Pepin K, Momose F, Ishida N, Nagata K
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Nagatsuta, Yokohama, Japan.
J Vet Med Sci. 2001 Feb;63(2):115-24. doi: 10.1292/jvms.63.115.
Heat shock protein 90 (Hsp90), a molecular chaperone, is ubiquitous and involved in numerous cellular processes. To contribute to the relatively small collection of vertebrate Hsp90 sequences in the gene data bank, we cloned and sequenced horse (Equus caballus) Hsp90 alpha and beta cDNAs. This enabled identification of horse-specific primers for development of a convenient PCR-based method that could monitor horse stress tolerance. We analyzed the sequence data comparatively and phylogenetically with other Hsp90 cDNA sequences, and identified vertebrate-specific and isoform-specific conserved regions to facilitate future molecular investigations of Hsp90 functions. We found 4 highly conserved regions to vertebrate Hsp90 exclusively and 27 amino acids conserved among but differing between Hsp90 alpha and Hsp90 beta sequences. Protein-based phylogenetic trees revealed high conservation between mammal species within Hsp90 alpha and beta clusters. Comparison of nucleotide and amino acid substitution levels suggests that horse Hsp90 beta has undergone strong purifying selection, while rat Hsp90 beta and hamster Hsp90 alpha have been positively selected. Surprisingly, fish Hsp90 alpha genes clearly clustered with Hsp90 beta genes, and no distinct placement of fish Hsp90 alpha protein was found. The Hsp90 alpha isoform is apparently the result of beta gene duplication. Our results highlight the importance of organism- and isoform-specific Hsp90 functional analyses in describing the role of Hsp90 in cells.
热休克蛋白90（Hsp90）是一种分子伴侣，广泛存在且参与众多细胞过程。为了丰富基因数据库中相对较少的脊椎动物Hsp90序列，我们克隆并测序了马（Equus caballus）的Hsp90α和β cDNA。这使得我们能够鉴定出马特异性引物，从而开发出一种便捷的基于PCR的方法来监测马的应激耐受性。我们将序列数据与其他Hsp90 cDNA序列进行了比较和系统发育分析，确定了脊椎动物特异性和亚型特异性保守区域，以促进未来对Hsp90功能的分子研究。我们发现了4个仅对脊椎动物Hsp90高度保守的区域，以及在Hsp90α和Hsp90β序列之间保守但不同的27个氨基酸。基于蛋白质的系统发育树显示，Hsp90α和β簇内的哺乳动物物种之间具有高度保守性。核苷酸和氨基酸替代水平的比较表明，马Hsp90β经历了强烈的纯化选择，而大鼠Hsp90β和仓鼠Hsp90α则受到了正选择。令人惊讶的是，鱼类Hsp90α基因明显与Hsp90β基因聚类，未发现鱼类Hsp90α蛋白的明显定位。Hsp90α亚型显然是β基因复制的结果。我们的结果突出了在描述Hsp90在细胞中的作用时进行生物体和亚型特异性Hsp90功能分析的重要性。
