Identification and stress-induced expression of three 3尾-hydroxysteroid dehydrogenases from Erysimum crepidifolium Rchb. and their putative role in cardenolide biosynthesis

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• 作者：Jennifer Munkert^a ; Mona Ernst^a ; Frieder Mü ; ller-Uri^a ; Wolfgang Kreis^a ; ^b ; ^{wolfgang.kreis@fau.de" class="auth_mail}
• 关键词：Erysimum crepidifolium ; Brassicaceae ; Wallflower ; Cardiac glycosides ; Gene expression ; Stress ; Cardenolide biosynthesis ; 3尾-Hydroxysteroid dehydrogenase
• 刊名：Phytochemistry
• 出版年：April, 2014
• 年：2014
• 卷：100
• 期：Complete
• 页码：26-33
• 全文大小：693 K

文摘

3尾-Hydroxysteroid dehydrogenases (3尾HSD) are supposed to be involved in cardenolide biosynthesis in plants. Erysimum crepidifolium Rchb., a member of the Brassicaceae accumulating cardenolides, is a close relative to Arabidopsis thaliana. Full length cDNAs encoding for three individual 3尾HSDs (EcHSD1, EcHSD2, EcHSD3) were isolated from E. crepidifolium leaves. EcHSD1 and EcHSD2 encode proteins assembled from 257 amino acids whereas EcHSD3 encodes a protein assembled from 260 amino acids. All three proteins qualify as members of the short-chain dehydrogenases/reductases family of proteins (SDRs). EcHSD1 and EcHSD2 shared a high amino acid sequence identity of about 86% and 91% with putative 3尾HSDs of A. thaliana ( and ). EcHSD3 showed high homology to the A. thaliana SDRs (74%) and (81%). All three EcHSD genes were expressed in Escherichia coli and the recombinant enzymes were characterized biochemically. All three recombinant EcHSDs catalyzed the dehydrogenation of pregnenolone and the 3-reduction of 5伪/尾-pregnane-3,20-dione when NAD and NADH were used as cosubstrates, respectively. After exposure to different stress conditions, no increased transcription was seen for EcHSD1 whereas EcHSD2 was expressed four times higher under osmotic stress than under control conditions. EcHSD3 expression was 10 times and 6 times higher after osmotic stress and MeJA treatment, respectively, than in controls.