Almouzni G, Wolffe A P
Laboratory of Molecular Embryology, National Institute of Child Health and Human Development, NIH, Bethesda, MD 20892, USA.
EMBO J. 1995 Apr 18;14(8):1752-65. doi: 10.1002/j.1460-2075.1995.tb07164.x.
We have examined the cause of transcriptional quiescence prior to the mid-blastula transition (MBT) in Xenopus laevis. We have found distinct requirements for transcription of class II and class III genes. An artificial increase of the amount of DNA present within the embryo over that found at the MBT allows precocious transcription of tRNA genes, but not of the adenovirus E4 or human cytomegalovirus (CMV) promoters. Thus titration of an inhibitor by exogenous DNA determines class III but not class II gene activation. We demonstrate that the action of the inhibitor depends on the association of core histones with DNA. The addition of exogenous TBP, together with an increase in the amount of DNA within the embryo, allows significant basal transcription of class II genes prior to the MBT, whereas it does not increase transcription of tRNA genes. To examine the activation of transcription above basal levels, we used a defined minimal promoter containing five Gal4 binding sites and the activator Gal4-VP16. Precocious transcriptional activation is directed by Gal4-VP16 prior to the MBT, demonstrating that a functional transcriptional machinery exists at this early developmental stage. Furthermore, since this activation can occur in the absence of exogenous TBP or chromatin titration, a transcription factor that can penetrate chromatin is sufficient for recruitment of this machinery to a promoter. Our results support the hypothesis that the temporal regulation of transcription during early embryogenesis in Xenopus reflects not only a titration of inhibitors by DNA, but also a deficiency in the activity of transcriptional activators prior to the MBT.
我们研究了非洲爪蟾中囊胚转换（MBT）之前转录静止的原因。我们发现了对II类和III类基因转录的不同要求。人为增加胚胎内DNA的量，使其超过在MBT时发现的量，可使tRNA基因提前转录，但腺病毒E4或人巨细胞病毒（CMV）启动子则不能。因此，外源DNA对抑制剂的滴定决定了III类而非II类基因的激活。我们证明抑制剂的作用取决于核心组蛋白与DNA的结合。在胚胎内DNA量增加的同时添加外源TBP，可使MBT之前的II类基因进行显著的基础转录，而这并不会增加tRNA基因的转录。为了检测基础水平之上的转录激活，我们使用了一个包含五个Gal4结合位点和激活剂Gal4-VP16的确定的最小启动子。在MBT之前，Gal4-VP16可引导早熟的转录激活，这表明在这个早期发育阶段存在功能性的转录机制。此外，由于这种激活可以在没有外源TBP或染色质滴定的情况下发生，一种能够穿透染色质的转录因子就足以将这种机制招募到启动子上。我们的结果支持这样一种假说，即非洲爪蟾早期胚胎发育过程中转录的时间调控不仅反映了DNA对抑制剂的滴定，还反映了MBT之前转录激活剂活性的不足。
