Nucleotide-binding site leucine-rich repeat receptors (NBS-LRR/NLRs) are crucial intracellular immune proteins in plants. Previous article reported a novel NLR protein SUT1 (SUPPRESSORS OF TOPP4-1, 1), which is involv...Nucleotide-binding site leucine-rich repeat receptors (NBS-LRR/NLRs) are crucial intracellular immune proteins in plants. Previous article reported a novel NLR protein SUT1 (SUPPRESSORS OF TOPP4-1, 1), which is involved in autoimmunity initiated by type one protein phosphatase 4 mutation (topp4-1) in Arabidopsis, however, its role in planta is still unclear. This study employed Nicotiana benthamiana, a model platform, to conduct an overall structural and functional analysis of SUT1 protein. The transient expression results revealed that SUT1 is a typical CNL (CC-NBS-LRR) receptor, both fluorescence data and biochemical results showed the protein is mainly anchored on the plasma membrane due to its N-terminal acylation site. Further truncation experiments announced that its CC (coiled-coil) domain possessed cell-death-inducing activity. The outcomes of point mutations analysis revealed that not only the CC domain, but also the full-length SUT1 protein, whose function and subcellular localization are influenced by highly conserved hydrophobic residues. These research outcomes provided favorable clues for elucidating the activation mechanism of SUT1.展开更多
根据抗病基因核苷酸结合位点(Nuc leotide b ind ing site,NBS)设计简并性引物,从陆地棉cDNA中进行RT-PCR扩增。获得含NBS保守域的EST,进一步用RACE技术和TAIL PCR技术获得其中1个EST的全长基因序列,并获得GHNBS基因的5′调控序列。此...根据抗病基因核苷酸结合位点(Nuc leotide b ind ing site,NBS)设计简并性引物,从陆地棉cDNA中进行RT-PCR扩增。获得含NBS保守域的EST,进一步用RACE技术和TAIL PCR技术获得其中1个EST的全长基因序列,并获得GHNBS基因的5′调控序列。此基因被命名为GHNBS。该基因的编码区长2 583 bp,编码861个氨基酸,GHNBS编码的氨基酸序列与拟南芥R基因具有28%的同源性。GHNBS与拟南芥的其他几个NBS-LRR基因比较发现,它们在保守区外的相似性相当低。Southern杂交和网上数据库搜索分析都表明GHNBS是1个寡拷贝基因。通过半定量RT-PCR分析发现GHNBS在棉花的蕾、花瓣、韧皮部、根及叶中均有表达且在根、叶中表达量比其它部位强,在木质部基本不表达。展开更多
Blackgram, an important legume crop, faces the constraint of Mungbean yellow mosaic India virus (MYMIV)-stress resulting in severe crop penalty. MYMIV-resistant plants exhibit incompatible response via a cognate CYR1 ...Blackgram, an important legume crop, faces the constraint of Mungbean yellow mosaic India virus (MYMIV)-stress resulting in severe crop penalty. MYMIV-resistant plants exhibit incompatible response via a cognate CYR1 gene-mediated interaction with virus effector molecule. In this study, we searched for the susceptible allele of the “R” gene in Cv. T9. Southern hybridization study confirmed presence of an allele in Cv. T9. However, transcripts of the CYR1 could not be detected either by RT-PCR or by Northern hybridization in Cv. T9 and also in other susceptible blackgram line. The allele was isolated, sequenced and referred as cyr1. In silico study revealed that cyr1 also encodes a CC-NBS-LRR protein like CYR1. However the CC domain of cyr1 is truncated by 128 amino acid residues indicating functional impairment with respect to the signal transduction after pathogen invasion. Comparative 3D structural modeling, hydrogen bonding and Van der Waals interaction studies revealed differences between CYR1 and cyr1. Lys519 and Thr490 present in the largest pockets of the CYR1 are the key interacting hotspots between CYR1 and MYMIV coat protein (CP). The weak Van der Waals interactions and intermolecular hydrogen bonding between cyr1 and CP confers less stability to the molecular recognition complex, unlike CYR1. Thus, the present investigation revealed Cv. T9 shows compatible interaction with MYMIV due to the truncation in the cyr1 sequence and consequent structural difference in the N-terminal of CC-domain.展开更多
文摘Nucleotide-binding site leucine-rich repeat receptors (NBS-LRR/NLRs) are crucial intracellular immune proteins in plants. Previous article reported a novel NLR protein SUT1 (SUPPRESSORS OF TOPP4-1, 1), which is involved in autoimmunity initiated by type one protein phosphatase 4 mutation (topp4-1) in Arabidopsis, however, its role in planta is still unclear. This study employed Nicotiana benthamiana, a model platform, to conduct an overall structural and functional analysis of SUT1 protein. The transient expression results revealed that SUT1 is a typical CNL (CC-NBS-LRR) receptor, both fluorescence data and biochemical results showed the protein is mainly anchored on the plasma membrane due to its N-terminal acylation site. Further truncation experiments announced that its CC (coiled-coil) domain possessed cell-death-inducing activity. The outcomes of point mutations analysis revealed that not only the CC domain, but also the full-length SUT1 protein, whose function and subcellular localization are influenced by highly conserved hydrophobic residues. These research outcomes provided favorable clues for elucidating the activation mechanism of SUT1.
文摘根据抗病基因核苷酸结合位点(Nuc leotide b ind ing site,NBS)设计简并性引物,从陆地棉cDNA中进行RT-PCR扩增。获得含NBS保守域的EST,进一步用RACE技术和TAIL PCR技术获得其中1个EST的全长基因序列,并获得GHNBS基因的5′调控序列。此基因被命名为GHNBS。该基因的编码区长2 583 bp,编码861个氨基酸,GHNBS编码的氨基酸序列与拟南芥R基因具有28%的同源性。GHNBS与拟南芥的其他几个NBS-LRR基因比较发现,它们在保守区外的相似性相当低。Southern杂交和网上数据库搜索分析都表明GHNBS是1个寡拷贝基因。通过半定量RT-PCR分析发现GHNBS在棉花的蕾、花瓣、韧皮部、根及叶中均有表达且在根、叶中表达量比其它部位强,在木质部基本不表达。
文摘Blackgram, an important legume crop, faces the constraint of Mungbean yellow mosaic India virus (MYMIV)-stress resulting in severe crop penalty. MYMIV-resistant plants exhibit incompatible response via a cognate CYR1 gene-mediated interaction with virus effector molecule. In this study, we searched for the susceptible allele of the “R” gene in Cv. T9. Southern hybridization study confirmed presence of an allele in Cv. T9. However, transcripts of the CYR1 could not be detected either by RT-PCR or by Northern hybridization in Cv. T9 and also in other susceptible blackgram line. The allele was isolated, sequenced and referred as cyr1. In silico study revealed that cyr1 also encodes a CC-NBS-LRR protein like CYR1. However the CC domain of cyr1 is truncated by 128 amino acid residues indicating functional impairment with respect to the signal transduction after pathogen invasion. Comparative 3D structural modeling, hydrogen bonding and Van der Waals interaction studies revealed differences between CYR1 and cyr1. Lys519 and Thr490 present in the largest pockets of the CYR1 are the key interacting hotspots between CYR1 and MYMIV coat protein (CP). The weak Van der Waals interactions and intermolecular hydrogen bonding between cyr1 and CP confers less stability to the molecular recognition complex, unlike CYR1. Thus, the present investigation revealed Cv. T9 shows compatible interaction with MYMIV due to the truncation in the cyr1 sequence and consequent structural difference in the N-terminal of CC-domain.
