title: Proteomic mapping of ER-PM junctions identifies STIMATE as a regulator of Ca²⁺ influx.
abstract: Specialized junctional sites that connect the plasma membrane (PM) and endoplasmic reticulum (ER) play critical roles in controlling lipid metabolism and Ca(2+) signalling. Store-operated Ca(2+) entry mediated by dynamic STIM1-ORAI1 coupling represents a classical molecular event occurring at ER-PM junctions, but the protein composition and how previously unrecognized protein regulators facilitate this process remain ill-defined. Using a combination of spatially restricted biotin labelling in situ coupled with mass spectrometry and a secondary screen based on bimolecular fluorescence complementation, we mapped the proteome of intact ER-PM junctions in living cells without disrupting their architectural integrity. Our approaches led to the discovery of an ER-resident multi-transmembrane protein that we call STIMATE (STIM-activating enhancer, encoded by TMEM110) as a positive regulator of Ca(2+) influx in vertebrates. STIMATE physically interacts with STIM1 to promote STIM1 conformational switch. Genetic depletion of STIMATE substantially reduces STIM1 puncta formation at ER-PM junctions and suppresses the Ca(2+)-NFAT signalling. Our findings enable further genetic studies to elucidate the function of STIMATE in normal physiology and disease, and set the stage to uncover more uncharted functions of hitherto underexplored ER-PM junctions.
authors: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, Texas 77030, USA.
authors: Ji
authors: J
authors: Jing
authors: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, Texas 77030, USA.
authors: Lian
authors: L
authors: He
authors: Beijing Key Laboratory of Gene Resource and Molecular Development, College of Life Sciences, Beijing Normal University, Beijing 100875, China.
authors: Aomin
authors: A
authors: Sun
authors: Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, California 92037, USA.
authors: Ariel
authors: A
authors: Quintana
authors: National Institute of Biological Sciences, Beijing 102206, China.
authors: Yuehe
authors: Y
authors: Ding
authors: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, Texas 77030, USA.
authors: Guolin
authors: G
authors: Ma
authors: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, Texas 77030, USA.
authors: Peng
authors: P
authors: Tan
authors: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, Texas 77030, USA.
authors: Xiaowen
authors: X
authors: Liang
authors: Institute of Molecular Medicine, Peking University, Beijing 100871, China.
authors: Xiaolu
authors: X
authors: Zheng
authors: Institute of Molecular Medicine, Peking University, Beijing 100871, China.
authors: Liangyi
authors: L
authors: Chen
authors: Department of Chemistry, West Virginia University, Morgantown, West Virginia 26506, USA.
authors: Xiaodong
authors: X
authors: Shi
authors: Department of Medical Physiology, College of Medicine, Texas A&M University Health Science Center, Temple, Texas 76504, USA.
authors: Shenyuan L
authors: SL
authors: Zhang
authors: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, Texas 77030, USA.
authors: Ling
authors: L
authors: Zhong
authors: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, Texas 77030, USA.
authors: Yun
authors: Y
authors: Huang
authors: National Institute of Biological Sciences, Beijing 102206, China.
authors: Meng-Qiu
authors: MQ
authors: Dong
authors: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, Texas 77030, USA.
authors: Cheryl L
authors: CL
authors: Walker
authors: Division of Signaling and Gene Expression, La Jolla Institute for Allergy and Immunology, La Jolla, California 92037, USA.
authors: Patrick G
authors: PG
authors: Hogan
authors: Beijing Key Laboratory of Gene Resource and Molecular Development, College of Life Sciences, Beijing Normal University, Beijing 100875, China.
authors: Youjun
authors: Y
authors: Wang
authors: Institute of Biosciences and Technology, Texas A&M University Health Science Center, Houston, Texas 77030, USA.|Department of Medical Physiology, College of Medicine, Texas A&M University Health Science Center, Temple, Texas 76504, USA.
authors: Yubin
authors: Y
authors: Zhou
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D019556:COS Cells; D002118:Calcium / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D002522:Chlorocebus aethiops; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D020869:Gene Expression Profiling; D020868:Gene Silencing; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence / Q000379:methods; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins / Q000235:genetics / Q000378:metabolism; D020543:Proteome / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D000071737:Stromal Interaction Molecule 1
chemical_list: D008565:Membrane Proteins; D009363:Neoplasm Proteins; D020543:Proteome; C103530:STIM1 protein, human; C000604313:STIMATE protein, human; D000071737:Stromal Interaction Molecule 1; D002118:Calcium