Immunolocalization of a Protein Disulfide Isomerase to Arabidopsis thaliana Chloroplasts and Its Association with Starch Biogenesis
Immunolocalization of a Protein Disulfide Isomerase to Arabidopsis thaliana Chloroplasts and Its Association with Starch Biogenesis
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DOI：
10.1086/498071
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发表时间：
2006-01
影响因子：
2.3
通讯作者：
Dongping Lu;D. Christopher
中科院分区：
文献类型：
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作者：
Dongping Lu;D. Christopher
蛋白质二硫键异构酶（PDIs）在蛋白质折叠和氧化还原信号转导中起重要作用。我们使用抗血清产生的叶绿体PDI，RB 60，从单细胞衣藻莱茵衣藻的PDI的分析，从维管植物拟南芥。免疫分析进行重组PDI和PDI梯度纯化的叶绿体亚组分，并通过荧光显微镜和透射电子显微镜的拟南芥叶片。通过免疫印迹分析总细胞蛋白和由pdi cDNA体外转录/翻译产生的65-kDa PDI，检测到丰富的65 kDa免疫相关PDI同种型。通过显微镜分析证实，PDI位于叶片的叶绿体中，主要与淀粉粒的基质边缘相关，特别是发育中的淀粉粒一端的基质-淀粉界面。在光诱导的叶绿体发育过程中，相对于黄化幼苗，光中pdi mRNA水平增加了50%。在黄化幼苗中未检测到65-kDa PDI蛋白，但在光照期间显著增加。相反，成熟的绿色叶绿体的暗适应并没有显着影响PDI水平。我们提出了植物中氧化还原调节PDI的新作用，即与植物叶片中的瞬时淀粉粒生物合成相关。
Protein disulfide isomerases (PDIs) play important roles in protein folding and redox signaling. We used antiserum generated against the chloroplast PDI, RB60, from the unicellular alga Chlamydomonas reinhardtii for the analysis of a PDI from the vascular plant Arabidopsis thaliana. Immunoanalysis was conducted on recombinant PDI and percoll gradient–purified chloroplast subfractions and via fluorescence microscopy and transmission electron microscopy of Arabidopsis leaves. An abundant immuno‐related PDI isoform of 65 kDa was detected via immunoblot analysis of total cell proteins and a 65‐kDa PDI produced by in vitro transcription/translation of a pdi cDNA. Analysis via microscopy verified that the PDI was located in chloroplasts of leaves, being mainly associated with the stromal fringes of the starch grain, in particular, the stromal‐starch interface at one end of the developing starch grain. During light‐induced chloroplast development, pdi mRNA levels increased by 50% in the light relative to etiolated seedlings. The 65‐kDa PDI protein was undetected in etiolated seedlings but increased significantly during illumination. In contrast, dark adaptation of mature green chloroplasts did not significantly affect PDI levels. We propose a new role for the redox‐regulatory PDIs in plants, that of being associated with transitory starch granule biogenesis in plant leaves.