拟南芥WHIRLY2基因超表达引起拟南芥角果发育异常的分子细胞学分析

摘要

To elucidate the role of WHIRLY2 in mitochondrial activity, overexpressing and knockout lines of Arabidopsis thaliana Co-lumbia were cultured. Phenotype analysis showed that silique of WHIRLY2-overexpressing line became dry, yellow and showed early senescence. Decreasing chloroplast number and increasing starch granule number were detected in pericarp cell of WHIRLY2 mutants compared to wild-type plant. Subsequent quantitative RT-PCR were conducted to analyze expression levels of genes related to silique development, energy metabolism, stability of mitochondrial genome and chloroplast differentiation in WHIRLY2 mutants. Results showed that transcription levels of genes involved in mitochondrial energy metabolism, including nad1, cytc382, rad50 and atp9, were increased in the silique of WHIRLY2-overexpressing lines. Mitochondrial genome structure and electrophoretic mobility shift as-says revealed that WHIRLY2 protein was able to bind to the 4×Telomere repeats of promoter region of nad1 and cytc382 genes. It suggested that WHIRLY2 may influence mitochondrial activity via gene expression of nad1 and cytc382 to regulate silique develop-ment in Arabidopsis.%通过构建拟南芥WHIRLY2基因超表达植株和鉴定这个基因的突变体，对WHIRLY2基因调控拟南芥角果的发育过程进行了分析。表型观察结果显示过量表达WHIRLY2基因的植株角果发育异常，出现变黄、干瘪和早衰现象，角果果皮细胞中叶绿体的数量明显比野生型少，叶绿体中淀粉粒的数目比野生型明显增多。进一步对角果发育、能量代谢、线粒体基因组稳定、叶绿体淀粉粒代谢相关基因的表达丰度进行qRT-PCR分析，发现WHIRLY2超表达植株角果中与线粒体能量代谢相关的nad1、cytc382、rad50和同时作用于线粒体和叶绿体的atp9基因的表达发生变化。通过线粒体基因组的结构分析和蛋白质凝胶迁移试验，发现WHIRLY2蛋白可以结合在线粒体基因组的nad1和cytc382基因启动子的4个端粒重复序列AAAATC-CCC上，推测WHIRLY2可能通过调控nad1和cytc382基因的转录从而参与角果发育的调控。