Wang Si-Jia, Wu Ming-Jie, Chen Xiang-Jun, Jiang Yan, Yan Yong-Bin
Key Laboratory of Bio-Resources and Eco-Environment of MOE, College of Life Science, Sichuan University, Chengdu 610064, China.
State Key Laboratory of Biomembrane and Membrane Biotechnology, Department of Biological Sciences and Biotechnology, Tsinghua University, Beijing 100084, China.
Int J Mol Sci. 2012;13(7):7963-7979. doi: 10.3390/ijms13077963. Epub 2012 Jun 27.
Heat shock protein 90 (Hsp90) is a molecular chaperone highly conserved across the species from prokaryotes to eukaryotes. Hsp90 is essential for cell viability under all growth conditions and is proposed to act as a hub of the signaling network and protein homeostasis of the eukaryotic cells. By interacting with various client proteins, Hsp90 is involved in diverse physiological processes such as signal transduction, cell mobility, heat shock response and osmotic stress response. In this research, we cloned the dshsp90 gene encoding a polypeptide composed of 696 amino acids from the halotolerant unicellular green algae Dunaliella salina. Sequence alignment indicated that DsHsp90 belonged to the cytosolic Hsp90A family. Further biophysical and biochemical studies of the recombinant protein revealed that DsHsp90 possessed ATPase activity and existed as a dimer with similar percentages of secondary structures to those well-studied Hsp90As. Analysis of the nucleotide sequence of the cloned genomic DNA fragment indicated that dshsp90 contained 21 exons interrupted by 20 introns, which is much more complicated than the other plant hsp90 genes. The promoter region of dshsp90 contained putative cis-acting stress responsive elements and binding sites of transcriptional factors that respond to heat shock and salt stress. Further experimental research confirmed that dshsp90 was upregulated quickly by heat and salt shock in the D. salina cells. These findings suggested that dshsp90 might serve as a component of the early response system of the D. salina cells against environmental stresses.
热休克蛋白90(Hsp90)是一种分子伴侣,从原核生物到真核生物在整个物种中高度保守。Hsp90在所有生长条件下对细胞活力至关重要,被认为是真核细胞信号网络和蛋白质稳态的枢纽。通过与各种客户蛋白相互作用,Hsp90参与多种生理过程,如信号转导、细胞迁移、热休克反应和渗透应激反应。在本研究中,我们从耐盐单细胞绿藻杜氏盐藻中克隆了编码由696个氨基酸组成的多肽的dshsp90基因。序列比对表明DsHsp90属于胞质Hsp90A家族。对重组蛋白的进一步生物物理和生化研究表明,DsHsp90具有ATP酶活性,以二聚体形式存在,其二级结构百分比与那些已深入研究的Hsp90A相似。对克隆的基因组DNA片段核苷酸序列的分析表明,dshsp90包含21个外显子,被20个内含子打断,这比其他植物hsp90基因复杂得多。dshsp90的启动子区域包含假定的顺式作用应激反应元件以及响应热休克和盐胁迫的转录因子结合位点。进一步的实验研究证实,dshsp90在杜氏盐藻细胞中受到热和盐胁迫后迅速上调。这些发现表明,dshsp90可能是杜氏盐藻细胞针对环境胁迫的早期反应系统的一个组成部分。