Lissemore J L, Quail P H
Department of Botany, University of Wisconsin, Madison.
Mol Cell Biol. 1988 Nov;8(11):4840-50. doi: 10.1128/mcb.8.11.4840-4850.1988.
We have examined phytochrome-regulated transcription of phytochrome (phy) and chlorophyll a/b binding protein (cab) genes in dark-grown Avena seedlings by using run-on transcription in isolated nuclei. Kinetic analysis of phy transcription following pulse-light treatments to produce various amounts of Pfr, the active form of phytochrome, leads to these conclusions. (i) Transcription decreases rapidly (discernible within 5 min) after Pfr formation, reaching an essentially undetectable level by 1 h. (ii) The response is very sensitive; less than 1% Pfr is sufficient to produce maximum feedback repression over the first 30 min. (iii) The duration of transcriptional repression is proportional to the Pfr concentration; derepression begins once the concentration falls below some saturation level because of degradation of Pfr. Concurrent analysis of cab transcription leads to these conclusions. (i) After Pfr formation, transcription increases approximately 10-fold by 3 h, but this response is not detectable until after a 30-min lag. (ii) Detectable induction of cab requires a greater than 30-fold-higher Pfr level than is needed to repress phy expression. (iii) Transcription returns to the preirradiation level considerably sooner than does phy transcription (less than 12 h versus greater than 24 h respectively), indicating that a high level of Pfr is needed to sustain the increased transcription of cab. Taken together, these results suggest that differences in the phytochrome signal transduction pathway are responsible for the distinct patterns of regulation of these genes. Full repression of phy occurs even when protein synthesis is inhibited greater than 90% by cycloheximide and chloramphenicol. In conjunction with the rapidity of the response to Pfr, this result provides evidence that feedback repression of phy gene transcription does not require expression of an intervening regulatory gene(s). Thus, phy is the first gene for which there is evidence for direct control of transcription by the phytochrome signal transduction chain.
我们通过在分离的细胞核中进行连续转录，研究了黑暗中生长的燕麦幼苗中光敏色素（phy）和叶绿素a/b结合蛋白（cab）基因的光敏色素调节转录。对经脉冲光照处理以产生不同量的Pfr（光敏色素的活性形式）后的phy转录进行动力学分析，得出以下结论。（i）Pfr形成后，转录迅速下降（5分钟内可察觉），到1小时时达到基本无法检测的水平。（ii）该反应非常敏感；在前30分钟内，不到1%的Pfr就足以产生最大程度的反馈抑制。（iii）转录抑制的持续时间与Pfr浓度成正比；一旦由于Pfr的降解，浓度降至某个饱和水平以下，去抑制就开始了。对cab转录的同时分析得出以下结论。（i）Pfr形成后，转录在3小时内增加约10倍，但直到30分钟的延迟后才能检测到这种反应。（ii）可检测到的cab诱导所需的Pfr水平比抑制phy表达所需的水平高30倍以上。（iii）转录恢复到照射前水平的时间比phy转录早得多（分别小于12小时和大于24小时），这表明需要高水平的Pfr来维持cab转录的增加。综上所述，这些结果表明，光敏色素信号转导途径的差异是这些基因不同调控模式的原因。即使在用环己酰亚胺和氯霉素将蛋白质合成抑制90%以上时，phy也会完全被抑制。结合对Pfr反应的快速性，这一结果证明phy基因转录的反馈抑制不需要中间调控基因的表达。因此，phy是第一个有证据表明其转录受光敏色素信号转导链直接控制的基因。
