage plaques revealed 2,540 clones that cross hybridized preferentially with [32P]cDNA derived from RNA of stimulated cells rather than with cDNA derived from nonstimulated cells. Cross hybridization of these clones identified 82 independent sequences, including c-fos and c-myc. Seventy-one clones were further studied. Analysis of the changes in transcription and mRNA levels after serum stimulation demonstrated that the kinetics and extent of the induction vary dramatically between the different genes. Cycloheximide in all cases superinduced the mRNA levels by two mechanisms, inhibiting the shutoff of transcription and prolonging the half-lives of the mRNAs. Our results showed that induction of proliferation is accompanied by the onset of a complex genetic program.A detailed knowledge of the biochemical events and the identification of the genes that are involved in the response to growth factors is currently being accumulated. A subset of these genes may play a critical role in cellular transformation. It is well established that the synthesis of new mRNA in quiescent cells is required for the cells to respond to mitogens and progress through Gl and enter the S phase (1). Furthermore, Gl seems to be a critical point of control of proliferation, where normal cells deprived of growth factors halt cycling, while transformed cells do not, suggesting that regulatory genes uncontrolled in the neoplastic phenotype are expressed during this transition. It has been estimated that in mouse fibroblasts, 3% of the mRNA species in logarithmically growing cells are absent in quiescent cells (49); thus, a set of these genes must be under the control of growth factors to allow cell proliferation. Evidence that gene expression could be controlled by the binding of a specific ligand to its receptor initially came from studies done with beta interferon (27,28) which demonstrated the transcriptional activation of two genes in human cells after interferon treatment. These findings were further supported by the interesting observation that the proto-oncogenes c-fos and c-myc present an immediate change in expression after stimulation of fibroblasts with growth factors (4,8,14,23,25,32), emphasizing also that important growth-controlling genes operate during the GO-to-Gl transition. Both genes are induced in the presence of protein synthesis inhibitors, demonstrating that their induction is a direct consequence of the growth factor-receptor interaction. Expression of c-fos is undetectable during the cell cycle (3), suggesting that its expression could be essential during the transition from GO to Gl and not for continuously growing ...

Molecular cloning, sequencing, and mapping of EGR2, a human early growth response gene encoding a protein with "zinc-binding finger" structure.

“…Genes controlling proliferation or differentiation of eukaryotic cells have been identified by differential screening (1)(2)(3)(4)(5) of cDNA libraries. We (6,7) and others (2,3) have identified cDNAs the expression of which is upregulated by serum stimulation of quiescent mouse fibroblasts.…”

Transformation by ras modifies AP1 composition and activity

“…Moreover, the jun and fos immediate early genes are di erentially expressed during development and in adult tissues (Dony and Gruss, 1987;Hirai et al, 1989;Wilkinson et al, 1989). They also respond di erently to mitogenic stimuli, genotoxic or differentiating agents (Almendral et al, 1988;de Groot et al, 1990;Devary et al, 1991;Lau and Nathans, 1987; Ras Protein Clone…”

The mammalian Jun proteins: redundancy and specificity

“…Upon serum stimulation, many quiescent cells displayed an increase in c-jun, junB and fos mRNAs (Herschman, 1991). Based on their rapid, high and transient transcriptional upregulation in response to mitogenic agents, these agents were de®ned as`immediate early genes' or`competence genes' and encode products necessary for entry into G1 and S phases of the cell cycle (Lau and Nathans, 1987;Almendral et al, 1988). In the following sections, we will review the function of the three members of the Jun family in di erent biological processes.…”