Plant growth and crop productivity are severely affected by abiotic stress on a global scale.WD40 repeat-containing proteins play a significant role in the development and environmental adaptation of eukaryotes.In thi...Plant growth and crop productivity are severely affected by abiotic stress on a global scale.WD40 repeat-containing proteins play a significant role in the development and environmental adaptation of eukaryotes.In this study,OsABT,a stress response gene,was cloned from rice(Oryza sativa L.cv.Nipponbare).OsABT encodes a protein containing seven WD40 domains.Expression analysis revealed that the OsABT gene was first up-regulated and then down-regulated following treatment with abscisic acid(ABA)and NaCl,but was down-regulated when treated with PEG8000.Subcellular localization results showed that OsABT was located in the cytoplasm and nucleus of Arabidopsis roots.OsABT transgenic Arabidopsis showed significantly increased tolerance to ABA and salt stress during plant seedling development.However,the transgenic lines were more sensitive to drought stress.Moreover,OsABT can interact with OsABI2,a component of ABA signaling pathway.These results showed that OsABT plays a positive regulatory role in response to salt stress and a negative role in response to drought stress in Arabidopsis.展开更多
In this study, we isolated a WD40-repeat gene from Artemisia annua glandular trichomes. This gene shows 69.97% sequence similarity to Arabidopsis TTG1 at aminoacid level. Sub-cellular localization study shows that AaW...In this study, we isolated a WD40-repeat gene from Artemisia annua glandular trichomes. This gene shows 69.97% sequence similarity to Arabidopsis TTG1 at aminoacid level. Sub-cellular localization study shows that AaWD40 protein diffuses in both cell nucleus and cytosol. The correct nuclear localization of AaWD40 was observed when co-expressed with AabHLH, a putative A. thaliana AtTTG1 homologue cloned from Artemisia annua glandular trichomes. When AaWD40 gene was ectopically over expressed in Arabidopsis transparent testa glabrous1-1 (ttg1-1) mutants of A. thaliana, PAs production in seeds was restored, and the trichomeless phenotypes of mutant were rescued. Real-time PCR analysis results revealed that ETC1, CPC, TTG2 and BAN (the downstream targets of AtTTG1 depend on regulatory complex), which regulate the epidermal differentiation and anthocyanin biosynthesis were differentially expressed as a result of AaWD40 over expression. Furthermore, the CLV1, CLV2, CLV3 and WUS, which are required to maintain the stem-cell niche of Arabidopsis shoot apex, were also modulated by AaWD40 and Arabidopsis TTG1. The transcriptions of AP2/ERF, bHLH, MYB, WRKY and NACs family proteins, which are mostly involved in defense, stress response and development regulation, were remarkably modulated by AaWD40 over expression. We hypothesize that WD40 repeat proteins act as a crucial factor in regulating a wide variety of cellular functions in A. thaliana.展开更多
AIM: To investigate role of putative mitogen-activated protein kinase activator with WD40 repeats (MAWD)/ MAWD binding protein (MAWBP) in gastric cancer (GC). METHODS: MAWBP and MAWD mRNA expression level was examined...AIM: To investigate role of putative mitogen-activated protein kinase activator with WD40 repeats (MAWD)/ MAWD binding protein (MAWBP) in gastric cancer (GC). METHODS: MAWBP and MAWD mRNA expression level was examined by real-time reverse transcriptasepolymerase chain reaction and semi-quantitative polymerase chain reaction in six GC cell lines. Western blotting was used to examine the protein expression levels. We developed GC cells that stably overexpressed MAWBP and MAWD, and downregulated expression by RNA interference assay. Proliferation and migration of these GC cells were analyzed by 3-(4,5-dimethyl2-thiazolyl)-2,5-diphenyl tetrazolium bromide (MTT), soft agar, tumorigenicity, migration and transwell assays. The effect of expression of MAWBP and MAWD on transforming growth factor (TGF)-β1-induced epithelialmesenchymal transition (EMT) was examined by transfection of MAWBP and MAWD into GC cells. We detected the levels of EMT markers E-cadherin, N-cadherin and Snail in GC cells overexpressing MAWBP and MAWD by Western blotting. The effect of MAWBP and MAWD on TGF-β signal was detected by analysis of phosphorylation level and nuclear translocation of Smad3 using Western blotting and immunofluorescence. RESULTS: Among the GC cell lines, expression of endogenous MAWBP and MAWD was lowest in SGC7901 cells and highest in BGC823 cells. MAWBP and MAWD were stably overexpressed in SGC7901 cells and knocked down in BGC823 cells. MAWBP and MAWD inhibited GC cell proliferation in vitro and in vivo . MTT assay showed that overexpression of MAWBP and MAWD suppressed growth of SGC7901 cells (P < 0.001), while knockdown of these genes promoted growth of BGC823 cells (P < 0.001). Soft agar colony formation experiments showed that overexpression of MAWBP and MAWD alone or together reduced colony formation compared with vector group in SGC7901 (86.25±8.43, 12.75±4.49, 30±6.41 vs 336.75±22.55, P < 0.001), and knocked-down MAWBP and MAWD demonstrated opposite effects (131.25±16.54, 88.75±11.12, 341.75±22.23 vs 30.25±8.07, P < 0.001). Tumorigenicity experiments revealed that overexpressed MAWBP and MAWD inhibited GC cell proliferation in vivo (P < 0.001). MAWBP and MAWD also inhibited GC cell invasion. Transwell assay showed that the number of traverse cells of MAWBP, MAWD and coexpression group were more than that in vector group (84±16.57, 98.33±9.8, 29±16.39 vs 298±11.86, P < 0.001). Coexpression of MAWBP and MAWD significantly decreased the cells traversing the matrix membrane. Conversely, knocked-down MAWBP and MAWD correspondingly promoted invasion of GC cells (100.67±14.57, 72.66±8.51, 330.67±20.55 vs 27±11.53, P < 0.001). More importantly, coexpression of MAWBP and MAWD promoted EMT. Cells that coexpressed MAWBP and MAWD displayed a pebble-like shape and tight cell-cell adhesion, while vector cells showed a classical mesenchymal phenotype. Western blotting showed that expression of E-cadherin was increased, and expression of N-cadherin and Snail was decreased when cells coexpressed MAWBP and MAWD and were treated with TGF-β1. Nuclear translocation of p-Smad3 was reduced by attenuating its phosphorylation. CONCLUSION: Coexpression of MAWBP and MAWD inhibited EMT, and EMT-aided malignant cell progression was suppressed.展开更多
WD40 repeat-containing proteins(WD40 proteins)serve as versatile scaffolds for protein-protein interac-tions,modulating a variety of cellular processes such as plant stress and hormone responses.Here we report the ide...WD40 repeat-containing proteins(WD40 proteins)serve as versatile scaffolds for protein-protein interac-tions,modulating a variety of cellular processes such as plant stress and hormone responses.Here we report the identification of a WD40 protein,XIW1(for XPO1-interacting WD40 protein 1),which positively regulates the abscisic acid(ABA)response in Arabidopsis.XIW1 is located in the cytoplasm and nucleus.We found that it interacts with the nuclear transport receptor XPO1 and is exported by XPO1 from the nucleus.Mutation of XIW1 reduces the induction of ABA-responsive genes and the accumulation of ABA Insensitive 5(ABI5),causing mutant plants with ABA-insensitive phenotypes during seed germination and seedling growth,and decreased drought stress resistance.ABA treatment upregulates the expression of XIW1,and both ABA and abiotic stresses promote XIW1 accumulation in the nucleus,where it interacts with ABI5.Loss of XIW1 function results in rapid proteasomal degradation of ABI5.Taken together,these findings suggest that XIW1 is a nucleocytoplasmic shuttling protein and plays a positive role in ABA responses by interacting with and maintaining the stability of ABI5 in the nucleus.展开更多
目的探讨F框/WD40域蛋白7(F-box and WD-40domain protein7,FBXW7)和胆固醇调节元件结合蛋白(sterol regulatory element binding proteins,SREBPs)基因多态性与新疆地区维吾尔族人群冠心病的相关性。方法 360例维吾尔族冠心病患者(冠...目的探讨F框/WD40域蛋白7(F-box and WD-40domain protein7,FBXW7)和胆固醇调节元件结合蛋白(sterol regulatory element binding proteins,SREBPs)基因多态性与新疆地区维吾尔族人群冠心病的相关性。方法 360例维吾尔族冠心病患者(冠心病组)和373例维吾尔族健康受试者(对照组),2组采用改良的多重高温连接酶检测反应基因分型技术分析FBXW7、SREBP-1和SREBP-2基因多态性位点,检测2组血压和血生化指标,比较2组基因频率分布情况,并采用多因素logistic回归分析冠心病的独立危险因素。结果冠心病组吸烟、饮酒比率,合并高血压、糖尿病、高脂血症发生率,空腹血糖、三酰甘油、总胆固醇、低密度脂蛋白胆固醇水平均高于对照组(P<0.05),高密度脂蛋白胆固醇水平低于对照组(P<0.05);2组SREBP-1基因rs9902941位点基因型分布和隐性模型(TT/CT+CC)分布、SREBP-2基因rs7288536位点基因型分布及显性模型(TT/CT+CC)和相加模型(CT/TT+CC)分布、FBXW7基因rs10033601位点基因型分布和隐性模型(GG/AG+AA)分布比较差异均有统计学意义(P<0.05);多因素logistic回归校正性别、年龄、吸烟、饮酒、血压、血糖及血脂等影响因素后,SREBP-1基因rs9902941位点隐性模型、FBXW7基因rs10033601位点隐性模型及SREBP-2基因rs7288536位点相加模型为冠心病的独立危险因素(OR=1.900,95%CI:1.041~3.467,P=0.036;OR=1.670,95%CI:1.054~2.645,P=0.029;OR=1.578,95%CI:1.133~2.197,P=0.007)。结论在新疆维吾尔族人群中,FBXW7基因rs10033601多态性、SREBP-1基因rs9902941多态性和SREBP-2基因rs7288536多态性与冠心病发生具有相关性。展开更多
Background:Accumulated studies have demonstrated that Kruppel‑like factor 5(KLF5),a transcription factor,plays an important role in regulating cell proliferation and tissue remodeling through the expression of its dow...Background:Accumulated studies have demonstrated that Kruppel‑like factor 5(KLF5),a transcription factor,plays an important role in regulating cell proliferation and tissue remodeling through the expression of its downstream genes.KLF5‑related factors are expected to be involved in the healing process after myocardial injury or myocardial ischemic changes,especially for the forensic diagnosis of myocardial ischemic physiopathology.Aim and Objectives:This study aimed to explore the discrimination ability and applicability of KLF5-related factors in SCD caused by MI compared with other causes of death to provide further insights into the forensic diagnosis of myocardial ischemic pathology.Materials and Methods:The relative quantification of F‑Box and WD Repeat Domain Containing 7(FBW7),KLF5,factor‑binding protein(FGFBP)1,and FGFBP2 messenger RNAs(mRNAs)in myocardial tissue samples was performed using real‑time fluorescence quantitative reverse transcription polymerase chain reaction.KLF5 and FGFBP1/2 protein levels were examined using immunohistochemistry(IHC).The forensic autopsy cases(27 in total,autopsy within 72 h postmortem)included seven cases of acute myocardial infarction and 10 cases of acute myocardial ischemia.There were 10 cases in the control group,including four cases of traffic injury one case of injury by fall from height,one case of electric death,and four cases of blunt force injury.Results:Characteristic results were found in myocardial samples from three groups of deaths:KLF5 and FGFBP1 mRNA levels were significantly elevated in the infarction and ischemia groups,while FBW7 mRNA levels were significantly decreased.FBW7 is an important ubiquitin ligase that can mediate the degradation of KLF5 protein.In addition,FBW7 and FGFBP2 mRNA levels were decreased in the infarction group compared with the ischemia group.The IHC results were consistent with the observed mRNA expression patterns.Conclusions:Quantitative detection of FBW7,KLF5,FGFBP1,and FGFBP2 mRNA transcripts in myocardial tissues supports the pathophysiological study of myocardial ischemic diseases and provides molecular pathological evidence for forensic discrimination of death causes.展开更多
基金supported by the Major Program of the Zhejiang Province for Food Crop Breeding(Grant No.2016C02050-6)the Key Program of Hangzhou Agricultural Scientific Research(Grant No.20191203B08)。
文摘Plant growth and crop productivity are severely affected by abiotic stress on a global scale.WD40 repeat-containing proteins play a significant role in the development and environmental adaptation of eukaryotes.In this study,OsABT,a stress response gene,was cloned from rice(Oryza sativa L.cv.Nipponbare).OsABT encodes a protein containing seven WD40 domains.Expression analysis revealed that the OsABT gene was first up-regulated and then down-regulated following treatment with abscisic acid(ABA)and NaCl,but was down-regulated when treated with PEG8000.Subcellular localization results showed that OsABT was located in the cytoplasm and nucleus of Arabidopsis roots.OsABT transgenic Arabidopsis showed significantly increased tolerance to ABA and salt stress during plant seedling development.However,the transgenic lines were more sensitive to drought stress.Moreover,OsABT can interact with OsABI2,a component of ABA signaling pathway.These results showed that OsABT plays a positive regulatory role in response to salt stress and a negative role in response to drought stress in Arabidopsis.
文摘In this study, we isolated a WD40-repeat gene from Artemisia annua glandular trichomes. This gene shows 69.97% sequence similarity to Arabidopsis TTG1 at aminoacid level. Sub-cellular localization study shows that AaWD40 protein diffuses in both cell nucleus and cytosol. The correct nuclear localization of AaWD40 was observed when co-expressed with AabHLH, a putative A. thaliana AtTTG1 homologue cloned from Artemisia annua glandular trichomes. When AaWD40 gene was ectopically over expressed in Arabidopsis transparent testa glabrous1-1 (ttg1-1) mutants of A. thaliana, PAs production in seeds was restored, and the trichomeless phenotypes of mutant were rescued. Real-time PCR analysis results revealed that ETC1, CPC, TTG2 and BAN (the downstream targets of AtTTG1 depend on regulatory complex), which regulate the epidermal differentiation and anthocyanin biosynthesis were differentially expressed as a result of AaWD40 over expression. Furthermore, the CLV1, CLV2, CLV3 and WUS, which are required to maintain the stem-cell niche of Arabidopsis shoot apex, were also modulated by AaWD40 and Arabidopsis TTG1. The transcriptions of AP2/ERF, bHLH, MYB, WRKY and NACs family proteins, which are mostly involved in defense, stress response and development regulation, were remarkably modulated by AaWD40 over expression. We hypothesize that WD40 repeat proteins act as a crucial factor in regulating a wide variety of cellular functions in A. thaliana.
基金Supported by The National Bio-Tech 863 Program, No.2006AA02A402National Natural Science Foundation of China, No. 30901717
文摘AIM: To investigate role of putative mitogen-activated protein kinase activator with WD40 repeats (MAWD)/ MAWD binding protein (MAWBP) in gastric cancer (GC). METHODS: MAWBP and MAWD mRNA expression level was examined by real-time reverse transcriptasepolymerase chain reaction and semi-quantitative polymerase chain reaction in six GC cell lines. Western blotting was used to examine the protein expression levels. We developed GC cells that stably overexpressed MAWBP and MAWD, and downregulated expression by RNA interference assay. Proliferation and migration of these GC cells were analyzed by 3-(4,5-dimethyl2-thiazolyl)-2,5-diphenyl tetrazolium bromide (MTT), soft agar, tumorigenicity, migration and transwell assays. The effect of expression of MAWBP and MAWD on transforming growth factor (TGF)-β1-induced epithelialmesenchymal transition (EMT) was examined by transfection of MAWBP and MAWD into GC cells. We detected the levels of EMT markers E-cadherin, N-cadherin and Snail in GC cells overexpressing MAWBP and MAWD by Western blotting. The effect of MAWBP and MAWD on TGF-β signal was detected by analysis of phosphorylation level and nuclear translocation of Smad3 using Western blotting and immunofluorescence. RESULTS: Among the GC cell lines, expression of endogenous MAWBP and MAWD was lowest in SGC7901 cells and highest in BGC823 cells. MAWBP and MAWD were stably overexpressed in SGC7901 cells and knocked down in BGC823 cells. MAWBP and MAWD inhibited GC cell proliferation in vitro and in vivo . MTT assay showed that overexpression of MAWBP and MAWD suppressed growth of SGC7901 cells (P < 0.001), while knockdown of these genes promoted growth of BGC823 cells (P < 0.001). Soft agar colony formation experiments showed that overexpression of MAWBP and MAWD alone or together reduced colony formation compared with vector group in SGC7901 (86.25±8.43, 12.75±4.49, 30±6.41 vs 336.75±22.55, P < 0.001), and knocked-down MAWBP and MAWD demonstrated opposite effects (131.25±16.54, 88.75±11.12, 341.75±22.23 vs 30.25±8.07, P < 0.001). Tumorigenicity experiments revealed that overexpressed MAWBP and MAWD inhibited GC cell proliferation in vivo (P < 0.001). MAWBP and MAWD also inhibited GC cell invasion. Transwell assay showed that the number of traverse cells of MAWBP, MAWD and coexpression group were more than that in vector group (84±16.57, 98.33±9.8, 29±16.39 vs 298±11.86, P < 0.001). Coexpression of MAWBP and MAWD significantly decreased the cells traversing the matrix membrane. Conversely, knocked-down MAWBP and MAWD correspondingly promoted invasion of GC cells (100.67±14.57, 72.66±8.51, 330.67±20.55 vs 27±11.53, P < 0.001). More importantly, coexpression of MAWBP and MAWD promoted EMT. Cells that coexpressed MAWBP and MAWD displayed a pebble-like shape and tight cell-cell adhesion, while vector cells showed a classical mesenchymal phenotype. Western blotting showed that expression of E-cadherin was increased, and expression of N-cadherin and Snail was decreased when cells coexpressed MAWBP and MAWD and were treated with TGF-β1. Nuclear translocation of p-Smad3 was reduced by attenuating its phosphorylation. CONCLUSION: Coexpression of MAWBP and MAWD inhibited EMT, and EMT-aided malignant cell progression was suppressed.
基金This work was supported by grants from the National Natural Science Foundation of China(31570250)the Science and Technology Project of Guangzhou City,China(201607020006,201804010377)the Natural Science Foundation of Guangdong Province,China(2015A030313410,2017A030313100).
文摘WD40 repeat-containing proteins(WD40 proteins)serve as versatile scaffolds for protein-protein interac-tions,modulating a variety of cellular processes such as plant stress and hormone responses.Here we report the identification of a WD40 protein,XIW1(for XPO1-interacting WD40 protein 1),which positively regulates the abscisic acid(ABA)response in Arabidopsis.XIW1 is located in the cytoplasm and nucleus.We found that it interacts with the nuclear transport receptor XPO1 and is exported by XPO1 from the nucleus.Mutation of XIW1 reduces the induction of ABA-responsive genes and the accumulation of ABA Insensitive 5(ABI5),causing mutant plants with ABA-insensitive phenotypes during seed germination and seedling growth,and decreased drought stress resistance.ABA treatment upregulates the expression of XIW1,and both ABA and abiotic stresses promote XIW1 accumulation in the nucleus,where it interacts with ABI5.Loss of XIW1 function results in rapid proteasomal degradation of ABI5.Taken together,these findings suggest that XIW1 is a nucleocytoplasmic shuttling protein and plays a positive role in ABA responses by interacting with and maintaining the stability of ABI5 in the nucleus.
文摘目的探讨F框/WD40域蛋白7(F-box and WD-40domain protein7,FBXW7)和胆固醇调节元件结合蛋白(sterol regulatory element binding proteins,SREBPs)基因多态性与新疆地区维吾尔族人群冠心病的相关性。方法 360例维吾尔族冠心病患者(冠心病组)和373例维吾尔族健康受试者(对照组),2组采用改良的多重高温连接酶检测反应基因分型技术分析FBXW7、SREBP-1和SREBP-2基因多态性位点,检测2组血压和血生化指标,比较2组基因频率分布情况,并采用多因素logistic回归分析冠心病的独立危险因素。结果冠心病组吸烟、饮酒比率,合并高血压、糖尿病、高脂血症发生率,空腹血糖、三酰甘油、总胆固醇、低密度脂蛋白胆固醇水平均高于对照组(P<0.05),高密度脂蛋白胆固醇水平低于对照组(P<0.05);2组SREBP-1基因rs9902941位点基因型分布和隐性模型(TT/CT+CC)分布、SREBP-2基因rs7288536位点基因型分布及显性模型(TT/CT+CC)和相加模型(CT/TT+CC)分布、FBXW7基因rs10033601位点基因型分布和隐性模型(GG/AG+AA)分布比较差异均有统计学意义(P<0.05);多因素logistic回归校正性别、年龄、吸烟、饮酒、血压、血糖及血脂等影响因素后,SREBP-1基因rs9902941位点隐性模型、FBXW7基因rs10033601位点隐性模型及SREBP-2基因rs7288536位点相加模型为冠心病的独立危险因素(OR=1.900,95%CI:1.041~3.467,P=0.036;OR=1.670,95%CI:1.054~2.645,P=0.029;OR=1.578,95%CI:1.133~2.197,P=0.007)。结论在新疆维吾尔族人群中,FBXW7基因rs10033601多态性、SREBP-1基因rs9902941多态性和SREBP-2基因rs7288536多态性与冠心病发生具有相关性。
基金supported by the Beijing Natural Science Foundation(grant number 7192121,China)General Program of National Natural Science Foundation of China(grant number 81971796,China).
文摘Background:Accumulated studies have demonstrated that Kruppel‑like factor 5(KLF5),a transcription factor,plays an important role in regulating cell proliferation and tissue remodeling through the expression of its downstream genes.KLF5‑related factors are expected to be involved in the healing process after myocardial injury or myocardial ischemic changes,especially for the forensic diagnosis of myocardial ischemic physiopathology.Aim and Objectives:This study aimed to explore the discrimination ability and applicability of KLF5-related factors in SCD caused by MI compared with other causes of death to provide further insights into the forensic diagnosis of myocardial ischemic pathology.Materials and Methods:The relative quantification of F‑Box and WD Repeat Domain Containing 7(FBW7),KLF5,factor‑binding protein(FGFBP)1,and FGFBP2 messenger RNAs(mRNAs)in myocardial tissue samples was performed using real‑time fluorescence quantitative reverse transcription polymerase chain reaction.KLF5 and FGFBP1/2 protein levels were examined using immunohistochemistry(IHC).The forensic autopsy cases(27 in total,autopsy within 72 h postmortem)included seven cases of acute myocardial infarction and 10 cases of acute myocardial ischemia.There were 10 cases in the control group,including four cases of traffic injury one case of injury by fall from height,one case of electric death,and four cases of blunt force injury.Results:Characteristic results were found in myocardial samples from three groups of deaths:KLF5 and FGFBP1 mRNA levels were significantly elevated in the infarction and ischemia groups,while FBW7 mRNA levels were significantly decreased.FBW7 is an important ubiquitin ligase that can mediate the degradation of KLF5 protein.In addition,FBW7 and FGFBP2 mRNA levels were decreased in the infarction group compared with the ischemia group.The IHC results were consistent with the observed mRNA expression patterns.Conclusions:Quantitative detection of FBW7,KLF5,FGFBP1,and FGFBP2 mRNA transcripts in myocardial tissues supports the pathophysiological study of myocardial ischemic diseases and provides molecular pathological evidence for forensic discrimination of death causes.
