We have isolated and sequenced a full-length cDNA clone encoding rat glyceraldehyde-3-phosphate-dehydrogenase (GAPDH, E.C.1.2.1.12). The entire mRNA is 1269 nucleotides long exclusive of poly(A) and contains respectively 71 and 196 bases of 5' and 3' non-coding regions. Primer extension as well as S1 nuclease protection experiments clearly established that a single (or at least a highly prominent) GAPDH mRNA species is expressed in all rat tissues examined. This sequence allowed the determination of the hitherto unknown primary structure of rat GAPDH which is 333 aminoacids long. Comparison between GAPDH sequences from rat, man and chicken revealed a high degree of sequence conservation at both nucleotide and protein levels.

To address the possibility that the expression of the myc gene might be regulated at a post-transcriptional level, we have investigated the half-life of myc mRNA in various cells. Our survey included normal human embryonic fibroblasts as well as transformed human cells of various origins: cervix carcinoma (HeLa), breast carcinoma (MCF7), Burkitt lymphoma (Daudi), and promyelocytic leukemia (HL60). All these cells revealed an extreme instability of myc mRNA (half-life, 10 min), suggesting that the control of myc mRNA degradation might be a general means (although not necessarily exclusive) of regulating both the level and the timing of myc gene expression. Inhibition of protein synthesis resulted in a dramatic stabilization of myc mRNA in HeLa, MCF7, and HL60 cells, suggesting that the controlling element might itself be, at least in these cells, a protein of rapid turnover. This finding opens the way to studying the mechanism of myc mRNA inactivation in these different cell types. However, protein synthesis inhibition had no effect on myc mRNA instability in other transformed (Daudi) cell lines as well as normal embryonic human fibroblasts. These different types of behavior suggest that the post-transcriptional control of myc gene expression might involve multiple factors that would be differently affected in various cell types.

The recent observation made in our laboratory that cellular myc (c-myc) mRNA has a very short half-life in a variety ofnormal and transformed human cells emphasized the potential importance of post-transcriptional regulation of c-myc gene expression. Jonak and Knight [Jonak, G. J. & Knight, E., Jr. (1984) Proc. NatI. Acad. Sci. USA 81, [1747][1748][1749][1750] have reported a selective reduction of c-myc mRNA accumulation in lymphoblastoid Daudi cells treated with human (3 interferon. This provided a suitable situation in which to examine a possible action of negative modulators of c-myc expression at the level of mRNA stability. Our results confirm the observation by Jonak and Knight that c-myc mRNA level is depressed in cells treated with i interferon and extend it to a2 interferon. Furthermore, we now demonstrate that interferon has no effect on c-myc transcription rate in isolated nuclei but rather reduces the half-life of its mRNA. Conversely, we show that it increases the level of HLA-A2 mRNA by stimulating its transcription.The pathway that leads from a normal cell to a fully transformed one is controlled by at least two distinct classes of oncogenes acting in concert (for a review, see ref. 1). c-myc, the cellular homologue ofthe avian myelocytomatosis virus (MC29) oncogene, belongs to the establishment-immortalization class along with adenovirus Ela and the polyoma gene encoding the large tumor (T) antigen. To the second class, referred to as transforming genes, belong the ras family of oncogenes and the polyoma gene encoding middle T antigen.The expression of the c-myc gene can be modulated in opposite directions. Enhanced expression of c-myc mRNA has been observed in a wide spectrum of neoplasms. It also can be induced experimentally early after mitogenic stimulation of lymphocytes by lipopolysaccharides or concanavalin A and offibroblasts by platelet-derived-growth factor (2). Conversely, HL60, a human promyelocytic leukemia cell line, has been shown to respond by an early decrease in myc expression to agents that induce its granulocytic differentiation (3,4). Along the same vein, Jonak and Knight (5) Several models of c-myc regulation by trans-acting elements, possibly repressors, have been proposed (6-8). On the other hand, we have shown that c-myc mRNA is extremely unstable in both normal and transformed cells (9), suggesting that it might, at least for a large part, be posttranscriptionally regulated at the level of its degradation. Thus, it was tempting to assess the contribution of transcriptional and post-transcriptional events to the effects of positive and negative modulators of c-myc expression. We selected IFNs as representatives of this latter type of modulators. The present results show that the reduced level of steady-state c-myc mRNA in IFN-treated cells can be accounted for by an increased rate of its degradation. A preliminary account of this work has recently been published in abstract form (10). Hu-IFN-a and -,/, human a and /3 interferons; GADPH, glyceraldehyde-3-phosphat...

c-myc RNA rapidly decreases to barely detectable levels in Friend erythroleukemia cells induced to differentiate upon the addition of dimethylsulfoxide. We show here that c-myc gene is down-regulated both at the transcriptional level presumably by a block in the elongation of primary transcripts and at the post-transcriptional level by an increase in the degradation of its mRNA.

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