Exosomes(EXOs)play an important role in the progression of breast cancer.EXOs,with a diameter of approximately 100 nm,have a simple structure but diverse functions,and can affect the development of breast cancer throu...Exosomes(EXOs)play an important role in the progression of breast cancer.EXOs,with a diameter of approximately 100 nm,have a simple structure but diverse functions,and can affect the development of breast cancer through signal transduction and molecular transfer,etc.Angiogenesis provides nutrients for the growth and metastasis of breast cancer and is a crucial part of tumor progression.The mechanism of tumor angiogenesis is complex.The VEGF/VEGFR pathway promotes angiogenesis by regulating the activities of ECs.Hypoxia,a common feature in the tumor microenvironment,as a key regulator,can affect angiogenesis in multiple aspects such as the transfer of miRNAs in EXOs,protein transport,extracellular matrix regulation,and metabolic adaptation.The Notch pathway has a bidirectional regulatory role in breast cancer angiogenesis,and different molecules can promote or inhibit angiogenesis.EXOs secreted by breast cancer cells are rich in angiogenic factors.Components such as proteins and nucleic acids in EXOs can affect the functions and behaviors of vascular ECs,thereby influencing breast cancer angiogenesis.Research on the mechanisms of EXOs in breast cancer angiogenesis is of great significance for tumor treatment.EXOs are expected to become biomarkers for breast cancer diagnosis/prognosis.This research provides potential targets for in-depth understanding of the biological characteristics of breast cancer and the development of new treatment strategies.展开更多
As a major subunit of the exocyst complex,members of the EXO70 family have mainly been shown to play roles in cell polarity and morphogenesis in Arabidopsis,but their roles in plant endosymbiosis,such as with arbuscul...As a major subunit of the exocyst complex,members of the EXO70 family have mainly been shown to play roles in cell polarity and morphogenesis in Arabidopsis,but their roles in plant endosymbiosis,such as with arbuscular mycorrhizal fungi(AMF),have rarely been reported.Here,using knockout and overexpression lines,we show that OsEXO70L2,which encodes a divergent EXO70 protein in rice,controls the number of primary roots and is essential for large lateral root formation.Furthermore,the OsEXO70L2 mutant sr1 displayed rare internal AMF hyphaeand no arbuscules.We also found that AMF sporulation can occur in roots despite low colonization and that AMF colonization and sporulation are modulated by photoperiod and co-culture with clover.Finally,genes related to auxin homeostasis were found to be affected in the OsEXO70L2 knockout or overexpression lines,suggesting that auxin is at least partly responsible for the phenotypes.This study provides new perspectives on the role of the exocyst complex during root development and AM in rice.展开更多
【目的】深入研究甘蓝自交不亲和信号传导关键元件S-位点受体激酶SRK与臂重复蛋白ARC1及ARC1与Exo70A1间相互识别的分子机理,鉴定SRK-ARC1及ARC1-Exo70A1之间的互作区段,并分析其作用强度,明确蛋白间互作功能域。【方法】通过生物信息...【目的】深入研究甘蓝自交不亲和信号传导关键元件S-位点受体激酶SRK与臂重复蛋白ARC1及ARC1与Exo70A1间相互识别的分子机理,鉴定SRK-ARC1及ARC1-Exo70A1之间的互作区段,并分析其作用强度,明确蛋白间互作功能域。【方法】通过生物信息学分析得到蛋白功能域,根据分析结果以典型的自交不亲和结球甘蓝E1为材料分别扩增SRK、ARC1和Exo70A1含不同功能域的截短体片段,利用分子克隆技术将SRK激酶域(SRKj)及其截短体SRKjΔ1—SRKjΔ4,Exo70A1全长及其截短体Exo70A1Δ1—Exo70A1Δ3的编码序列分别亚克隆至p GADT7(AD)质粒,将ARC1及其截短体ARC1Δ1—ARC1Δ8的编码序列分别亚克隆至载体p GBKT7(BD)质粒。用PEG/Li Ac法将获得的AD和BD重组质粒两两组合分别共转化到酵母AH109感受态中,观察融合菌株在SD/-Leu-Trp-His-Ade/X-α-gal/25 m M 3-AT平板上的菌落生长情况和颜色变化情况,进一步测定其β-半乳糖苷酶活性。最后通过原核表达体外孵育检测蛋白质相互作用的方法对SRK-ARC1及ARC1-Exo70A1的相互作用进行验证。【结果】DNA测序和内切酶分析显示成功构建18个酵母双杂交表达载体,且无自激活能力。在SRK-ARC1的10个试验组合中,只有ARC1Δ4、ARC1Δ8、ARC1与SRKj组合的融合菌株在SD/-Leu-Trp-His-Ade/X-α-gal/25 m M 3-AT培养基上长出蓝色菌落,激活报告基因HIS3、ADE2和MEL1。随着SRKj或ARC1截短体片段的延长,二者的β-半乳糖苷酶活性逐渐增加,其中,ARC1Δ4与SRKj组合的β-半乳糖苷酶活性最高(酶活为15.98)。在ARC1-Exo70A1 16个试验组合中,Exo70A1Δ3与ARC1Δ1Δ3都相互作用,其融合菌株在SD/-Leu-Trp-His-Ade/X-α-gal/25 m M 3-AT培养基上长出蓝色菌落,激活报告基因HIS3、ADE2和MEL1。随着ARC1或Exo70A1截短体片段的延长,二者的β-半乳糖苷酶活性呈现先增加后降低的趋势,其中ARC1Δ2与Exo70A1Δ3组合的β-半乳糖苷酶活性最大(酶活性为25.07)。说明ARC1的N端和Exo70A1的N端发生了互作,而ARC1的C端、全长与Exo70A1都不发生互作。体外表达检测蛋白相互作用发现,SRKj与ARC1Δ4、ARC1Δ2与Exo70A1Δ3均可以直接发生相互作用。【结论】SRK的激酶域(SRKj)与ARC1的C端臂重复区发生互作,缩短SRK激酶域中的任何结构域或者缩短ARC1的臂重复区,二者都不会发生互作。ARC1的亮氨酸拉链和蜷曲螺旋与Exo70A1的N端结构域(去除pfam Exo70A1域)介导了二者的相互作用。SRK-ARC1的作用力强度小于ARC1-Exo70A1的作用力强度。展开更多
文摘Exosomes(EXOs)play an important role in the progression of breast cancer.EXOs,with a diameter of approximately 100 nm,have a simple structure but diverse functions,and can affect the development of breast cancer through signal transduction and molecular transfer,etc.Angiogenesis provides nutrients for the growth and metastasis of breast cancer and is a crucial part of tumor progression.The mechanism of tumor angiogenesis is complex.The VEGF/VEGFR pathway promotes angiogenesis by regulating the activities of ECs.Hypoxia,a common feature in the tumor microenvironment,as a key regulator,can affect angiogenesis in multiple aspects such as the transfer of miRNAs in EXOs,protein transport,extracellular matrix regulation,and metabolic adaptation.The Notch pathway has a bidirectional regulatory role in breast cancer angiogenesis,and different molecules can promote or inhibit angiogenesis.EXOs secreted by breast cancer cells are rich in angiogenic factors.Components such as proteins and nucleic acids in EXOs can affect the functions and behaviors of vascular ECs,thereby influencing breast cancer angiogenesis.Research on the mechanisms of EXOs in breast cancer angiogenesis is of great significance for tumor treatment.EXOs are expected to become biomarkers for breast cancer diagnosis/prognosis.This research provides potential targets for in-depth understanding of the biological characteristics of breast cancer and the development of new treatment strategies.
基金supported by the National Natural Science Foundation of China(32171970)the Chongqing Talent Program,China(cstc2022ycjh-bgzxm0073)+1 种基金the Natural Science Foundation of Chongqing,China(cstc2021jcyjcxtt X0004)the Rice Innovative Research Team of Chongqing Modern Agricultural Industrial Technology System,China(CQMAITS202301)。
文摘As a major subunit of the exocyst complex,members of the EXO70 family have mainly been shown to play roles in cell polarity and morphogenesis in Arabidopsis,but their roles in plant endosymbiosis,such as with arbuscular mycorrhizal fungi(AMF),have rarely been reported.Here,using knockout and overexpression lines,we show that OsEXO70L2,which encodes a divergent EXO70 protein in rice,controls the number of primary roots and is essential for large lateral root formation.Furthermore,the OsEXO70L2 mutant sr1 displayed rare internal AMF hyphaeand no arbuscules.We also found that AMF sporulation can occur in roots despite low colonization and that AMF colonization and sporulation are modulated by photoperiod and co-culture with clover.Finally,genes related to auxin homeostasis were found to be affected in the OsEXO70L2 knockout or overexpression lines,suggesting that auxin is at least partly responsible for the phenotypes.This study provides new perspectives on the role of the exocyst complex during root development and AM in rice.
文摘【目的】深入研究甘蓝自交不亲和信号传导关键元件S-位点受体激酶SRK与臂重复蛋白ARC1及ARC1与Exo70A1间相互识别的分子机理,鉴定SRK-ARC1及ARC1-Exo70A1之间的互作区段,并分析其作用强度,明确蛋白间互作功能域。【方法】通过生物信息学分析得到蛋白功能域,根据分析结果以典型的自交不亲和结球甘蓝E1为材料分别扩增SRK、ARC1和Exo70A1含不同功能域的截短体片段,利用分子克隆技术将SRK激酶域(SRKj)及其截短体SRKjΔ1—SRKjΔ4,Exo70A1全长及其截短体Exo70A1Δ1—Exo70A1Δ3的编码序列分别亚克隆至p GADT7(AD)质粒,将ARC1及其截短体ARC1Δ1—ARC1Δ8的编码序列分别亚克隆至载体p GBKT7(BD)质粒。用PEG/Li Ac法将获得的AD和BD重组质粒两两组合分别共转化到酵母AH109感受态中,观察融合菌株在SD/-Leu-Trp-His-Ade/X-α-gal/25 m M 3-AT平板上的菌落生长情况和颜色变化情况,进一步测定其β-半乳糖苷酶活性。最后通过原核表达体外孵育检测蛋白质相互作用的方法对SRK-ARC1及ARC1-Exo70A1的相互作用进行验证。【结果】DNA测序和内切酶分析显示成功构建18个酵母双杂交表达载体,且无自激活能力。在SRK-ARC1的10个试验组合中,只有ARC1Δ4、ARC1Δ8、ARC1与SRKj组合的融合菌株在SD/-Leu-Trp-His-Ade/X-α-gal/25 m M 3-AT培养基上长出蓝色菌落,激活报告基因HIS3、ADE2和MEL1。随着SRKj或ARC1截短体片段的延长,二者的β-半乳糖苷酶活性逐渐增加,其中,ARC1Δ4与SRKj组合的β-半乳糖苷酶活性最高(酶活为15.98)。在ARC1-Exo70A1 16个试验组合中,Exo70A1Δ3与ARC1Δ1Δ3都相互作用,其融合菌株在SD/-Leu-Trp-His-Ade/X-α-gal/25 m M 3-AT培养基上长出蓝色菌落,激活报告基因HIS3、ADE2和MEL1。随着ARC1或Exo70A1截短体片段的延长,二者的β-半乳糖苷酶活性呈现先增加后降低的趋势,其中ARC1Δ2与Exo70A1Δ3组合的β-半乳糖苷酶活性最大(酶活性为25.07)。说明ARC1的N端和Exo70A1的N端发生了互作,而ARC1的C端、全长与Exo70A1都不发生互作。体外表达检测蛋白相互作用发现,SRKj与ARC1Δ4、ARC1Δ2与Exo70A1Δ3均可以直接发生相互作用。【结论】SRK的激酶域(SRKj)与ARC1的C端臂重复区发生互作,缩短SRK激酶域中的任何结构域或者缩短ARC1的臂重复区,二者都不会发生互作。ARC1的亮氨酸拉链和蜷曲螺旋与Exo70A1的N端结构域(去除pfam Exo70A1域)介导了二者的相互作用。SRK-ARC1的作用力强度小于ARC1-Exo70A1的作用力强度。
