Lee Jaesung, Herrin David L
Molecular Cell and Developmental Biology Section and Institute for Cellular and Molecular Biology, School of Biological Sciences, University of Texas at Austin, Austin, TX, 78712, USA,
Photosynth Res. 2002;72(3):295-306. doi: 10.1023/A:1019881306640.
Previous work has shown that transcription of a number of chloroplast-encoded genes, including those for photosynthesis, are under circadian clock control in Chlamydomonas reinhardtii. However, some of these genes encode long-lived mRNAs that are also subject to translational control. Rates of synthesis of the major chloroplast translation products vary dramatically (10-20-fold) during light-dark (LD) cycles, peaking in the light period. To determine whether this pattern reflects circadian clock control, LD-grown cells were shifted to continuous light (LL) and chloroplast protein synthesis monitored by periodic pulse-labeling in the presence of cycloheximide; chloroplast protein synthesis in LD was also examined for comparison. The LD patterns of synthesis of the major polypeptides (including D1, D2, and the large subunit of ribulose-1,5-bisphosphate carboxylase (LS)) were similar to those obtained previously in the absence of cycloheximide. In the LL condition, rates of synthesis of the major chloroplast translation products were high throughout the period examined ( approximately 36 h), fluctuating > 3-fold, although they were generally higher in the subjective light period. LD-grown cells were also shifted to continuous dark (DD) and chloroplast protein synthesis analyzed for approximately 24 h starting from the mid-dark period. There was a gradual decline in synthesis of the major proteins during the first subjective light period, which was followed by a very small peak in synthesis around the second subjective dark --> light transition. RNA blot analysis showed that the mRNAs for D1, D2 and LS were present at high levels during the period of declining translation. These results indicate that with photoautotrophic growth in LD cycles, the illumination conditions per se are more important than the clock in determining chloroplast translation, but the clock may contribute to this regulation. The advantages of controlling translation by a direct light response and transcription primarily by the circadian clock are discussed. Finally, evidence of translational control of elongation factor Tu synthesis was obtained.
先前的研究表明，莱茵衣藻中一些叶绿体编码基因（包括光合作用相关基因）的转录受生物钟控制。然而，其中一些基因编码的是长寿命mRNA，它们也受到翻译调控。在明暗（LD）循环过程中，主要叶绿体翻译产物的合成速率变化显著（10 - 20倍），在光照期达到峰值。为了确定这种模式是否反映了生物钟控制，将在LD条件下生长的细胞转移到持续光照（LL）条件下，并在存在环己酰亚胺的情况下通过定期脉冲标记监测叶绿体蛋白质合成；同时也检测了LD条件下的叶绿体蛋白质合成以作比较。主要多肽（包括D1、D2和1,5 - 二磷酸核酮糖羧化酶大亚基（LS））的LD合成模式与先前在不存在环己酰亚胺的情况下获得的模式相似。在LL条件下，在所检测的整个时期（约36小时）内，主要叶绿体翻译产物的合成速率都很高，波动超过3倍，尽管在主观光照期通常更高。将在LD条件下生长的细胞也转移到持续黑暗（DD）条件下，并从黑暗中期开始分析约24小时的叶绿体蛋白质合成。在第一个主观光照期，主要蛋白质的合成逐渐下降，随后在第二个主观黑暗到光照转变前后合成出现一个非常小的峰值。RNA印迹分析表明，在翻译下降期间，D1、D2和LS的mRNA水平很高。这些结果表明，在LD循环中进行光合自养生长时，光照条件本身在决定叶绿体翻译方面比生物钟更重要，但生物钟可能有助于这种调节。文中讨论了通过直接光反应控制翻译和主要通过生物钟控制转录的优势。最后，获得了延伸因子Tu合成受翻译调控的证据。
