Manrow R E, Jacobson A
Department of Molecular Genetics and Microbiology, University of Massachusetts Medical School, Worcester 01655.
Mol Cell Biol. 1988 Oct;8(10):4088-97. doi: 10.1128/mcb.8.10.4088-4097.1988.
We reevaluated the use of 32PO4 pulse-chases for analyzing mRNA decay rates in late-developing Dictyostelium cells. We found that completely effective PO4 chases could not be obtained in developing cells and that, as a consequence, the decay rates exhibited by some mRNAs were influenced by the rates at which they were transcribed. In developing cells disaggregated in the presence of cyclic AMP, the poly(A)+ mRNA population turned over with an apparent half-life of 4 h, individual mRNA decay rates were heterogeneous, and some prestalk and prespore mRNAs appeared to decay with biphasic kinetics. In cells disaggregated in the absence of cyclic AMP, all prestalk and prespore mRNAs decayed with biphasic kinetics. During the first 1 to 1.5 h after disaggregation in the absence of cyclic AMP, the cell-type-specific mRNAs were selectively degraded, decaying with half-lives of 20 to 30 min; thereafter, the residual prestalk and prespore mRNA molecules decayed at rates that were similar to those measured in the presence of cyclic AMP. This short-term labilization of cell-type-specific mRNAs was observed even for those species not requiring cyclic AMP for their accumulation in developing cells. The observation that cell-type specific mRNAs can decay at similar rates in disaggregated cells with or without cyclic AMP indicates that this compound does not act directly to stabilize prestalk and prespore mRNAs during development and that its primary role in the maintenance of cyclic-AMP-dependent mRNAs is likely to be transcriptional.
我们重新评估了使用³²PO₄脉冲追踪法来分析发育后期盘基网柄菌细胞中mRNA的衰变率。我们发现,在发育中的细胞中无法获得完全有效的PO₄追踪,因此,一些mRNA显示出的衰变率受到其转录速率的影响。在存在环磷酸腺苷(cAMP)的情况下分散培养的发育细胞中,聚腺苷酸(poly(A)⁺)mRNA群体以约4小时的表观半衰期周转,单个mRNA的衰变率是异质的,一些前柄和前孢子mRNA似乎以双相动力学衰变。在不存在cAMP的情况下分散培养的细胞中,所有前柄和前孢子mRNA均以双相动力学衰变。在不存在cAMP的情况下分散培养后的最初1至1.5小时内,细胞类型特异性mRNA被选择性降解,半衰期为20至30分钟;此后,残留的前柄和前孢子mRNA分子的衰变速率与在存在cAMP的情况下测得的速率相似。即使对于那些在发育细胞中积累不需要cAMP的mRNA种类,也观察到了这种细胞类型特异性mRNA的短期不稳定现象。在有或没有cAMP的分散细胞中,细胞类型特异性mRNA可以以相似的速率衰变,这一观察结果表明,该化合物在发育过程中并不直接作用于稳定前柄和前孢子mRNA,其在维持依赖cAMP的mRNA中的主要作用可能是转录方面的。