Identification and analysis of Chlamydomonas mutants with defects in chlorophyll synthesis

衣藻的突变体耐受叶绿素合成的遗传损伤，只要积累叶绿素前体不能产生光氧化应激。因此，叶绿素合成基因的敲除突变体可以在黑暗中在额外的碳源上生长并被分析，而相应的植物突变体将是胚胎致死的。我们继续筛选一系列黑暗生长的有趣的光敏突变体，棕色，黄色和黄绿色色素沉着，这都是由不同的积累叶绿素前体，光敏这些突变体，使他们严格的光合自养增长指定。主要目的是详细分析三个突变体，无论是删除GUN4或HEM4基因或显示镁原卟啉单甲酯的积累，表明受损的环化酶反应。基因的表达以及四吡咯生物合成的酶活性将在这些突变体中进行研究，从而预测衣原体影响四吡咯生物合成和光合作用的调控的转录和翻译后调控机制，这可能不同于先前分析的维管植物。其他研究包括评估（i）LHC表达在增加光强度响应受损的叶绿素生物合成和（ii）蛋白质之间的相互作用的酶和Mg分支的因素作为一个先决条件，有效的基板通道和避免四吡咯依赖性光氧化应激。

Mutants of Chlamydomonas reinhardtii tolerate genetic lesions in chlorophyll synthesis as long as accumulating chlorophyll precursors cannot generate photooxidative stress. Therefore, knock-out mutants for genes in chlorophyll synthesis can grow in darkness on an additional carbon source and be analyzed, while corresponding plant mutants would be embryo-lethal. We continue to screen a collection of dark-grown interesting photosensitive mutants with brown, yellow and yellow-green pigmentation, which are all specified by diverse accumulating chlorophyll precursors that photosensitize these mutants and disable them to grow strictly photoautotrophically. The main objective is the detailed analysis of three mutants with either deleted GUN4 or HEM4 genes or displaying an accumulation of Mg protoporphyrin monomethylester indicating an impaired cyclase reaction. The expression of genes as well as enzyme activities of tetrapyrrole biosynthesis will be studied in these mutants allowing a prediction of transcriptional and posttranslational regulatory mechanisms of Chlamydomonas affecting regulation of tetrapyrrole biosynthesis and photosynthesis, which might be different from those of previously analyzed vascular plants. Other studies include assessments of (i) LHC expression during increasing light intensities in response to impaired chlorophyll biosynthesis and (ii) protein interactions among enzymes and factors of the Mg branch as a prerequisite for efficient substrate channeling and avoidance of tetrapyrrole-dependent photooxidative stress.