Stromal interaction molecule 1 (STIM1), a transmembrane protein with growth suppressor activity, contains an extracellular SAM domain modified by N-linked glycosylation.
Stromal interaction molecule 1 (STIM1) is a cell surface transmembrane glycoprotein implicated in tumour growth control and stromal-haematopoietic cell interactions. A single sterile alpha motif (SAM) protein-protein interaction domain is modelled within its extracellular region, a subcellular localisation not previously described for other SAM domain-containing proteins. We have defined the transmembrane topology of STIM1 by determining the sites of N-linked glycosylation. We have confirmed that STIM1 is modified by N-linked glycosylation at two sites within the SAM domain itself, deduced as asparagine residues N131 and N171, demonstrating that STIM1 is translocated across the membrane of the endoplasmic reticulum such that the SAM domain resides within the endoplasmic reticulum (ER) lumen. Both N-linked oligosaccharides remain endoglycosidase H-sensitive, indicating absence of full processing within the ER and Golgi. This immature modification is nevertheless sufficient and critical for cell surface expression of STIM1. We show that STIM1-STIM1 homotypic interactions are mediated via the cytoplasmic rather than the extracellular region of STIM1, excluding an essential role for the SAM domain in these protein interactions. These studies provide the first evidence for an extracellular localisation of a SAM domain within any protein, and the first example of a SAM domain modified by N-linked glycosylation.
Williams
Richard T
RT
Department of Anatomy and Cell Biology, University of Melbourne, Melbourne, Victoria 3010, Australia.
Senior
Paul V
PV
Van Stekelenburg
Leonie
L
Layton
Judith E
JE
Smith
Peter J
PJ
Dziadek
Marie A
MA
D003001:Cloning, Molecular; D003593:Cytoplasm / Q000737:chemistry; D005110:Extracellular Space / Q000737:chemistry; D005786:Gene Expression Regulation; D006031:Glycosylation; D016179:Granulocyte Colony-Stimulating Factor / Q000096:biosynthesis / Q000737:chemistry; D006801:Humans; D015151:Immunoblotting; D008565:Membrane Proteins; D016297:Mutagenesis, Site-Directed; D009363:Neoplasm Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D000071737:Stromal Interaction Molecule 1; D014162:Transfection
D008565:Membrane Proteins; D009363:Neoplasm Proteins; C103530:STIM1 protein, human; D000071737:Stromal Interaction Molecule 1; D016179:Granulocyte Colony-Stimulating Factor