We have previously reported that tumor necrosis factor I8 (TNFO) expression is induced by interleukin-2 (IL-2) in the murine lymphocytic T-cell line CTLL-2. In this study, we have characterized the nuclear and cytoplasmic TNFi transcripts and assessed their role in TNF(8 gene expression. A unique feature of TNFji expression was the accumulation of nuclear precursors, which reflected a slow nuclear RNA processing. As a consequence, there was a delay in the appearance of cytoplasmic messengers after the transcriptional induction of TNF,3 by IL-2. We also found that two messengers, the fully spliced messenger and an intron 3-retaining messenger, were exported to the cytoplasm and actively translated. The same pattern of expression was observed in concanavalin A-stimulated splenocytes, although the level of expression was much lower than in CTLL-2 cells. The simple genetic structure and the high level of accumulation of nuclear precursors make TNF,3 a particularly attractive model system to use for studies of RNA processing and cytoplasmic transport of partially spliced messengers.
The glycolytic enzyme enolase (EC 4.2.1.11) exists as dimers formed from three structurally related subunits alpha, beta, and gamma, encoded by separate genes. The gene encoding the beta-subunit is expressed only in striated muscles. We have previously shown that the beta-enolase gene belongs to a small subset of muscle-specific genes showing transcriptional activity in cultured myoblasts, prior to withdrawal from the cell cycle. An increase in the level of beta-enolase mRNA occurs during terminal differentiation of myoblasts. To investigate the mechanisms underlying this increase, we have simultaneously estimated, under steady state conditions, the rate of synthesis and the stability of beta-enolase mRNA in proliferating C2.7 myoblasts as well as in differentiating myotubes. The method used is based on the isolation of newly synthesized RNA from the total RNA pool, following pulse-labeling of intact cells in the presence of 4-thiouridine. The results described here demonstrate a coordinate increase in newly synthesized and total beta-enolase mRNA, while the mRNA half-life, about 4 hr, remains unchanged in the course of terminal differentiation. The expression of the gene for insulin-like growth factor-II (IGF-II), a major positive regulator of myogenesis, was analyzed using the same approach. It is concluded that the up-regulation of beta-enolase as well as IGF-II gene expression in differentiating muscle cells reflects an increased rate of entry of newly synthesized mRNAs into the general pool of transcripts without changes in their respective half-lives.
We have previously reported that tumor necrosis factor beta (TNF beta) expression is induced by interleukin-2 (IL-2) in the murine lymphocytic T-cell line CTLL-2. In this study, we have characterized the nuclear and cytoplasmic TNF beta transcript and assessed their role in TNF beta gene expression. A unique feature of TNF beta expression was the accumulation of nuclear precursors, which reflected a slow nuclear RNA processing. As a consequence, there was a delay in the appearance of cytoplasmic messengers after the transcriptional induction of TNF beta by IL-2. We also found that two messengers, the fully spliced messenger and an intron 3-retaining messenger, were exported to the cytoplasm and actively translated. The same pattern of expression was observed in concanavalin A-stimulated splenocytes, although the level of expression was much lower than in CTLL-2 cells. The simple genetic structure and the high level of accumulation of nuclear precursors make TNF beta a particularly attractive model system to use for studies of RNA processing and cytoplasmic transport of partially spliced messengers.
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