Central Salt and Marine Chemicals Research Institute, Gujarat, India.
Mol Biotechnol. 2013 Jun;54(2):412-25. doi: 10.1007/s12033-012-9579-7.
The pathogenesis-related proteins have a broad spectrum of roles, ranging from seed germination, development to resistance. The PR-10 is a multigene family differing from other PR proteins in being intracellular, small and acidic with similar 3D structures. We have isolated JcPR-10a cDNA with an ORF of 483 bp from J. curcas, an important biofuel crop grown in the wastelands of India. JcPR-10a gets clustered with dicots in phylogenetic tree. The genomic organisation analysis of JcPR-10a revealed the presence of an intron at conserved 185 bp position. Transcript expression of JcPR-10a was upregulated in response to different stimuli such as NaCl, salicylic acid, methyl jasmonate and M. phaseolina. In response to SA and Macrophomina the transcript was found increased at 48 h, however, in case of NaCl and MeJa a strong induction was observed at 12 h which decreased at 48 h. We first time report the transcript up regulation of PR-10 gene by Macrophomina, a pathogen causing collar rot in Jatropha. The recombinant E. coli cells showed better growth in LB medium supplemented with NaCl, whereas growth of recombinant cells was inhibited in LB medium supplemented with KCl, mannitol, sorbitol, methyl jasmonate and salicylic acid. The JcPR-10a protein was overexpressed in E. coli cells, and was purified to homogeneity, the purified protein exhibited RNase and DNase activity. Furthermore, the protein also showed antifungal activity against Macrophomina, indicating that JcPR-10a can serve as an important candidate to engineer stress tolerance in Jatropha as well as other plants susceptible to collar rot by Macrophomina.
发病相关蛋白具有广泛的作用,从种子萌发、发育到抗性。PR-10 是一个多基因家族,与其他 PR 蛋白不同,它是细胞内的,体积小,酸性,具有相似的 3D 结构。我们从印度荒地种植的重要生物燃料作物麻疯树中分离到了 JcPR-10a cDNA,其 ORF 长 483bp。JcPR-10a 在系统发育树中与双子叶植物聚类。JcPR-10a 的基因组组织分析显示,在保守的 185bp 位置存在内含子。JcPR-10a 的转录表达在受到不同刺激时上调,如 NaCl、水杨酸、茉莉酸甲酯和 M 期。在对 SA 和 Macrophomina 的反应中,发现转录本在 48 小时增加,然而,在 NaCl 和 MeJa 的情况下,在 12 小时观察到强烈诱导,随后在 48 小时下降。我们首次报道了 PR-10 基因在 Macrophomina(一种引起麻疯树颈腐病的病原体)作用下的转录上调。重组大肠杆菌细胞在添加 NaCl 的 LB 培养基中表现出更好的生长,而在添加 KCl、甘露醇、山梨醇、茉莉酸甲酯和水杨酸的 LB 培养基中重组细胞的生长受到抑制。JcPR-10a 蛋白在大肠杆菌细胞中过表达,并被纯化至均一性,纯化的蛋白表现出 RNase 和 DNase 活性。此外,该蛋白还表现出对 Macrophomina 的抗真菌活性,表明 JcPR-10a 可以作为一个重要的候选基因,用于通过 Macrophomina 工程提高麻疯树以及其他易患颈腐病的植物的抗胁迫能力。