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phosphorylation of the LCB1 subunit of Arabidopsis serine palmitoyltransferase stimulates its activity and modulates sphingolipid biosynthesis

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Journal of Integrative Plant Biology 2023年第6期65卷 1585-1601页

作者： Yuan Li Hanwei Cao Tingting Dong Xiaoke Wang Liang Ma Kun Li Huiqiang Lou Chun-Peng Song Dongtao Ren State Key Laboratory of Plant Environmental Resilience College of Biological SciencesChina Agricultural UniversityBeijing 100193China State Key Laboratory of Agro-Biotechnology and Beijing Advanced Innovation Center for Food Nutrition and Human Health College of Biological SciencesChina Agricultural UniversityBeijing 100193China Collaborative Innovation Center of Crop Stress Biology Henan Province.Institute of Plant Stress BiologySchool of Life ScienceHenan UniversityKaifeng 475001China

Sphingolipids are the structural components of membrane lipid bilayers and act as signaling molecules in many cellular processes.Serine palmitoyltransferase(SPT)is the first committed and rate-limiting enzyme in the de novo sphingolipids biosynthetic pathway.The core SPT enzyme is a heterodimer consisting of LONG-CHAIN BASE1(LCB1)and LCB2 subunits.SPT activity is inhibited by orosomucoid proteins and stimulated by small subunits of SPT(ssSPTs).However,whether LCB1 is modified and how such modification might regulate SPT activity have to date been unclear.Here,we show that activation of MITOGEN-ACTIVATED PROTEIN KINASE 3(MPK3)and MPK6 by upstream MKK9 and treatment with Flg22(a pathogen-associated molecular pattern)increases SPT activity and induces the accumulation of sphingosine long-chain base t18:0 in Arabidopsis thaliana,with activated MPK3and MPK6 phosphorylating AtLCB1.phosphorylation of AtLCB1 strengthened its binding with AtLCB2b,promoted its binding with ssSPTs,and stimulated the formation of higher order oligomeric and active SPT complexes.Our findings therefore suggest a novel regulatory mechanism for SPT activity.

关键词： Arabidopsis MAPK phosphorylation sphingolipids

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phosphorylation of a wheat aquaporin at two sites enhances both plant growth and defense

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Molecular Plant 2022年第11期15卷 1772-1789页

作者： Kai Lu Xiaochen Chen Xiaohui Yao Yuyan An Xuan Wang Lina Qin Xiaoxu Li Zuodong Wang Shuo Liu Zhimao Sun Liyuan Zhang Lei Chen Baoyan Li Baoyou Liu Weiyang Wang Xinhua Ding Yonghua Yang Meixiang Zhang Shenshen Zou Hansong Dong College of Plant Protection State Key Laboratory of Crop BiologyQilu CollegeShandong Agricultural UniversityTaian 271018China College of Life Sciences Shaanxi Normal UniversityXi’an 710019China Institute of Plant Molecular Biology State Key Laboratory of Pharmaceutical BiotechnologySchool of Life SciencesNanjing UniversityNanjing 210023China nstitute of Plant Protection&Resource and Environment Yantai Academy of Agricultural SciencesYantai 265599China

Eukaryotic aquaporins share the characteristic of functional multiplicity in transporting distinct substrates and regulating various processes,but the underlying molecular basis for this is largely unknown.Here,we report that the wheat(Triticum aestivum)aquaporin TaPIP2;10 undergoes phosphorylation to promote photosynthesis and productivity and to confer innate immunity against pathogens and a generalist aphid pest.In response to elevated atmospheric CO_(2)concentrations,TaPIP2;10 is phosphorylated at the serine residue S280 and thereafter transports CO_(2)into wheat cells,resulting in enhanced photosynthesis and increased grain yield.In response to apoplastic H_(2)O_(2) induced by pathogen or insect attacks,TaPIP2;10 is phosphorylated at S121 and this phosphorylated form transports H_(2)O_(2) into the cytoplasm,where H_(2)O_(2)intensifies host defenses,restricting further attacks.Wheat resistance and grain yield could be simultaneously increased by TaPIP2;10 overexpression or by expressing a TaPIP2;10 phosphomimic with aspartic acid substitutions at S121 and S280,thereby improving both crop productivity and immunity.

关键词： aquaporin phosphorylation gating photosynthesis immunity growth-defense tradeoffs

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phosphorylation of Ago2 is required for its role in DNA double-strand break repair

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Journal of Genetics and Genomics 2021年第4期48卷 333-340页

作者： Xiaolu Hu Yan Li Tianfang Zhang Lin Li She Chen Xiaohong Wu Haijun Li Binjie Qi Zuobing Chen Department of Rehabilitation Medicine The First Affiliated HospitalCollege of MedicineZhejiang UniversityHangzhou 310000China Center for Plant Biology School of Life SciencesTsinghua UniversityBeijing 100084China Tsinghua-Peking Center for Life Sciences Beijing 100084China National Institute of Biological Sciences Zhongguancun Life Science ParkBeijing 102206China

Repair of DNA double-strand break(DSB)is critical for the maintenance of genome integrity.A class of DSB-induced small RNAs(di RNAs)has been shown to play an important role in DSB repair.In humans,di RNAs are associated with Ago2 and guide the recruitment of Rad51 to DSB sites to facilitate repair by homologous recombination(HR).Ago2 activity has been reported to be regulated by phosphorylation under normal and hypoxic conditions.However,the role of Ago2 phosphorylation in DNA damage repair is unexplored.Here,we show that S672,S828,T830,and S831 of human Ago2 are phosphorylated in response to ionizing radiation(IR).S672 A mutation of Ago2 leads to significant reduction in Rad51 foci formation and HR efficiency.We further show that defective association of Ago2 S672 A variant with DSB sites,instead of defects in di RNA and Rad51 binding,may account for decreased Rad51 foci formation and HR efficiency.Our study reveals a novel regulatory mechanism for the function of Ago2 in DNA repair.

关键词： Ago2 diRNA DNA repair phosphorylation Small RNA

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phosphorylation regulates cullin-based ubiquitination in tumorigenesis

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Acta Pharmaceutica Sinica B 2021年第2期11卷 309-321页

作者： Yifan Chen Xuejing Shao Ji Cao Hong Zhu Bo Yang Qiaojun He Meidan Ying Zhejiang Province Key Laboratory of Anti-Cancer Drug Research College of Pharmaceutical SciencesZhejiang UniversityHangzhou 310058China

Cullin-RING ligases(CRLs)recognize and interact with substrates for ubiquitination and degradation,and can be targeted for disease treatment when the abnormal expression of substrates involves pathologic processes.phosphorylation,either of substrates or receptors of CRLs,can alter their interaction.phosphorylation-dependent ubiquitination and proteasome degradation influence various cellular processes and can contribute to the occurrence of various diseases,most often tumorigenesis.These processes have the potential to be used for tumor intervention through the regulation of the activities of related kinases,along with the regulation of the stability of specific oncoproteins and tumor suppressors.This review describes the mechanisms and biological functions of crosstalk between phosphorylation and ubiquitination,and most importantly its influence on tumorigenesis,to provide new directions and strategies for tumor therapy.

关键词： phosphorylation Cullin-RING ligases Ubiquitination Tumorigenesis Targeted therapy Degradation Crosstalk Kinases

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phosphorylation of OsFD1 by OsCIPK3 promotes the formation of RFT1-containing florigen activation complex for long-day flowering in rice

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Molecular Plant 2021年第7期14卷 1135-1148页

作者： Qiang Peng Chunmei Zhu Tao Liu Shuo Zhang Shijing Feng Changyin Wu National Key Laboratory of Crop Genetic Improvement National Center of Plant Gene Research(Wuhan)Huazhong Agricultural UniversityWuhan 430070China Guizhou Rice Research Institute Guizhou Academy of Agricultural SciencesGuiyang 550006China

Heading date is a critical trait that determines the regional adaptability and grain productivity of many crops.Although rice is a facultative short-day plant,its domestication led to the Ghd7-Ehd1-Hd3a/RFT1 pathway for adaptation to long-day conditions(LDs).The formation of the"florigen activation complex"(FAC)containing florigen Hd3a has been characterized.However,the molecular composition of the FAC that contains RFT1 for long-day flowering is unclear.We show here that RFT1 forms a ternary FAC with 14-3-3 proteins and OsFD1 to promote flowering under LDs.We identified a calcineurin B-like-interacting protein kinase,OsCIPK3,which directly interacts with and phosphorylates OsFD1,thereby facilitating the localization of the FAC to the nucleus.Mutation in OsCIPK3 results in a late heading date under LDs but a normal heading date under short-day conditions.Collectively,our results suggest that OsCIPK3 phosphorylates OsFD1 to promote RFT1-containing FAC formation and consequently induce flowering in rice under LDs.

关键词： rice heading date florigen activation complex OsCIPK3 phosphorylation

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phosphorylation of PLIN3 by AMPK promotes dispersion of lipid droplets during starvation

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Protein & Cell 2019年第5期10卷 382-387页

作者： Jianxi Zhu Mingyang Xu Yi Liu Lisha Zhuang Kejun Ying Feng Liu Dan Liu Wenbin Ma Zhou Songyang MOE Key Laboratory of Gene Function and Regulation State Key Laboratory for BiocontrolSchool of Life SciencesSun Yat-sen UniversityGuangzhou 510275China Guangzhou Regenerative Medicine and Health-Guangdong Laboratory(GRMH-GDL) Guangzhou 510530China Verna and Marrs McLean Department of Biochemistry and Molecular Biology Baylor College of MedicineOne Baylor PlazaHoustonTX 77030USA

Dear Editor,Lipid droplets(LDs)are dynamic lipid-storage organelles of storage depots and sources of essential substrates for myriad cellular processes and protect cells from lipotoxicity(Ohsaki et al.,2006).Disrupted LD and fat storage homeostasis has been linked to metabolic diseases such as atherosclerosis,obesity,and type II diabetes(Levin et al.,2001).Structurally,the core of neutral lipids in LDs is surroun ded by a phospholipid mono layer and coated with specific proteins(Storey et al.,2011).Perilipin family of proteins are the predominant LD-associated proteins.

关键词： phosphorylation PLIN3 AMPK

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phosphorylation of PB2 at serine 181 restricts viral replication and virulence of the highly pathogenic H5N1 avian influenza virus in mice

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Virologica Sinica 2024年第1期39卷 97-112页

作者： Jiao Hu Zixiong Zeng Xia Chen Manyu Zhang Zenglei Hu Min Gu Xiaoquan Wang Ruyi Gao Shunlin Hu Yu Chen Xiaowen Liu Daxin Peng Xiufan Liu Animal Infectious Disease Laboratory School of Veterinary MedicineYangzhou UniversityYangzhou225009China Joint International Research Laboratory of Agriculture and Agri-Product Safety The Ministry of Education of ChinaYangzhou University225009China Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonosis Yangzhou UniversityYangzhou225009China Key Laboratory of Prevention and Control of Biological Hazard Factors(Animal Origin)for Agri-food Safety and Quality Ministry of Agriculture of China(26116120)Yangzhou UniversityYangzhou225009China

Influenza A virus(IAV)continues to pose a pandemic threat to public health,resulting a high mortality rate annually and during pandemic years.Posttranslational modification of viral protein plays a substantial role in regulating IAV infection.Here,based on immunoprecipitation(IP)-based mass spectrometry(MS)and purified virus-coupled MS,a total of 89 phosphorylation sites distributed among 10 encoded viral proteins of IAV were identified,including 60 novel phosphorylation sites.Additionally,for the first time,we provide evidence that PB2 can also be acetylated at site K187.Notably,the PB2 S181 phosphorylation site was consistently identified in both IP-based MS and purified virus-based MS.Both S181 and K187 are exposed on the surface of the PB2 protein and are highly conserved in various IAV strains,suggesting their fundamental importance in the IAV life cycle.Bioinformatic analysis results demonstrated that S181E/A and K187Q/R mimic mutations do not significantly alter the PB2 protein structure.While continuous phosphorylation mimicked by the PB2 S181E mutation substantially decreases viral fitness in mice,PB2 K187Q mimetic acetylation slightly enhances viral virulence in mice.Mechanistically,PB2 S181E substantially impairs viral polymerase activity and viral replication,remarkably dampens protein stability and nuclear accumulation of PB2,and significantly weakens IAV-induced inflammatory responses.Therefore,our study further enriches the database of phosphorylation and acetylation sites of influenza viral proteins,laying a foundation for subsequent mechanistic studies.Meanwhile,the unraveled antiviral effect of PB2 S181E mimetic phosphorylation may provide a new target for the subsequent study of antiviral drugs.

关键词： H5N1 influenza virus PB2 phosphorylation Acetylation Viral fitness Mice

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Sucrose-associated SnRK1a1-mediated phosphorylation of Opaque2 modulates endosperm filling in maize

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Molecular Plant 2024年第5期17卷 788-806页

作者： Tao Yang Yunqin Huang Longyu Liao Shanshan Wang Haoyu Zhang Jingying Pan Yongcai Huang Xiaoling Li Di Chen Tao Liu Xiaoduo Lu Yongrui Wu State Key Laboratory of Crop Gene Exploration and Utilization in Southwest China Sichuan Agricultural UniversityChengdu 611130SichuanChina National Key Laboratory of Plant Molecular Genetics CAS Center for Excellence in Molecular Plant SciencesShanghai lnstitute of Plant Physiology and EcologyChinese Academy of SciencesShanghai 200032China Institute of Molecular Breeding for Maize Qilu Normal UniversityJinanChina

During maize endosperm filling,sucrose not only serves as a source of carbon skeletons for storage-reserve synthesis but also acts as a stimulus to promote this process.However,the molecular mechanisms underlying sucrose and endosperm filling are poorly understood.In this study,we found that sucrose promotes the expression of endosperm-filling hub gene Opaque2(O2),coordinating with storage-reserve accumulation.We showed that the protein kinase SnRK1a1 can attenuate O2-mediated transactivation,but sucrose can release this suppression.Biochemical assays revealed that SnRK1a1 phosphorylates O2 at serine 41(S41),negatively affecting its protein stability and transactivation ability.We observed that mutation of SnRK1a1 results in larger seeds with increased kernel weight and storage reserves,while overexpression of SnRK1a1 causes the opposite effect.Overexpression of the native O2(O2-OE),phospho-dead(O2-SA),and phospho-mimetic(O2-SD)variants all increased 100-kernel weight.Although O2-SA seeds exhibit smaller kernel size,they have higher accumulation of starch and proteins,resulting in larger vitreous endosperm and increased test weight.O2-SD seeds display larger kernel size but unchanged levels of storage reserves and test weight.O2-OE seeds show elevated kernel dimensions and nutrient storage,like a mixture of O2-SA and O2-SD seeds.Collectively,our study discovers a novel regulatory mechanism of maize endosperm filling.Identification of S41 as a SnRK1-mediated phosphorylation site in O2 offers a potential engineering target for enhancing storage-reserve accumulation and yield in maize.

关键词： maize Opaque2 SnRK1 endosperm phosphorylation sucrose

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phosphorylation of sarcoplasmic and myofibrillar proteins in three ovine muscles during postmortem ageing

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Journal of Integrative Agriculture 2019年第7期18卷 1643-1651页

作者： WANG Ying LI Xin LI Zheng DU Man-ting ZHU Jie ZHANG She-qi ZHANG De-quan Laboratory of Biomechanics and Engineering Institute of Biophysics and College of ScienceNorthwest A&F UniversityYangling 712100P.R. China Institute of Food Science and Technology Chinese Academy of Agricultural Sciences/Key Laboratory of Agro-Products ProcessingMinistry of AgricultureBeijing 100193P.R. China

This study aimed to examine changes in phosphorylation of sarcoplasmic and myofibrillar proteins from longissimus lumborum,semitendinosus,and psoas major muscles during postmortem ageing for 5 d.These sarcoplasmic and myofibrillar proteins were separated using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and stained with phosphorous and protein specific stains.Myofibril fragmentation index,pH,the content of lactic acid and the relative activity of μ-calpain in three ovine muscles were measured.These results showed that the relative phosphorylation level of sarcoplasmic and myofibrillar proteins of psoas major muscle were lower compared with longissimus lumborum and semitendinosus muscles(Pphosphorylation may regulate the rate of pH decline to influence the μ-calpain activity and then proteolysis of proteins consequently.This study gives a new perspective of the mechanism of postmortem meat tenderization.

关键词： ovine muscle protein phosphorylation μ-calpain myofibril fragmentation index

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phosphorylation of Serine 186 of bHLH Transcription Factor SPEECHLESS Promotes Stomatal Development in Arabidopsis

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Molecular Plant 2015年第5期8卷 783-795页

作者： Ke-Zhen Yang Min Jiang Ming Wang Shan Xue Ling-Ling Zhu Hong-Zhe Wang Jun-Jie Zou Eun-Kyoung Lee Fred Sack Jie Le Key Laboratory of Plant Molecular Physiology Institute of Botany Chinese Academy of Sciences 20 Nanxincun Beijing 100093 China Department of Botany University of British Columbia Vancouver British Columbia V6T 1Z4 Canada

The initiation of stomatal lineage and subsequent asymmetric divisions in Arabidopsis require the activity of the basic helix-loop-helix transcription factor SPEECHLESS （SPCH）. It has been shown that SPCH controls entry into the stomatal lineage as a substrate either of the MITOGEN-ACTIVATED PROTEIN KINASE （MAPK） cascade or GSK3-1ike kinase BRASSlNOSTEROID INSENSITIVE 2 （BIN2）. Here we show that three serine residues of SPCH appear to be the primary phosphorylation targets of Cyclin-Dependent Kinases A;1 （CDKA;1） in vitro, and among them Serine 186 plays a crucial role in stomatal formation. Expression of an SPCH construct harboring a mutation that results in phosphorylation deficiencies on Serine 186 residue failed to rescue stomatal defects in spch null mutants. Expression of a phosphorylation-mimic mutant SPCHS186D complemented stomatal production defects in the transgenic lines harboring the targeted expression of dominant-negative CDKA;1.N146. Therefore, in addition to MAPK- and BIN2-mediated phos- phorylation on SPCH, phosphorylation at Serine 186 is positively required for SPCH function in regulating stomatal development.

关键词： Arabidopsis stomata development transcription factor phosphorylation

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