Functional characterization of Arabidopsis calreticulin1a:: A key alleviator of endoplasmic reticulum stress
Functional characterization of Arabidopsis calreticulin1a:: A key alleviator of endoplasmic reticulum stress
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DOI：
10.1093/pcp/pcn065
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发表时间：
2008-06-01
影响因子：
4.9
通讯作者：
Sommarin, Marianne
中科院分区：
文献类型：
--
作者：
Christensen, Anna;Svensson, Karin;Sommarin, Marianne
钙网蛋白分子伴侣在动物细胞内质网中的钙信号转导和蛋白质折叠等过程中发挥重要作用。虽然钙网蛋白的功能在动物中得到了很好的表征，但在植物中只有间接的证据。为了增加我们对植物钙网蛋白的理解，我们将拟南芥中的一种同种型AtCRT 1a引入钙网蛋白缺陷（crt（-/-））小鼠胚胎成纤维细胞中。由于钙网蛋白缺乏，小鼠crt（-/-）成纤维细胞内质网中Ca ~（2+）水平降低，蛋白质折叠能力受损。AtCRT 1a在小鼠crt（-/-）成纤维细胞中的表达挽救了这些表型，即AtCRT 1a恢复了小鼠crt（-/-）成纤维细胞ER中的Ca 2+保持能力和伴侣蛋白功能，表明动物分选机制对植物蛋白也有功能，并且基本钙网蛋白功能在整个界中是保守的。使用β-葡萄糖醛酸酶（GUS）AtCRT 1a启动子构建的表达分析显示，CRT 1a在根尖，花组织和与维管束的高表达。为了评估AtCRT 1a在植物中的影响，我们产生了Atcrt 1a突变体植物。Atcrt 1a突变体表现出对药物衣霉素的敏感性增加，衣霉素是未折叠蛋白反应的诱导剂。因此，我们得出结论，AtCRT 1a是衣霉素诱导的未折叠蛋白反应的一个放大器，并建议使用小鼠crt（-/-）成纤维细胞作为钙网蛋白表达系统可能被证明是有用的，以评估不同物种的钙网蛋白的功能。
The chaperone calreticulin plays important roles in a variety of processes in the endoplasmic reticulum (ER) of animal cells, such as Ca2+ signaling and protein folding. Although the functions of calreticulin are well characterized in animals, only indirect evidence is available for plants. To increase our understanding of plant calreticulins we introduced one of the Arabidopsis isoforms, AtCRT1a, into calreticulin-deficient (crt(-/-)) mouse embryonic fibroblasts. As a result of calreticulin deficiency, the mouse crt(-/-) fibroblasts have decreased levels of Ca2+ in the ER and impaired protein folding abilities. Expression of the AtCRT1a in mouse crt(-/-) fibroblasts rescued these phenotypes, i.e. AtCRT1a restored the Ca2+-holding capacity and chaperone functions in the ER of the mouse crt(-/-) fibroblasts, demonstrating that the animal sorting machinery was also functional for a plant protein, and that basic calreticulin functions are conserved across the Kingdoms. Expression analyses using a beta-glucuronidase (GUS)AtCRT1a promoter construct revealed high expression of CRT1a in root tips, floral tissues and in association with vascular bundles. To assess the impact of AtCRT1a in planta, we generated Atcrt1a mutant plants. The Atcrt1a mutants exhibited increased sensitivity to the drug tunicamycin, an inducer of the unfolded protein response. We therefore conclude that AtCRT1a is an alleviator of the tunicamycin-induced unfolded protein response, and propose that the use of the mouse crt(-/-) fibroblasts as a calreticulin expression system may prove useful to assess functionalities of calreticulins from different species.