Increasing plant density is an effective strategy for enhancing crop yield per unit land area.A key architectural trait for crops adapting to high planting density is a smaller leaf angle(LA).Previous studies have dem...Increasing plant density is an effective strategy for enhancing crop yield per unit land area.A key architectural trait for crops adapting to high planting density is a smaller leaf angle(LA).Previous studies have demonstrated that LG1,a SQUAMOSA BINDING PROTEIN(SBP)transcription factor,plays a critical role in LA establishment.However,the molecular mechanisms underlying the regulation of LG1 on LA formation remain largely unclear.In this study,we conduct comparative RNA-seq analysis of the preligule band(PLB)region of wild type and lg1 mutant leaves.Gene Ontology(GO)term enrichment analysis reveals enrichment of phytohormone pathways and transcription factors,including three auxin transporter genes ZmPIN1a,ZmPIN1b,and ZmPIN1c.Further molecular experiments demonstrate that LG1 can directly bind to the promoter region of these auxin transporter genes and activate their transcription.We also show that double and triple mutants of these ZmPINs genes exhibit varying degrees of auricle size reduction and thus decreased LA.On the contrary,overexpression of ZmPIN1a causes larger auricle and LA.Taken together,our findings establish a functional link between LG1 and auxin transport in regulating PLB formation and provide valuable targets for genetic improvement of LA for breeding high-density tolerant maize cultivars.展开更多
目的克隆并表达具有生物学活性的人层黏连蛋白α4链LG1组件(hum an lam in in alpha4 chain LG1 modu le,hLNα4LG1)蛋白。方法RT-PCR扩增hLNα4LG1的cDNA片段并将其插入pMD-18T载体进行测序。将测序片段亚克隆入原核表达载体pET-28 a并...目的克隆并表达具有生物学活性的人层黏连蛋白α4链LG1组件(hum an lam in in alpha4 chain LG1 modu le,hLNα4LG1)蛋白。方法RT-PCR扩增hLNα4LG1的cDNA片段并将其插入pMD-18T载体进行测序。将测序片段亚克隆入原核表达载体pET-28 a并在BL21(DE3)大肠杆菌中进行表达,W estern b lot对表达蛋白进行鉴定。用N i-NTA亲和柱对目的蛋白进行纯化,并通过细胞黏附与伸展实验对其生物学活性进行检测。结果成功克隆了hLNα4LG1的cDNA片段,SDS-PAGE及W estern b lot分析显示,经IPTG诱导后BL21(DE3)/pET28 a-LG1总蛋白中出现一条分子量为26×103的新蛋白带。与对照组相比,N i-NTA亲和层析纯化的目的蛋白有明显促进A549细胞伸展与黏附的作用。结论克隆并原核表达了hLNα4LG1蛋白,目的蛋白具有促进细胞黏附与伸展的活性,为深入研究hLNα4LG1的功能奠定了基础。展开更多
Leaf angle(LA)and tassel branch angle(TBA)are two important agronomic traits influencing maize plant architecture,thereby affecting its adaptability to high-density planting.Liguleless1(LG1)acts as a key regulator of ...Leaf angle(LA)and tassel branch angle(TBA)are two important agronomic traits influencing maize plant architecture,thereby affecting its adaptability to high-density planting.Liguleless1(LG1)acts as a key regulator of LA and TBA,yet its precise regulatory mechanism remains largely obscure.In this study,we have identified ZmTCP23,a teosinte branched1/CYCLOIDEA/proliferating cell factors(TCP)transcription factor that is highly expressed in tassel and leaf primordia,serving as a pivotal upstream transcriptional regulator of LG1.The functional loss of ZmTCP23 results in a significant reduction in both TBA and LA ranges.Moreover,in vitro and in vivo studies revealed that LG1 directly represses the expression of ZmXERICO1,a gene encoding an inhibitor of abscisic acid(ABA)degradation that can also influence LA and TBA upon overexpression.Additionally,ZmTCP23 physically interacts with the previously identified TBA regulator BAD1,forming a complex that co-activates the expression of LG1 via direct binding to its promoter.This dynamic duo established a positive feedback loop,mutually enhancing each other's expression within the tassels,and consequently influencing TBA.Our findings establish a ZmTCP23-LG1-ZmXERICO1 transcriptional regulatory cascade that orchestrates LA and TBA through influencing ABA content,and provide new targets for the genetic manipulation of LA and TBA for molecular breeding of high-density tolerant maize cultivars.展开更多
目的:利用巴斯德毕赤酵母表达系统获得重组的人层黏连蛋白LG1-3组件蛋白,为进一步研究LG1-3的结构和功能间的关系奠定基础。方法:利用RT-PCR从人胎盘组织总RNA中扩增LG1-3组件cDNA,并重组入pPICZαA,电转化入毕赤酵母GS115,利用甲醇诱...目的:利用巴斯德毕赤酵母表达系统获得重组的人层黏连蛋白LG1-3组件蛋白,为进一步研究LG1-3的结构和功能间的关系奠定基础。方法:利用RT-PCR从人胎盘组织总RNA中扩增LG1-3组件cDNA,并重组入pPICZαA,电转化入毕赤酵母GS115,利用甲醇诱导表达目的蛋白。结果和结论:获得了1 800 bp LG1-3组件cDNA,利用体外定点突变将SacⅠ酶切位点GAGCTC突变为GAGCGC,经SDS-PAGE和W estern印迹检测到相对分子质量为67.48×103的LG1-3组件蛋白的表达。展开更多
The LGS1(Large grain size 1)gene,also known as GS2/GL2/Os GRF4,is involved in regulating grain size and quality in rice,but the mechanism governing grain size has not been elucidated.We performed transcriptomic,proteo...The LGS1(Large grain size 1)gene,also known as GS2/GL2/Os GRF4,is involved in regulating grain size and quality in rice,but the mechanism governing grain size has not been elucidated.We performed transcriptomic,proteomic,and phosphoproteomic analyses of young rice panicles in Samba(a wild-type cultivar with extra-small grain)and NIL-LGS1(a nearly isogenic line of LGS1 with large grain in the Samba genetic background)at three developmental stages(4–6)to identify internal dynamic functional networks determining grain size that are mediated by LGS1.Differentially expressed proteins formed seven highly functionally correlated clusters.The concordant regulation of multiple functional clusters may be key features of the development of grain length in rice.In stage 5,16 and 24 phosphorylated proteins were significantly up-regulated and down-regulated,and dynamic phosphorylation events may play accessory roles in determining rice grain size by participating in protein–protein interaction networks.Transcriptomic analysis in stage 5 showed that differentially expressed alternative splicing events and dynamic gene regulatory networks based on 39 transcription factors and their highly correlated target genes might contribute to rice grain development.Integrative multilevel omics analysis suggested that the regulatory network at the transcriptional and posttranscriptional levels could be directly manifested at the translational level,and this analysis also suggested a regulatory mechanism,regulation of protein translation levels,in the biological process that extends from transcript to protein to the development of grain.Functional analysis suggested that biological processes including MAPK signaling,calcium signaling,cell proliferation,cell wall,energy metabolism,hormone pathway,and ubiquitin-proteasome pathway might be involved in LGS1-mediated regulation of grain length.Thus,LGS1-mediated regulation of grain size is affected by dynamic transcriptional,posttranscriptional,translational and posttranslational changes.展开更多
基金supported by the National Key Research and Development Program of China(2021YFF1000301)the National Natural Science Foundation of China(32472179,32130077,32201835)+2 种基金the Natural Science Foundation of Hebei Province(C2022407068)the Natural Science Foundation of Guangdong Province(2024A1515030237,2022A1515011002)the Hainan Yazhou Bay Seed Lab(B21HJ8101).
文摘Increasing plant density is an effective strategy for enhancing crop yield per unit land area.A key architectural trait for crops adapting to high planting density is a smaller leaf angle(LA).Previous studies have demonstrated that LG1,a SQUAMOSA BINDING PROTEIN(SBP)transcription factor,plays a critical role in LA establishment.However,the molecular mechanisms underlying the regulation of LG1 on LA formation remain largely unclear.In this study,we conduct comparative RNA-seq analysis of the preligule band(PLB)region of wild type and lg1 mutant leaves.Gene Ontology(GO)term enrichment analysis reveals enrichment of phytohormone pathways and transcription factors,including three auxin transporter genes ZmPIN1a,ZmPIN1b,and ZmPIN1c.Further molecular experiments demonstrate that LG1 can directly bind to the promoter region of these auxin transporter genes and activate their transcription.We also show that double and triple mutants of these ZmPINs genes exhibit varying degrees of auricle size reduction and thus decreased LA.On the contrary,overexpression of ZmPIN1a causes larger auricle and LA.Taken together,our findings establish a functional link between LG1 and auxin transport in regulating PLB formation and provide valuable targets for genetic improvement of LA for breeding high-density tolerant maize cultivars.
文摘目的克隆并表达具有生物学活性的人层黏连蛋白α4链LG1组件(hum an lam in in alpha4 chain LG1 modu le,hLNα4LG1)蛋白。方法RT-PCR扩增hLNα4LG1的cDNA片段并将其插入pMD-18T载体进行测序。将测序片段亚克隆入原核表达载体pET-28 a并在BL21(DE3)大肠杆菌中进行表达,W estern b lot对表达蛋白进行鉴定。用N i-NTA亲和柱对目的蛋白进行纯化,并通过细胞黏附与伸展实验对其生物学活性进行检测。结果成功克隆了hLNα4LG1的cDNA片段,SDS-PAGE及W estern b lot分析显示,经IPTG诱导后BL21(DE3)/pET28 a-LG1总蛋白中出现一条分子量为26×103的新蛋白带。与对照组相比,N i-NTA亲和层析纯化的目的蛋白有明显促进A549细胞伸展与黏附的作用。结论克隆并原核表达了hLNα4LG1蛋白,目的蛋白具有促进细胞黏附与伸展的活性,为深入研究hLNα4LG1的功能奠定了基础。
基金supported by projects from the National Key R&D Program of China(Grant No.2021YFF1000301)the National Natural Science Foundation of China(Grant Nos.31970202,32370349,32130077,and 32372209)the Natural Science Foundation of Guangdong Province(Grant No.2023A1515010493)。
文摘Leaf angle(LA)and tassel branch angle(TBA)are two important agronomic traits influencing maize plant architecture,thereby affecting its adaptability to high-density planting.Liguleless1(LG1)acts as a key regulator of LA and TBA,yet its precise regulatory mechanism remains largely obscure.In this study,we have identified ZmTCP23,a teosinte branched1/CYCLOIDEA/proliferating cell factors(TCP)transcription factor that is highly expressed in tassel and leaf primordia,serving as a pivotal upstream transcriptional regulator of LG1.The functional loss of ZmTCP23 results in a significant reduction in both TBA and LA ranges.Moreover,in vitro and in vivo studies revealed that LG1 directly represses the expression of ZmXERICO1,a gene encoding an inhibitor of abscisic acid(ABA)degradation that can also influence LA and TBA upon overexpression.Additionally,ZmTCP23 physically interacts with the previously identified TBA regulator BAD1,forming a complex that co-activates the expression of LG1 via direct binding to its promoter.This dynamic duo established a positive feedback loop,mutually enhancing each other's expression within the tassels,and consequently influencing TBA.Our findings establish a ZmTCP23-LG1-ZmXERICO1 transcriptional regulatory cascade that orchestrates LA and TBA through influencing ABA content,and provide new targets for the genetic manipulation of LA and TBA for molecular breeding of high-density tolerant maize cultivars.
文摘目的:利用巴斯德毕赤酵母表达系统获得重组的人层黏连蛋白LG1-3组件蛋白,为进一步研究LG1-3的结构和功能间的关系奠定基础。方法:利用RT-PCR从人胎盘组织总RNA中扩增LG1-3组件cDNA,并重组入pPICZαA,电转化入毕赤酵母GS115,利用甲醇诱导表达目的蛋白。结果和结论:获得了1 800 bp LG1-3组件cDNA,利用体外定点突变将SacⅠ酶切位点GAGCTC突变为GAGCGC,经SDS-PAGE和W estern印迹检测到相对分子质量为67.48×103的LG1-3组件蛋白的表达。
基金the National Key Research and Development Program of China(2017YFD0100103)the Seed Industry Innovation and Industrialization Project of Fujian Province(fjzycxny2017004,zycxny2021004)+1 种基金the Program on Technology of Fujian Province(2020NZ08016,2020N0049)the Open Program of State Key Laboratory of Rice Biology of China(170101)。
文摘The LGS1(Large grain size 1)gene,also known as GS2/GL2/Os GRF4,is involved in regulating grain size and quality in rice,but the mechanism governing grain size has not been elucidated.We performed transcriptomic,proteomic,and phosphoproteomic analyses of young rice panicles in Samba(a wild-type cultivar with extra-small grain)and NIL-LGS1(a nearly isogenic line of LGS1 with large grain in the Samba genetic background)at three developmental stages(4–6)to identify internal dynamic functional networks determining grain size that are mediated by LGS1.Differentially expressed proteins formed seven highly functionally correlated clusters.The concordant regulation of multiple functional clusters may be key features of the development of grain length in rice.In stage 5,16 and 24 phosphorylated proteins were significantly up-regulated and down-regulated,and dynamic phosphorylation events may play accessory roles in determining rice grain size by participating in protein–protein interaction networks.Transcriptomic analysis in stage 5 showed that differentially expressed alternative splicing events and dynamic gene regulatory networks based on 39 transcription factors and their highly correlated target genes might contribute to rice grain development.Integrative multilevel omics analysis suggested that the regulatory network at the transcriptional and posttranscriptional levels could be directly manifested at the translational level,and this analysis also suggested a regulatory mechanism,regulation of protein translation levels,in the biological process that extends from transcript to protein to the development of grain.Functional analysis suggested that biological processes including MAPK signaling,calcium signaling,cell proliferation,cell wall,energy metabolism,hormone pathway,and ubiquitin-proteasome pathway might be involved in LGS1-mediated regulation of grain length.Thus,LGS1-mediated regulation of grain size is affected by dynamic transcriptional,posttranscriptional,translational and posttranslational changes.
