Brassinosteroids （BRs） are perceived by transmembrane receptors and play vital roles in plant growth and development, as well as cell in responses to environmental stimuli. The transmemhrane receptor BRI1 can direct...Brassinosteroids （BRs） are perceived by transmembrane receptors and play vital roles in plant growth and development, as well as cell in responses to environmental stimuli. The transmemhrane receptor BRI1 can directly bind to brassinolide （BL）, and BAK1 interacts with BRI1 to enhance the BRI1-mediated BR signaling. Our previous studies indicated that a membrane steroid-binding protein 1 （MSBP1） could bind to BL in vitro and is negatively involved in BR signaling. To further elucidate the underlying mechanism, we here show that MSBPI specifically interacts with the extraeellular domain of BAK1 in vivo in a BL-independent manner. Suppressed cell expansion and BR responses by increased expression of MSBP1 can be recovered by overexpressing BAK1 or its intracellnlar kinase domain, sug- gesting that MSBP1 may suppress BR signaling through interacting with BAK1. Subcellular localization studies re- vealed that both MSBPI and BAK1 are localized to plasma membrane and endocytic vesicles and MSBP1 accelerates BAK1 endocytosis, which results in suppressed BR signaling by shifting the equilibrium of BAKI toward endosomes. Indeed, enhanced MSBP1 expression reduces the interaction between BRI1 and BAK1 in vivo, demonstrating that MSBP1 acts as a negative factor at an early step of the BR signaling pathway.展开更多
Objective:To investigate the effects of silencing BAK1 and BCL2 gene expression on proliferation,invasion and metastasis of hepatocellular carcinoma(HCC)HepG2 cells.Methods:30 HCC HepG2 cells were randomly divided int...Objective:To investigate the effects of silencing BAK1 and BCL2 gene expression on proliferation,invasion and metastasis of hepatocellular carcinoma(HCC)HepG2 cells.Methods:30 HCC HepG2 cells were randomly divided into groups and received the corresponding treatments,namely,control group,NC-siRNA group,BAK1-siRNA group,BCL2-siRNA group and BAK1+BCL2 group,with 6 strains in each group.ThenqRT-PCR,CCK8,Transwell chamber invasion and scratch assay were used to detect the expression,proliferation,invasion and metastasis of BAK1 and BCL2 genes in HepG2 cells.Results:The mRNA expression,cell proliferation rate,cell migration rate and cell invasion ability of BAK1 and BCL2 in HepG2 cells were lowest in the BAK1+BCL2 siRNA group,followed by BCL2-siRNA group,BAK1-siRNA group,NC-shRNA group and control group(P<0.05).The proliferation rate of HepG2 cells in the BAK1+BCL2 siRNA group decreased significantly with time(P<0.05).Conclusion:Silencing the expression of BAK1 and BCL2 genes can inhibit the proliferation and invasion of HCC HepG2 cells and promote their apoptosis.展开更多
Maize(Zea mays)is one of the most important crops in the world,but its yield and quality are seriously affected by diverse diseases.Identifying broad-spectrum resistance genes is crucial for developing effective strat...Maize(Zea mays)is one of the most important crops in the world,but its yield and quality are seriously affected by diverse diseases.Identifying broad-spectrum resistance genes is crucial for developing effective strategies to control the disease in maize.In a genome-wide study in maize,we identified a G-type lectin receptor kinase ZmLecRK1,as a new resistance protein against Pythium aphanidermatum,one of the causal pathogens of stalk rot in maize.Genetic analysis showed that the specific ZmLecRK1 allele can confer resistance to multiple pathogens in maize.The cell death and disease resistance phenotype mediated by the resistant variant of ZmLecRK1 requires the co-receptor ZmBAK1.A naturally occurring A404S variant in the extracellular domain of ZmLecRK1 determines the ZmLecRK1-ZmBAK1 interaction and the formation of ZmLecRK1-related protein complexes.Interestingly,the ZmLecRK1 susceptible variant was found to possess the amino acid S404 that is present in the ancestral variants of ZmLecRK1 and conserved among the majority of grass species,while the resistance variant of ZmLecRK1 with A404 is only present in a few maize inbred lines.Substitution of S by A at position 404 in ZmLecRK1-like proteins of sorghum and rice greatly enhances their ability to induce cell death.Further transcriptomic analysis reveals that ZmLecRK1 likely regulates gene expression related to the pathways in cell wall organization or biogenesis in response to pathogen infection.Taken together,these results suggest that the ZmLecRK1 resistance variant enhances its binding affinity to the co-receptor ZmBAK1,thereby enhancing the formation of active complexes for defense in maize.Our work highlights the biotechnological potential for generating disease-resistant crops by precisely modulating the activity of ZmLecRK1 and its homologs through targeted base editing.展开更多
文摘目的 探讨miR-125b通过靶向抑制Bak1表达影响人髓系白血病(AML)细胞增殖的机制及意义。方法选择miR-125b mimics、miR-125b inhibitor、miR-125b NC分别转染人AML细胞株THP-1(miR-125b mimics组、miR-125b inhibitor组、NC组)。采用荧光定量PCR检测miR-125b mRNA相对表达量, MTT法检测细胞增殖,流式细胞仪检测细胞凋亡,Western-blot法检测蛋白表达水平,利用生物信息学分析和荧光素酶双报告系统明确miR-125b的下游靶基因。结果 miR-125b mimics转染后THP-1细胞中miR-125b表达水平显著上调( P <0.05),miR-125b inhibitor转染后细胞中miR-125b水平则显著降低( P <0.05)。转染24 h、48 h后,MTT法检测显示miR-125b mimics组的细胞增殖活性显著低于miR-125b inhibitor组与NC组( P <0.05),细胞凋亡率显著高于miR-125b inhibitor组与NC组( P <0.05),后两组对比差异无统计学意义( P >0.05)。转染48 h后,Western-blot法检测显示miR-125b mimics组的 Bak1蛋白表达水平显著高于miR-125b inhibitor组与NC组( P <0.05),三组PI3K、AKT蛋白表达水平对比差异无统计学意义( P >0.05)。在HEK293细胞株中,miR-125b mimic转染48 h后可以显著降低Bak1萤火虫荧光素酶活性( P <0.05),而对于含有突变型载体的Bak1萤火虫荧光素酶活性无显著影响( P >0.05)。结论 过表达miR-125b可通过靶向抑制Bak1的活性,抑制人AML细胞株的增殖活性,促进细胞凋亡。
基金Acknowledgments This study was supported by the Chinese Academy of Sciences and National Natural Science Foundation of China （Grants 30425029, 30421001, 90717001）. We greatly thank Prof Hong Ma （Penn. State University, USA） for critical reading and writing improvement and Prof Nam-Hai Chua （The Rockefeller University, USA） for helpful comments. We thank the Salk Institute Genomic Analysis Laboratory for providing the sequence-indexed Arabidopsis T-DNA insertion mutants, and Prof Sheng Luan （University of California, Berkeley, USA） for providing the construct pATC940. We thank Prof Hong-Quan Yang （SIPPE, CAS） for providing LexA yeast two-hybrid system and Prof Zhi-Yong Wang （The Stanford University, USA） for providing the BRI1 antibody. We thank Mr Xiao-Shu Gao for the help on Confocal Laser Scanning Microscopy.
文摘Brassinosteroids （BRs） are perceived by transmembrane receptors and play vital roles in plant growth and development, as well as cell in responses to environmental stimuli. The transmemhrane receptor BRI1 can directly bind to brassinolide （BL）, and BAK1 interacts with BRI1 to enhance the BRI1-mediated BR signaling. Our previous studies indicated that a membrane steroid-binding protein 1 （MSBP1） could bind to BL in vitro and is negatively involved in BR signaling. To further elucidate the underlying mechanism, we here show that MSBPI specifically interacts with the extraeellular domain of BAK1 in vivo in a BL-independent manner. Suppressed cell expansion and BR responses by increased expression of MSBP1 can be recovered by overexpressing BAK1 or its intracellnlar kinase domain, sug- gesting that MSBP1 may suppress BR signaling through interacting with BAK1. Subcellular localization studies re- vealed that both MSBPI and BAK1 are localized to plasma membrane and endocytic vesicles and MSBP1 accelerates BAK1 endocytosis, which results in suppressed BR signaling by shifting the equilibrium of BAKI toward endosomes. Indeed, enhanced MSBP1 expression reduces the interaction between BRI1 and BAK1 in vivo, demonstrating that MSBP1 acts as a negative factor at an early step of the BR signaling pathway.
基金Science and technology introduction and innovation project of Xinjiang uygur autonomous region people's hospital(No.20170305).
文摘Objective:To investigate the effects of silencing BAK1 and BCL2 gene expression on proliferation,invasion and metastasis of hepatocellular carcinoma(HCC)HepG2 cells.Methods:30 HCC HepG2 cells were randomly divided into groups and received the corresponding treatments,namely,control group,NC-siRNA group,BAK1-siRNA group,BCL2-siRNA group and BAK1+BCL2 group,with 6 strains in each group.ThenqRT-PCR,CCK8,Transwell chamber invasion and scratch assay were used to detect the expression,proliferation,invasion and metastasis of BAK1 and BCL2 genes in HepG2 cells.Results:The mRNA expression,cell proliferation rate,cell migration rate and cell invasion ability of BAK1 and BCL2 in HepG2 cells were lowest in the BAK1+BCL2 siRNA group,followed by BCL2-siRNA group,BAK1-siRNA group,NC-shRNA group and control group(P<0.05).The proliferation rate of HepG2 cells in the BAK1+BCL2 siRNA group decreased significantly with time(P<0.05).Conclusion:Silencing the expression of BAK1 and BCL2 genes can inhibit the proliferation and invasion of HCC HepG2 cells and promote their apoptosis.
基金supported by Biological Breeding-National Science and Technology Major Project(no.2023ZD04070,W.Z.)the National Key Research and Development Program,Ministry of Science and Technology of China(no.2022YFD1201802,W.Z.)+1 种基金the National Natural Science Foundation of China(no.32472499,W.Z.)the Pinduoduo-China Agricultural University Research Fund(no.PC2023A01005,Y.-L.P.).
文摘Maize(Zea mays)is one of the most important crops in the world,but its yield and quality are seriously affected by diverse diseases.Identifying broad-spectrum resistance genes is crucial for developing effective strategies to control the disease in maize.In a genome-wide study in maize,we identified a G-type lectin receptor kinase ZmLecRK1,as a new resistance protein against Pythium aphanidermatum,one of the causal pathogens of stalk rot in maize.Genetic analysis showed that the specific ZmLecRK1 allele can confer resistance to multiple pathogens in maize.The cell death and disease resistance phenotype mediated by the resistant variant of ZmLecRK1 requires the co-receptor ZmBAK1.A naturally occurring A404S variant in the extracellular domain of ZmLecRK1 determines the ZmLecRK1-ZmBAK1 interaction and the formation of ZmLecRK1-related protein complexes.Interestingly,the ZmLecRK1 susceptible variant was found to possess the amino acid S404 that is present in the ancestral variants of ZmLecRK1 and conserved among the majority of grass species,while the resistance variant of ZmLecRK1 with A404 is only present in a few maize inbred lines.Substitution of S by A at position 404 in ZmLecRK1-like proteins of sorghum and rice greatly enhances their ability to induce cell death.Further transcriptomic analysis reveals that ZmLecRK1 likely regulates gene expression related to the pathways in cell wall organization or biogenesis in response to pathogen infection.Taken together,these results suggest that the ZmLecRK1 resistance variant enhances its binding affinity to the co-receptor ZmBAK1,thereby enhancing the formation of active complexes for defense in maize.Our work highlights the biotechnological potential for generating disease-resistant crops by precisely modulating the activity of ZmLecRK1 and its homologs through targeted base editing.
