彩叶草中一个富亮氨酸重复相关蛋白基因SsLRP的克隆和分析

Molecular Cloning and Analysis of a Novel Leucine-Rich Repeat Related Protein Gene,SsLRP,from Solenostemon scutellarioides (L.) Codd

根据彩叶草叶片小型EST库中一条具有1个富亮氨酸重复(leucine-rich repeat,LRR)结构域的EST序列,采用RACE与文库结合的方法,克隆了1个具有5个LRR结构域的全长cDNA,SsLRP(LRR-Related Protein)(GenBank登录号FJ787729)。SsLRP cDNA全长1024bp,包含一个657bp的ORF框,编码218个氨基酸。其5’-UTR区含有2个终止子TAG,3’-UTR区具有推测的加尾信号AATAAA。SsLRP蛋白N端具有的信号肽和保守的亮氨酸拉链结构域,具有5个保守的LRR结构域,多个磷酸化位点和N-糖基化位点。多序列比对和系统进化分析表明,SsLRP与番茄SlLRP同源性最高。二级结构和三级结构预测表明,SsLRP的功能可能与保守的LRR结构域密切相关,推测该基因可能参与蛋白间的相互作用与信号识别。RT-PCR分析表明,SsLRP与番茄SlLRP具有相似的表达模式,在正常植株的根、茎、叶和花中都有表达,在受菌核病感染植株的茎和叶中表达上调。

According to a EST sequence with a LRR （leucine-rich repeat） domain from Solenostemon scutellarioides, a novel leucine-rich repeat prelated rotein （LRP） gene with 5 LRRs, denoted SsLRP （GenBank Accession No. FJ787729）, was rapidly cloned using a strategy of RACE combined with cDNA library. The full length of SsLRP was 1 024 bp and contained a 657 bp open reading frame （ORF） encoding a 218 amino acid protein. Two stop codons （TAG） were found in 5＇-UTR and one possible polyadenylation signal, AATAAA, was found in 3＇-UTR. The SsLRP contained a predicted signal peptide and a conserved LZ （leucine zipper）domain in the N-terminal region, five conserved LRR domains, and multiple phosphorylation sites and N-linked glycosylation sites. Multiple sequence alignment and phylogenetic analysis showed that SsLRP shared high homology with tomato S1LRP, and belonged to the same subfamilies of LRPs. The prediction analysis of secondary structure and three dimensional structures of SsLRP showed that the possible function of the protein was decided by the multiple LRR domains, and involved in protein-protein interactions and signal perception. The RT-PCR analysis showed that SsLRP and SlLRP had the same expression pattern. SsLRP was expressed in all normal tissues, and up-regulated at high levels in diseased steam and leaves infected with Sclerotinia sclerotiorum.