Cloning and gene expression analysis of two cDNA of cysteine proteinase genes involved in programmed cell death in the inner integument from developing seeds of Jatropha curcas L

摘要

In this paper, two cysteine proteinases were cloned from Jatropha curcas seeds. The full length cDNAs obtained from cloning of Jc-CysEP1 and Jc-CysEP2 genes were 1.516bp and 1500 pb, respectively. The Jc-CysEP1 contained a 1083bp open reading frame (ORF) coding for 360 amino acids. The JcCysEP1 protein sequence had an estimated native molecular weight of 36.89 kDa, with a predicted isoelectric point of 4.55. The average lengths of JcCysEP1 5' UTR and 3' UTR were 269 bp and 167bp, respectively. The Jc-CysEP2 contained a 1077 pb open reading frame (ORF) that encoded 358 amino acids. We also identified UTRs with lengths of 229 pb (5'UTR) and 194 pb (3'UTR). The Jc-CysEP2 sequence had a native molecular weight of 39.94 kDa, with a predicted isoelectric point of 6.19. Real-time PCR analyses of developing seeds (stages I-VII) showed that most cysteine proteinase genes were expressed at stage IV (middle stage) revealing peculiar spatio-temporal differences. JcCysEP2 was the cysteine proteinase gene with the highest expression in inner integument tissue, while JcCysEP1 was expressed in lower levels. Our results suggest that JcCysEP2 could be the major cysteine proteinase gene involved in PCD events in inner integument tissue, playing a critical role in PCD events during seed development, while Jc-CyEP1 and JcCysEP2 genes act cooperatively in stages IV-VII. JcCysEP2 is important to complete their participation in PCD until development of seeds.