Bhadula S K, Elthon T E, Habben J E, Helentjaris T G, Jiao S, Ristic Z
Department of Biology, University of South Dakota, Vermillion 57069, USA.
Planta. 2001 Feb;212(3):359-66. doi: 10.1007/s004250000416.
A heat-tolerant maize (Zea mays L.) line, ZPBL 1304, synthesizes a unique set of five heat-shock polypeptides of 45 kDa. Previous studies suggested that these polypeptides might play a role in the development of thermotolerance in maize (Ristic et al., 1996, J. Plant Physiol. 149:424-432; Ristic et al., 1998, J. Plant Physiol. 153:497-505). In the present study, we isolated these polypeptides, sequenced them, and investigated their subcellular distribution and origin. Of the five polypeptides of 45 kDa, three polypeptides, including the two most abundant ones, yielded amino acid sequences similar to the chloroplast and bacterial protein synthesis elongation factor (EF-Tu). This was further confirmed using an antibody raised against maize EF-Tu, which showed a very strong reaction with the 45-kDa heatshock protein(s). Studies on subcellular distribution and origin revealed that the 45-kDa polypeptides were localized to the chloroplasts, and were likely of nuclear origin. A full-length maize EF-Tu cDNA (Zmeftu1), previously isolated from the B73 line of maize, was used as a probe for northern blot analysis of RNA extracted from the ZPBL 1304 maize line (the nucleotide and deduced amino acid sequences of Zmeftu1 are 88% identical to the rice EF-Tu sequence). Northern blots showed a 1.85-fold increase in steady-state levels of EF-Tu mRNA during heat stress. An increase in EF-Tu transcript levels during heat stress was accompanied by increased levels of the EF-Tu protein. Isolated chloroplasts from heat-stressed plants also had higher levels of EF-Tu as compared to control chloroplasts. The maize EF-Tu polypeptides showed > 80% sequence similarity with the bacterial EF-Tu, which has recently been shown to function as a molecular chaperone and to play a role in the protection of other proteins from thermal denaturation (Caldas et al., 1998, J. Biol. Chem. 273:11478-11482). It is hypothesized that chloroplast EF-Tu of the ZPBL 1304 maize line plays an important role in the development of thermotolerance.
一个耐热玉米（Zea mays L.）品系ZPBL 1304能合成一套独特的五种45 kDa热激多肽。先前的研究表明，这些多肽可能在玉米耐热性的形成中发挥作用（里斯蒂克等人，1996年，《植物生理学杂志》149:424 - 432；里斯蒂克等人，1998年，《植物生理学杂志》153:497 - 505）。在本研究中，我们分离了这些多肽，测定了它们的序列，并研究了它们的亚细胞分布和来源。在这五种45 kDa的多肽中，有三种多肽，包括含量最高的两种，其氨基酸序列与叶绿体和细菌蛋白质合成延伸因子（EF - Tu）相似。用针对玉米EF - Tu产生的抗体进一步证实了这一点，该抗体与45 kDa热激蛋白有非常强烈的反应。对亚细胞分布和来源的研究表明，45 kDa的多肽定位于叶绿体，并且可能起源于细胞核。一个先前从玉米B73品系中分离得到的玉米EF - Tu全长cDNA（Zmeftu1），被用作探针，对从ZPBL 1304玉米品系中提取的RNA进行Northern印迹分析（Zmeftu1的核苷酸和推导氨基酸序列与水稻EF - Tu序列的同源性为88%）。Northern印迹显示，热胁迫期间EF - Tu mRNA的稳态水平增加了1.85倍。热胁迫期间EF - Tu转录本水平的增加伴随着EF - Tu蛋白水平的升高。与对照叶绿体相比，热胁迫植物分离出的叶绿体中EF - Tu水平也更高。玉米EF - Tu多肽与细菌EF - Tu的序列相似性> 80%，最近已证明细菌EF - Tu可作为分子伴侣发挥作用，并在保护其他蛋白质免受热变性方面发挥作用（卡尔达斯等人，1998年，《生物化学杂志》273:11478 - 11482）。据推测，ZPBL 1304玉米品系的叶绿体EF - Tu在耐热性的形成中起重要作用。
