Sar1-dependent trafficking of the human calcium receptor to the cell surface.
The molecular mechanisms underlying the exit from the endoplasmic reticulum (ER) for cell surface trafficking of the human calcium receptor (hCaR) remain poorly understood. We investigated the role of the Sar1 small GTP-binding protein in cell surface transport of the hCaR. Disruptions of endogenous Sar1 function with the constitutively active Sar1H79G mutant or depletion using small interfering RNA, attenuates cell surface expression of the hCaR. Mutation of several putative di-acidic ER export motifs in the carboxyl-tail of the receptor revealed no apparent defect in cell surface expression. Truncated mutants lacking most of the carboxyl-terminal sequences or all intracellular domains also showed no impairment in cell surface expression at steady state. A truncated receptor containing only the large amino-terminal extracellular ligand-binding domain (ECD) is secreted into the culture medium and Sar1H79G inhibits this secretion. ECD receptor variants with the cysteines essential for intermolecular disulfide-linked dimerization mutated to serine or four of the asparagine sites for N-glycosylation mutated to alanine also disrupt secretion, indicating proper ECD conformation is critical for forward transport of this receptor.
Zhuang
Xiaolei
X
Laboratory of Cellular Biology, NIDCD, National Institutes of Health, Bethesda, MD 20892, USA.
Chowdhury
Shoaib
S
Northup
John K
JK
Ray
Kausik
K
D000595:Amino Acid Sequence; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D020559:Monomeric GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000235:genetics; D044169:Receptors, Calcium-Sensing / Q000378:metabolism*; D022161:Transport Vesicles / Q000378:metabolism
D034741:RNA, Small Interfering; D044169:Receptors, Calcium-Sensing; C493083:SAR1A protein, human; D020559:Monomeric GTP-Binding Proteins
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