Key Laboratory of Ministry of Education for Bio-resources and Eco-environment, School of Life Sciences, Sichuan University, Chengdu 610064, People's Republic of China.
Mol Biol Rep. 2011 Jun;38(5):3113-20. doi: 10.1007/s11033-010-9981-9. Epub 2010 Feb 3.
Three coding region cDNAs of duplicated PISTILLATA-like (PI-like) MADS-box genes, BnPI-1, BnPI-2 and BnPI-3, were isolated from B. napus by RT-PCR. The sequence analysis showed that the three PI cDNAs possessed 627, 627 and 625 nucleotides, respectively, and their nucleotide sequences had 96.49-98.72% similarity. Due to a deletion of two nucleotides, the protein sequence in the downstream of the frameshift site was altered in BnPI-3. Therefore, there were only 171 amino acids coded by BnPI-3, while there were 208 ones coded by BnPI-1 or BnPI-2. The deduced amino acid identity between BnPI-1 and BnPI-2 was 97.6% and the amino acid sequence of BnPI-1 and BnPI-2 shared 72.6% identity with BnPI-3. The deduced amino acid sequences of the coded proteins indicated high homology with the members of the PI family of MADS-box proteins. RT-PCR analysis showed that BnPI transcription was only detectable in petals and stamens. The yeast two-hybrid assays results showed that the three BnPI proteins exhibited different dimerization affinities with three BnAP3. BnPI-1 and BnPI-2 could form strong heterodimers with BnAP3. The dimerization affinity of BnPI-1 with BnAP3-4 is the strongest in all the combinations, while the affinity of BnPI-3 with BnAP3-4 is the weakest. The dimerization affinity to BnAP3-4 of BnPI-1 is 3.5 times of that of BnPI-3. The distinguished weak interaction to AP3 of BnPI-3 is probably due to the loss of the PI motif. The divergences of sequence and affinity of protein interaction might reflect some functional divergence of the three PI genes in B. napus.
从油菜中通过 RT-PCR 分离了三个重复的 PISTILLATA 样(PI 样)MADS 框基因 BnPI-1、BnPI-2 和 BnPI-3 的编码区 cDNA。序列分析表明,这三个 PI cDNA 分别具有 627、627 和 625 个核苷酸,其核苷酸序列具有 96.49-98.72%的相似性。由于两个核苷酸的缺失,移码位点下游的蛋白质序列在 BnPI-3 中发生了改变。因此,BnPI-3 仅编码 171 个氨基酸,而 BnPI-1 或 BnPI-2 编码 208 个氨基酸。BnPI-1 和 BnPI-2 之间的推导氨基酸同一性为 97.6%,BnPI-1 和 BnPI-2 的氨基酸序列与 BnPI-3 共享 72.6%的同一性。编码蛋白的推导氨基酸序列与 PI 家族的 MADS 框蛋白成员具有高度同源性。RT-PCR 分析表明,BnPI 转录仅在花瓣和雄蕊中检测到。酵母双杂交分析结果表明,这三个 BnPI 蛋白与三个 BnAP3 表现出不同的二聚化亲和力。BnPI-1 和 BnPI-2 可以与 BnAP3 形成强异源二聚体。BnPI-1 与 BnAP3-4 的二聚化亲和力在所有组合中最强,而 BnPI-3 与 BnAP3-4 的亲和力最弱。BnPI-1 与 BnAP3-4 的二聚化亲和力是 BnPI-3 的 3.5 倍。BnPI-3 对 AP3 的区分较弱的相互作用可能是由于 PI 基序的缺失。序列和蛋白相互作用亲和力的差异可能反映了油菜中这三个 PI 基因的一些功能分化。