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To investigate the function of the MsphyA gene and its molecular mechanism in response to abiotic stress in alfalfa (Medicago sativa L.), this study conducted a bioinformatic analysis of the cloned MsphyA gene and detected its expression patterns under various abiotic stresses using qRT-PCR. Bioinformatic analysis revealed that the MsphyA gene has a CDS length of 3375 bp, encoding 1124 amino acids, and possesses typical PAS, GAF, and PHY motifs characteristic of the PHY family. Protein analysis indicated that MsphyA has a relative molecular mass of 124.3 kDa and a theoretical isoelectric point of 6.1. It is classified as an unstable, hydrophilic, non-secretory protein without a signal peptide and lacks transmembrane domains. Subcellular localization prediction suggested that MsphyA is localized to the nucleus. Phylogenetic analysis showed that MsphyA is most closely related to MtphyA from M. truncatula. qRT-PCR results demonstrated that salt, drought, acid, high-temperature, and low-temperature stresses all induced MsphyA expression. These findings indicate that MsphyA is widely involved in the abiotic stress responses in alfalfa. This study provides a foundation for further exploration of the MsphyA gene’s function., authors=FAN Wen-na1, ZHANG Meng-yao1, SHI Ya-qi1, YANG Yi-xin1, authorsList=FAN Wen-na, ZHANG Meng-yao, SHI Ya-qi, YANG Yi-xin, authorCompany=1 College of Animal Science and Technology, Henan University of Science and Technology, Luoyang, Henan Province 471023, China, correspAuthors=null, authorNote=null, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, fund=null), CN=ArticleExt(id=1309575153256858599, articleId=1309575152845816806, tenantId=1146029695717560320, journalId=1308397196074385509, language=CN, title=紫花苜蓿MsphyA基因鉴定分析及非生物胁迫下表达, columnId=1309565024058835950, journalTitle=草地学报, columnName=研究论文, runingTitle=null, highlight=null, articleAbstract=光敏色素A(phyA)是植物感知远红光信号并调控生长发育及逆境应答的关键受体。为探究MsphyA基因功能及其在紫花苜蓿(Medicago sativa L.)中响应非生物胁迫的分子机制,本研究对已克隆的MsphyA基因全长质粒进行测序鉴定及完整结构域生物信息学分析,并通过qRT-PCR技术检测该基因在不同非生物胁迫下的表达模式。生物信息学分析显示MsphyA基因编码序列(Coding sequence,CDS)区全长3375 bp,编码1124个氨基酸,具有典型的PHY家族的PAS,PHY和GAF motifs特殊结构域;蛋白分析显示其相对分子量为124.3 kDa,理论等电点为6.1,属于不含信号肽的非分泌不稳定亲水性蛋白且不具备跨膜结构;亚细胞定位预测该蛋白定位于细胞核;系统进化树表明MsphyA蛋白与蒺藜苜蓿MtphyA亲缘关系最近。qRT-PCR结果表明,盐、干旱、酸、高温、低温处理均能诱导MsphyA的表达,MsphyA广泛参与紫花苜蓿的非生物胁迫应答过程,本研究为进一步探究紫花苜蓿MsphyA基因功能提供了参考。, authors=樊文娜1, 张梦瑶1, 石亚奇1, 杨怡欣1, authorsList=樊文娜, 张梦瑶, 石亚奇, 杨怡欣, authorCompany=1 河南科技大学动物科技学院, 河南 洛阳 471023, correspAuthors=null, authorNote=樊文娜(1981-),女,汉族,河南许昌人,博士,副教授,主要从事饲料饲草资源开发与利用研究,E-mail:chou0516@163.com, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=ES7vYia/19QMkgIkL8zuxA==, pdfFileSize=7299609, 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Plant Physiology,2009,150(2):1083-1092 BAE G,CHOI G. Decoding of light signals by plant phytochromes and their interacting proteins[J]. Annual Review of Plant Biology,2008,59:281-311 TANG Y L,QIN H S,WANG T Q,et al. Research progress on light quality regulating plant abiotic stress resistance[J]. Jiangsu Agricultural Sciences,2025,53(18):62-68 唐艳领,秦恒山,王腾起,等.光质调控植物非生物胁迫抗性的研究进展[J].江苏农业科学,2025,53(18):62-68 CHEN Y,WANG Y Q. Cloning and function analysis of rice phytochrome a gene in photomorphogenesis[J]. Journal of South China Normal University (Natural Science Edition),2014,46(1):71-76 陈由,王亚琴.水稻光敏色素A基因克隆及其在光形态建成中的作用[J]. 华南师范大学学报,2014,46(1):71-76 ABDELLATIF I M Y,YUAN S Z,NA R H,et al. Functional characterization of tomato phytochrome A and B1B2 mutants in response to heat stress[J]. International Journal of Molecular Sciences,2022,23(3): 1681 WANG F,GUO Z X,LI H Z,et al. Phytochrome A and B function antagonistically to regulate cold tolerance via abscisic acid-dependent jasmonate signaling[J]. Plant physiology,2016,170(1): 459-471 BOGGS J Z,LOEWY K,BIBEE K,et al. Phytochromes influence stomatal conductance plasticity in Arabidopsis thaliana[J]. Plant Growth Regulation,2010,60(2):77-81 MA L,HAN R,YANG Y Q,et al. Phytochromes enhance SOS2-mediated PIF1 and PIF3 phosphorylation and degradation to promote Arabidopsis salt tolerance[J]. The Plant Cell,2023,35(8):2997-3020 DU X F,SHEN M M,WANG Z L,et al. Responses of SiPHYA and SiPHYB gene to light quality,photoperiod and abiotic stresses in foxtail millet (Setaria italica) [J]. Journal of Agricultural Biotechnology,2024,32(5):983-994 杜晓芬,沈蒙蒙,王智兰,等.谷子SiPHYA和SiPHYB基因对光质、光周期和非生物胁迫的响应[J]. 农业生物技术学报,2024,32(5):983-994 YANG T,LV R H,LI J,et al. Phytochrome A and B negatively regulate salt stress tolerance of Nicotiana tobacum via ABA-jasmonic acid synergistic cross-talk[J]. Plant and Cell Physiology,2018,59(11): 2381-2393 ZHANG T,LENG H J,CUI J,et al. Identification of synaptotagmin gene family members in alfalfa and their transcript profiles under abiotic stresses[J]. Acta Prataculturae Sinica,2026,35(4):158-168 张钿,冷华娟,崔婧,等.紫花苜蓿突触结合蛋白家族成员鉴定与非生物胁迫下的表达分析[J]. 草业学报,2026,35(4):158-168 ZOU W P,LIU Y,ZHAI J X,et al. Cloning of MsNAC053 from alfalfa and analysis of its transcript profile in response to abiotic stresses[J]. Acta Prataculturae Sinica,2025,34(9):121-133 邹苇鹏,刘怡,翟佳兴,等.紫花苜蓿MsNAC053基因克隆及其对非生物胁迫的响应分析[J]. 草业学报,2025,34(9):121-133 WANG M J,PENG J,QI L J,et al. Research advances on light control of plant responses to abiotic stresses[J]. Plant Physiology Journal,2023,59(4):682-704 王美娇,彭晶,齐立娟,等.光信号调控植物响应非生物胁迫的研究进展[J]. 植物生理学报,2023,59(4):682-704 LI P,YANG L L,CHEN Q X,et al. Two strategies of cloning Medicago sativa phytochrome A and B genes[J]. Acta Prataculturae Sinica,2011,20(6):85-92 李平,杨玲玲,陈其新,等.两种策略分别克隆紫花苜蓿光敏色素A、B基因[J]. 草业学报,2011,20(6):85-92 YANG L L. Cloning of alfalfa phytochrome a gene using rapid amplification of cDNA ends[D]. Zhengzhou:Henan Agricultural University,2008:29-46 杨玲玲.RACE法克隆紫花苜蓿光敏色素A基因[D]. 郑州:河南农业大学,2008:29-46 YANG Y X. Screening of alfalfa internal reference genes and determination of antioxidant indexes under different stress conditions[D]. Luoyang: Henan University of Science and Technology,2024:21-35 杨怡欣.不同胁迫条件下苜蓿内参基因的筛选及抗氧化指标的测定[D]. 洛阳:河南科技大学,2024:21-35 CHEN F,LI B,DEMONE J,et al. Photoreceptor partner FHY1 has an independent role in gene modulation and plant development under far-red light[J]. Proceedings of the National Academy of Sciences of the United States of America,2014,111(32):11888-11893 QUAIL P H. Phytochrome photosensory signalling networks[J]. Nature Reviews Molecular Cell biology,2002,3(2):85-93 CHEN Q,YU Q W,ZHENG D F,et al. Regulation of PHYA gene on photothermal sensitivity and related traits in tobacco[J]. Molecular Plant Breeding,2021,19(13):4392-4400 陈倩,喻奇伟,郑登峰,等. 烟草PHYA基因的光温敏感性及生物学功能分析[J].分子植物育种,2021,19(13):4392-4400 YUE L Q,XING Q J,ZHANG X L,et al. Research progress on the effect of phytochrome-interacting factors in plant resistance to abiotic stress[J]. Acta Horticulturae Sinica,2021,48(4):632-646 岳玲琦,邢巧娟,张晓兰,等. 光敏色素互作因子在植物抵御逆境胁迫中的作用研究进展[J]. 园艺学报,2021,48(4):632-646 SHICHIJO C,KATADA K,TANAKA O,et al. Phytochrome A-mediated inhibition of seed germination in tomato[J]. Planta,2001,213(5):764-769 JOHNSON E,BRADLEY M,HARBERD N P,et al. Photo responses of light-grown phyA mutants of Arabidopsis (phytochrome A is required for the perception of daylength extensions) [J]. Plant physiology,1994,105(1):141-149 WEI S,XU X H,LU X B,et al. The rice phytochrome genes,PHYA and PHYB have synergistic effects on anther development and pollen viability[J]. Scientific Reports,2017,7:6439 KIRCHER S,GIL P,KOZMA-BOGNÁR L,et al. Nucleocytoplasmic partitioning of the plant photoreceptors phytochrome A,B,C,D,and E is regulated differentially by light and exhibits a diurnal rhythm[J]. The Plant Cell,2002,14(7):1541-1555 LI Y K. Functional analysis of three photosensitive pigment genes in rye during photomorphogenesis[D]. Zhengzhou: Henan Agricultural University,2025:27-39 李彦坤.黑麦3个光敏色素基因在光形态建成中的功能分析[D]. 郑州:河南农业大学,2025:27-39 HISADA A,HANZAWA H,WELLER J L,et al. Light-induced nuclear translocation of endogenous pea phytochrome A visualized by immunocytochemical procedures[J]. The Plant Cell,2000,12(7):1063-1078 RÖSLER J,KLEIN I,ZEIDLER M. Arabidopsis fhl/fhy1 double mutant reveals a distinct cytoplasmic action of phytochrome A[J]. Proceedings of the National Academy of Sciences of the United States of America,2007,104(25):10737-10742 HAN Y J,KIM H S,KIM Y M,et al. Functional characterization of phytochrome autophosphorylation in plant light signaling[J]. Plant and Cell Physiology,2010,51(4):596-609 ROCKWELL N C,SU Y S,LAGARIAS J C. Phytochrome structure and signaling mechanisms[J]. Annual Review of Plant Biology,2006,57(1): 837-858 XU J W,YOU Y S,XU Y W,et al. Research progress on the function of PIFs transcription factors[J]. Protection Forest Science and Technology,2025(4):74-80 许建闻,由云生,徐亚伟,等.PIFs转录因子功能研究进展[J]. 防护林科技,2025(4):74-80 WANG F. Roles and mechanisms of phyA-,HY5- and PIF4- mediated light quality-regulated cold tolerance in tomato[D]. Hangzhou: Zhejiang University,2017:19-55 王峰. PhyA、HY5和PIF4在光质调控番茄低温抗性中的机制研究[D]. 杭州:浙江大学,2017:19-55 JIA X P,ZHANG B,HE Z X,et al. The responsive characteristics of phytochrome genes to photoperiod,abiotic stresses and identification of their key natural variation sites in foxtail millet (Setaria italica L.) [J]. Chinese Journal of Biotechnology,2022,38(5):1929-1945 贾小平,张博,何占祥,等. 谷子光敏色素基因光周期、非生物胁迫响应特性及关键自然变异位点鉴定[J]. 生物工程学报,2022,38(5):1929-1945 ABDELLATIF I M Y,YUAN S Z,YOSHIHARA S,et al. Stimulation of tomato drought tolerance by PHYTOCHROME A and B1B2 mutations[J]. International Journal of Molecular Sciences,2023,24(2):1560 GONZÁLEZ C V,IBARRA S E,PICCOLI P N,et al. Phytochrome B increases drought tolerance by enhancing ABA sensitivity in Arabidopsis thaliana[J]. Plant,Cell & Environment,2012,35(11):1958-1968 PENG Y X,ZHU H Y,WANG Y T,et al. Revisiting the role of light signaling in plant responses to salt stress[J]. Horticulture Research,2025,12:uhae26 LI H,ZHOU Y Y,QIN X Y,et al. Reconstitution of phytochrome A-mediated light modulation of the ABA signaling pathways in yeast[J]. Proceedings of the National Academy of Sciences of the United States of America,2023,120(34):e2302901120 OLSEN J E,JUNTTILA O,NILSEN J,et al. Ectopic expression of oat phytochrome A in hybrid aspen changes critical daylength for growth and prevents cold acclimatization[J]. The Plant Journal,1997,12(6):1339-1350 FAN W N,REN B B,SHI P F,et al. Advance in alfalfa of phytochrome A regulating fall dormancy[J]. China Feed,2022,1(5):112-116 樊文娜,任冰冰,史鹏飞,等. 光敏色素A参与苜蓿秋眠调控分子机制的研究进展[J]. 中国饲料,2022,1(5):112-116 NIETO C,LÓPEZ-SALMERÓN V,DAVIÈRE J M,et al. ELF3-PIF4 interaction regulates plant growth independently of the evening complex[J]. Current Biology,2015,25(2):187-193 YANG T,ZHANG H,JIANG X P,et al. Phytochrome alleviates cadmium toxicity by regulating gibberellic acid and brassinolide in Nicotiana tabacum[J]. Plant Physiology and Biochemistry,2025,219:109375)
Phytochrome A (phyA) is a key receptor that enables plants to perceive far-red light signals and regulates their growth, development, and stress responses. To investigate the function of the MsphyA gene and its molecular mechanism in response to abiotic stress in alfalfa (Medicago sativa L.), this study conducted a bioinformatic analysis of the cloned MsphyA gene and detected its expression patterns under various abiotic stresses using qRT-PCR. Bioinformatic analysis revealed that the MsphyA gene has a CDS length of 3375 bp, encoding 1124 amino acids, and possesses typical PAS, GAF, and PHY motifs characteristic of the PHY family. Protein analysis indicated that MsphyA has a relative molecular mass of 124.3 kDa and a theoretical isoelectric point of 6.1. It is classified as an unstable, hydrophilic, non-secretory protein without a signal peptide and lacks transmembrane domains. Subcellular localization prediction suggested that MsphyA is localized to the nucleus. Phylogenetic analysis showed that MsphyA is most closely related to MtphyA from M. truncatula. qRT-PCR results demonstrated that salt, drought, acid, high-temperature, and low-temperature stresses all induced MsphyA expression. These findings indicate that MsphyA is widely involved in the abiotic stress responses in alfalfa. This study provides a foundation for further exploration of the MsphyA gene’s function.
FAN Wen-na, ZHANG Meng-yao, SHI Ya-qi, YANG Yi-xin.
Identification and Analysis of MsphyA Gene in Alfalfa and its Expression under Abiotic Stress[J].
Acta Agrestia Sinica,
2026
, 34
(9)
: 3211
-3222
.
DOI: 10.11733/j.issn.1007-0435.2026.09.007
基金
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国家自然科学基金 (32102585)
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Cloning of alfalfa phytochrome a gene using rapid amplification of cDNA ends[D]. Zhengzhou:Henan Agricultural University,2008:29-46 杨玲玲.RACE法克隆紫花苜蓿光敏色素A基因[D]. 郑州:河南农业大学,2008:29-46 YANG Y X. Screening of alfalfa internal reference genes and determination of antioxidant indexes under different stress conditions[D]. Luoyang: Henan University of Science and Technology,2024:21-35 杨怡欣.不同胁迫条件下苜蓿内参基因的筛选及抗氧化指标的测定[D]. 洛阳:河南科技大学,2024:21-35 CHEN F,LI B,DEMONE J,et al. Photoreceptor partner FHY1 has an independent role in gene modulation and plant development under far-red light[J]. Proceedings of the National Academy of Sciences of the United States of America,2014,111(32):11888-11893 QUAIL P H. Phytochrome photosensory signalling networks[J]. Nature Reviews Molecular Cell biology,2002,3(2):85-93 CHEN Q,YU Q W,ZHENG D F,et al. Regulation of PHYA gene on photothermal sensitivity and related traits in tobacco[J]. Molecular Plant Breeding,2021,19(13):4392-4400 陈倩,喻奇伟,郑登峰,等. 烟草PHYA基因的光温敏感性及生物学功能分析[J].分子植物育种,2021,19(13):4392-4400 YUE L Q,XING Q J,ZHANG X L,et al. Research progress on the effect of phytochrome-interacting factors in plant resistance to abiotic stress[J]. Acta Horticulturae Sinica,2021,48(4):632-646 岳玲琦,邢巧娟,张晓兰,等. 光敏色素互作因子在植物抵御逆境胁迫中的作用研究进展[J]. 园艺学报,2021,48(4):632-646 SHICHIJO C,KATADA K,TANAKA O,et al. Phytochrome A-mediated inhibition of seed germination in tomato[J]. Planta,2001,213(5):764-769 JOHNSON E,BRADLEY M,HARBERD N P,et al. Photo responses of light-grown phyA mutants of Arabidopsis (phytochrome A is required for the perception of daylength extensions) [J]. Plant physiology,1994,105(1):141-149 WEI S,XU X H,LU X B,et al. The rice phytochrome genes,PHYA and PHYB have synergistic effects on anther development and pollen viability[J]. Scientific Reports,2017,7:6439 KIRCHER S,GIL P,KOZMA-BOGNÁR L,et al. Nucleocytoplasmic partitioning of the plant photoreceptors phytochrome A,B,C,D,and E is regulated differentially by light and exhibits a diurnal rhythm[J]. The Plant Cell,2002,14(7):1541-1555 LI Y K. Functional analysis of three photosensitive pigment genes in rye during photomorphogenesis[D]. Zhengzhou: Henan Agricultural University,2025:27-39 李彦坤.黑麦3个光敏色素基因在光形态建成中的功能分析[D]. 郑州:河南农业大学,2025:27-39 HISADA A,HANZAWA H,WELLER J L,et al. Light-induced nuclear translocation of endogenous pea phytochrome A visualized by immunocytochemical procedures[J]. The Plant Cell,2000,12(7):1063-1078 RÖSLER J,KLEIN I,ZEIDLER M. Arabidopsis fhl/fhy1 double mutant reveals a distinct cytoplasmic action of phytochrome A[J]. Proceedings of the National Academy of Sciences of the United States of America,2007,104(25):10737-10742 HAN Y J,KIM H S,KIM Y M,et al. Functional characterization of phytochrome autophosphorylation in plant light signaling[J]. Plant and Cell Physiology,2010,51(4):596-609 ROCKWELL N C,SU Y S,LAGARIAS J C. Phytochrome structure and signaling mechanisms[J]. Annual Review of Plant Biology,2006,57(1): 837-858 XU J W,YOU Y S,XU Y W,et al. Research progress on the function of PIFs transcription factors[J]. Protection Forest Science and Technology,2025(4):74-80 许建闻,由云生,徐亚伟,等.PIFs转录因子功能研究进展[J]. 防护林科技,2025(4):74-80 WANG F. Roles and mechanisms of phyA-,HY5- and PIF4- mediated light quality-regulated cold tolerance in tomato[D]. Hangzhou: Zhejiang University,2017:19-55 王峰. PhyA、HY5和PIF4在光质调控番茄低温抗性中的机制研究[D]. 杭州:浙江大学,2017:19-55 JIA X P,ZHANG B,HE Z X,et al. The responsive characteristics of phytochrome genes to photoperiod,abiotic stresses and identification of their key natural variation sites in foxtail millet (Setaria italica L.) [J]. Chinese Journal of Biotechnology,2022,38(5):1929-1945 贾小平,张博,何占祥,等. 谷子光敏色素基因光周期、非生物胁迫响应特性及关键自然变异位点鉴定[J]. 生物工程学报,2022,38(5):1929-1945 ABDELLATIF I M Y,YUAN S Z,YOSHIHARA S,et al. Stimulation of tomato drought tolerance by PHYTOCHROME A and B1B2 mutations[J]. International Journal of Molecular Sciences,2023,24(2):1560 GONZÁLEZ C V,IBARRA S E,PICCOLI P N,et al. Phytochrome B increases drought tolerance by enhancing ABA sensitivity in Arabidopsis thaliana[J]. Plant,Cell & Environment,2012,35(11):1958-1968 PENG Y X,ZHU H Y,WANG Y T,et al. Revisiting the role of light signaling in plant responses to salt stress[J]. Horticulture Research,2025,12:uhae26 LI H,ZHOU Y Y,QIN X Y,et al. Reconstitution of phytochrome A-mediated light modulation of the ABA signaling pathways in yeast[J]. Proceedings of the National Academy of Sciences of the United States of America,2023,120(34):e2302901120 OLSEN J E,JUNTTILA O,NILSEN J,et al. Ectopic expression of oat phytochrome A in hybrid aspen changes critical daylength for growth and prevents cold acclimatization[J]. The Plant Journal,1997,12(6):1339-1350 FAN W N,REN B B,SHI P F,et al. Advance in alfalfa of phytochrome A regulating fall dormancy[J]. China Feed,2022,1(5):112-116 樊文娜,任冰冰,史鹏飞,等. 光敏色素A参与苜蓿秋眠调控分子机制的研究进展[J]. 中国饲料,2022,1(5):112-116 NIETO C,LÓPEZ-SALMERÓN V,DAVIÈRE J M,et al. ELF3-PIF4 interaction regulates plant growth independently of the evening complex[J]. Current Biology,2015,25(2):187-193 YANG T,ZHANG H,JIANG X P,et al. Phytochrome alleviates cadmium toxicity by regulating gibberellic acid and brassinolide in Nicotiana tabacum[J]. Plant Physiology and Biochemistry,2025,219:109375