In several filamentous fungi,incident light and environmental stress signaling share the mitogen-activated protein kinase(MAPK)HOG(SAK)pathway.It has been revealed that short-term illumination with blue light triggers...In several filamentous fungi,incident light and environmental stress signaling share the mitogen-activated protein kinase(MAPK)HOG(SAK)pathway.It has been revealed that short-term illumination with blue light triggers the activation of the HOG pathway in Trichoderma spp.In this study,we demonstrate the crucial role of the basic leucine zipper transcription factor ATF1 in blue light responses and signaling downstream of the MAPK HOG1 in Trichoderma guizhouense.The lack of ATF1 severely impaired photoconidiation and delayed vegetative growth and conidial germination.Upon blue light or H2O2 stimuli,HOG1 interacted with ATF1 in the nucleus.Genome-wide transcriptome analyses revealed that 61.8%(509 out of 824)and 85.2%(702 out of 824)of blue light-regulated genes depended on ATF1 and HOG1,respectively,of which 58.4%(481 out of 824)were regulated by both of them.Our results also show that blue light promoted conidial germination and HOG1 and ATF1 played opposite roles in controlling conidial germination in the dark.Additionally,the lack of ATF1 led to reduced oxidative stress resistance,probably because of the downregulation of catalase-encoding genes.Overall,our results demonstrate that ATF1 is the downstream component of HOG1 and is responsible for blue light responses,conidial germination,vegetative growth,and oxidative stress resistance in T.guizhouense.展开更多
目的:探讨内质网氧化还原酶-1α(endoplasmic reticulum oxidoreductin 1α,ERO1α)在同型半胱氨酸(homocysteine,Hcy)诱导的肝细胞内质网应激(endoplasmic reticulum stress,ERS)中的作用及机制.方法:培养人肝细胞,用Hcy(100μmol/L)干...目的:探讨内质网氧化还原酶-1α(endoplasmic reticulum oxidoreductin 1α,ERO1α)在同型半胱氨酸(homocysteine,Hcy)诱导的肝细胞内质网应激(endoplasmic reticulum stress,ERS)中的作用及机制.方法:培养人肝细胞,用Hcy(100μmol/L)干预,同时设对照组,使用酶联免疫法(ELISA)检测ERS相关蛋白葡萄糖调节蛋白78(glucoseregulated protein 78,GRP78)、X盒结合蛋白-1(X-box binding protein-1,XBP-1)、内质网类似激酶(protein kinase RNA-like endoplasmic r e t i c u l u m k i n a s e,P E R K)及激活转录因子6(activating transcription factor 6,ATF6)的水平;用不同浓度Hcy(0、50、100、200、500μmol/L)和100μmol/L Hcy+叶酸+维生素B12(VB12)干预,运用实时定量PCR和Western blot检测ERO1αm RNA和蛋白水平;构建ERO1α基因重组质粒和RNA干扰片段并感染肝细胞,检测ERO1αm RNA和蛋白表达变化;采用ELISA法检测其对ERS相关蛋白的调控作用.结果:与对照组比较,Hcy组GRP78、ATF6、P E R K、X B P-1水平升高(P<0.01,P<0.01,P<0.05,P<0.01);不同浓度Hcy干预后,肝细胞内ERO1αm RNA及蛋白水平呈下降趋势,且随着Hcy浓度的增加而减少(P<0.01),呈剂量依赖关系;将ERO1α重组质粒转染肝细胞后,ERO1αm RNA及蛋白表达明显增加(P<0.01);将三个不同的ERO1αsi RNA片段转染肝细胞后,ERO1αm RNA及蛋白表达降低(P<0.01),其中si RNA2作用最明显(P<0.01);与Hcy组相比,Hcy+p ERO1α组GRP78、XBP-1、PERK及ATF6均明显降低(P<0.01),而Hcy+si RNA2组均明显升高(P<0.01).结论:Hcy可能通过下调ERO1α引起肝细胞GRP78-XBP-1/PERK/ATF6表达增加促进ERS发生.展开更多
Aberrant chromosomal fusion of the Ewing's sarcoma oncogene(EWS)to several different cellular partners produces the Ewing's family of oncoproteins(EWSfusion-proteins,EFPs)and associated tumors(EFTs).EFPs are p...Aberrant chromosomal fusion of the Ewing's sarcoma oncogene(EWS)to several different cellular partners produces the Ewing's family of oncoproteins(EWSfusion-proteins,EFPs)and associated tumors(EFTs).EFPs are potent transcriptional activators,dependent on the N-terminal region of EWS(the EWS-activationdomain,EAD)and this function is thought to be central to EFT oncogenesis and maintenance.Thus EFPs are promising therapeutic targets,but detailed molecular studies will be pivotal for exploring this potential.Such studies have so far largely been restricted to intact mammalian cells while recent evidence has indicated that a mammalian cell-free transcription system may not support bona fide EAD function.Therefore,the lack of manipulatable assays for the EAD presents a significant barrier to progress.Using Xenopus laevis oocytes we describe a plasmid-based micro-injection assay that supports efficient,bona fide EAD transcriptional activity and hence provides a new vehicle for molecular dissection of the EAD.展开更多
基金supported by the National Natural Science Foundation of China (NSFC) (Grant Nos.32070101 and 32270053)the Fundamental Research Funds for the Central Universities (Grant Nos.XUEKEN2023039,XUEKEN2023041,RENCAI2022005,and KYT2023001)the Jiangsu Agriculture Science and Technology Innovation fund (JASTIF) (Grant No.CX (21)2018).
文摘In several filamentous fungi,incident light and environmental stress signaling share the mitogen-activated protein kinase(MAPK)HOG(SAK)pathway.It has been revealed that short-term illumination with blue light triggers the activation of the HOG pathway in Trichoderma spp.In this study,we demonstrate the crucial role of the basic leucine zipper transcription factor ATF1 in blue light responses and signaling downstream of the MAPK HOG1 in Trichoderma guizhouense.The lack of ATF1 severely impaired photoconidiation and delayed vegetative growth and conidial germination.Upon blue light or H2O2 stimuli,HOG1 interacted with ATF1 in the nucleus.Genome-wide transcriptome analyses revealed that 61.8%(509 out of 824)and 85.2%(702 out of 824)of blue light-regulated genes depended on ATF1 and HOG1,respectively,of which 58.4%(481 out of 824)were regulated by both of them.Our results also show that blue light promoted conidial germination and HOG1 and ATF1 played opposite roles in controlling conidial germination in the dark.Additionally,the lack of ATF1 led to reduced oxidative stress resistance,probably because of the downregulation of catalase-encoding genes.Overall,our results demonstrate that ATF1 is the downstream component of HOG1 and is responsible for blue light responses,conidial germination,vegetative growth,and oxidative stress resistance in T.guizhouense.
文摘目的:探讨内质网氧化还原酶-1α(endoplasmic reticulum oxidoreductin 1α,ERO1α)在同型半胱氨酸(homocysteine,Hcy)诱导的肝细胞内质网应激(endoplasmic reticulum stress,ERS)中的作用及机制.方法:培养人肝细胞,用Hcy(100μmol/L)干预,同时设对照组,使用酶联免疫法(ELISA)检测ERS相关蛋白葡萄糖调节蛋白78(glucoseregulated protein 78,GRP78)、X盒结合蛋白-1(X-box binding protein-1,XBP-1)、内质网类似激酶(protein kinase RNA-like endoplasmic r e t i c u l u m k i n a s e,P E R K)及激活转录因子6(activating transcription factor 6,ATF6)的水平;用不同浓度Hcy(0、50、100、200、500μmol/L)和100μmol/L Hcy+叶酸+维生素B12(VB12)干预,运用实时定量PCR和Western blot检测ERO1αm RNA和蛋白水平;构建ERO1α基因重组质粒和RNA干扰片段并感染肝细胞,检测ERO1αm RNA和蛋白表达变化;采用ELISA法检测其对ERS相关蛋白的调控作用.结果:与对照组比较,Hcy组GRP78、ATF6、P E R K、X B P-1水平升高(P<0.01,P<0.01,P<0.05,P<0.01);不同浓度Hcy干预后,肝细胞内ERO1αm RNA及蛋白水平呈下降趋势,且随着Hcy浓度的增加而减少(P<0.01),呈剂量依赖关系;将ERO1α重组质粒转染肝细胞后,ERO1αm RNA及蛋白表达明显增加(P<0.01);将三个不同的ERO1αsi RNA片段转染肝细胞后,ERO1αm RNA及蛋白表达降低(P<0.01),其中si RNA2作用最明显(P<0.01);与Hcy组相比,Hcy+p ERO1α组GRP78、XBP-1、PERK及ATF6均明显降低(P<0.01),而Hcy+si RNA2组均明显升高(P<0.01).结论:Hcy可能通过下调ERO1α引起肝细胞GRP78-XBP-1/PERK/ATF6表达增加促进ERS发生.
基金supported by The Association for International Cancer Research(AICR)(grant 03-131 to K.A.W.L.).
文摘Aberrant chromosomal fusion of the Ewing's sarcoma oncogene(EWS)to several different cellular partners produces the Ewing's family of oncoproteins(EWSfusion-proteins,EFPs)and associated tumors(EFTs).EFPs are potent transcriptional activators,dependent on the N-terminal region of EWS(the EWS-activationdomain,EAD)and this function is thought to be central to EFT oncogenesis and maintenance.Thus EFPs are promising therapeutic targets,but detailed molecular studies will be pivotal for exploring this potential.Such studies have so far largely been restricted to intact mammalian cells while recent evidence has indicated that a mammalian cell-free transcription system may not support bona fide EAD function.Therefore,the lack of manipulatable assays for the EAD presents a significant barrier to progress.Using Xenopus laevis oocytes we describe a plasmid-based micro-injection assay that supports efficient,bona fide EAD transcriptional activity and hence provides a new vehicle for molecular dissection of the EAD.
