Yu Fei, Park Sungsoon, Rodermel Steven R
Department of Genetics, Development and Cell Biology, Iowa State University, Ames, IA 50011, USA.
Plant J. 2004 Mar;37(6):864-76. doi: 10.1111/j.1365-313x.2003.02014.x.
The Arabidopsis At filamentation temperature sensitive (FtsH) metalloprotease gene family comprises 12 members (AtFtsH1-AtFtsH12), including three pairs of closely related genes that are targeted to chloroplasts (AtFtsH2 and AtFtsH8; AtFtsH1 and AtFtsH5; and AtFtsH7 and AtFtsH9). Mutations in AtFtsH5 (var1) and AtFtsH2 (var2) give rise to variegated plants with green- and white-sectored leaves. Cells in the green sectors contain morphologically normal chloroplasts, whereas cells in the white sectors are blocked in chloroplast biogenesis. A major question is how chloroplasts arise in cells that have a mutant genotype. We have found by two-dimensional (2-D) green gel and gel filtration analyses that AtFtsH2/VAR2 forms oligomeric complexes. Two bands in the 2-D green gels that correspond to AtFtsH5/VAR1 + AtFtsH1 and AtFtsH2/VAR2 + AtFtsH8 have been identified, and these bands are coordinately reduced in amount in var1 and var2 thylakoids that lack AtFtsH5/VAR1 and AtFtsH2/VAR2, respectively. These reductions are not because of alterations in transcript abundance. Overexpression of AtFtsH8 in var2-4 (a putative null allele) normalizes the variegation phenotype of the mutant and restores the two bands to their wild-type levels. These results suggest that AtFtsH8 is interchangeable with AtFtsH2/VAR2 in AtFtsH-containing oligomers, and that the two proteins have redundant functions. Consistent with this hypothesis, AtFtsH2 and AtFtsH8 have similar expression patterns, as monitored by promoter-beta-glucuronidase (GUS) fusion and RT-PCR experiments. Based on our findings, we propose that AtFtsH1, AtFtsH2/VAR2, AtFtsH5/VAR1, and AtFtsH8 interact to form oligomeric structures, and that subunit stoichiometry is controlled post-transcriptionally in var1 and var2, perhaps by turnover. A threshold model is presented to explain the pattern of variegation in var2 in which AtFtsH8 provides a compensating activity in the green sectors of the mutant.
拟南芥丝状温度敏感(FtsH)金属蛋白酶基因家族由12个成员(AtFtsH1 - AtFtsH12)组成,包括三对靶向叶绿体的密切相关基因(AtFtsH2和AtFtsH8;AtFtsH1和AtFtsH5;以及AtFtsH7和AtFtsH9)。AtFtsH5(var1)和AtFtsH2(var2)中的突变会导致植物叶片出现绿色和白色扇形区域的斑驳现象。绿色区域的细胞含有形态正常的叶绿体,而白色区域的细胞在叶绿体生物发生过程中受阻。一个主要问题是具有突变基因型的细胞中叶绿体是如何产生的。我们通过二维(2 - D)绿色凝胶和凝胶过滤分析发现AtFtsH2/VAR2形成寡聚复合物。在二维绿色凝胶中已鉴定出对应于AtFtsH5/VAR1 + AtFtsH1和AtFtsH2/VAR2 + AtFtsH8的两条带,并且在分别缺乏AtFtsH5/VAR1和AtFtsH2/VAR2的var1和var2类囊体中,这些带的量协同减少。这些减少并非由于转录本丰度的改变。在var2 - 4(一个推定的无效等位基因)中过表达AtFtsH8可使突变体的斑驳表型正常化,并使这两条带恢复到野生型水平。这些结果表明,在含AtFtsH的寡聚体中,AtFtsH8可与AtFtsH2/VAR2互换,并且这两种蛋白质具有冗余功能。与此假设一致,通过启动子 - β - 葡萄糖醛酸酶(GUS)融合和RT - PCR实验监测发现,AtFtsH2和AtFtsH8具有相似的表达模式。基于我们的发现,我们提出AtFtsH1、AtFtsH2/VAR2、AtFtsH5/VAR1和AtFtsH8相互作用形成寡聚结构,并且在var1和var2中,亚基化学计量在转录后受到控制,可能是通过周转实现的。提出了一个阈值模型来解释var2中的斑驳模式,其中AtFtsH8在突变体的绿色区域提供补偿活性。