PMID:12944413

Citation	Ozaslan, D, Wang, S, Ahmed, BA, Kocabas, AM, McCastlain, JC, Bene, A and Kilic, F (2003) Glycosyl modification facilitates homo- and hetero-oligomerization of the serotonin transporter. A specific role for sialic acid residues. J. Biol. Chem. 278:43991-4000
Abstract	The serotonin transporter (SERT) is an oligomeric glycoprotein with two sialic acid residues on each of two complex oligosaccharide molecules. In this study, we investigated the contribution of N-glycosyl modification to the structure and function of SERT in two model systems: wild-type SERT expressed in sialic acid-defective Lec4 Chinese hamster ovary (CHO) cells and a mutant form (after site-directed mutagenesis of Asn-208 and Asn-217 to Gln) of SERT, QQ, expressed in parental CHO cells. In both systems, SERT monomers required modification with both complex oligosaccharide residues to associate with each other and to function in homo-oligomeric forms. However, defects in sialylated N-glycans did not alter surface expression of the SERT protein. Furthermore, in heterologous (CHO and Lec4 cells) and endogenous (placental choriocarcinoma JAR cells) expression systems, we tested whether glycosyl modification also manipulates the hetero-oligomeric interactions of SERT, specifically with myosin IIA. SERT is phosphorylated by cGMP-dependent protein kinase G through interactions with anchoring proteins, and myosin is a protein kinase G-anchoring protein. A physical interaction between myosin and SERT was apparent; however, defects in sialylated N-glycans impaired association of SERT with myosin as well as the stimulation of the serotonin uptake function in the cGMP-dependent pathway. We propose that sialylated N-glycans provide a favorable conformation to SERT that allows the transporter to function most efficiently via its protein-protein interactions.
Links	PubMed PMC3042025 Online version:10.1074/jbc.M306360200
Keywords	Animals; CHO Cells; Carrier Proteins/chemistry; Carrier Proteins/genetics; Carrier Proteins/metabolism; Choriocarcinoma; Cricetinae; Cyclic GMP/pharmacology; Cyclic GMP-Dependent Protein Kinases/metabolism; Female; Gene Expression; Glycosylation; Macromolecular Substances; Membrane Glycoproteins/chemistry; Membrane Glycoproteins/genetics; Membrane Glycoproteins/metabolism; Membrane Transport Proteins; Mutagenesis, Site-Directed; N-Acetylneuraminic Acid/chemistry; N-Acetylneuraminic Acid/genetics; Nerve Tissue Proteins; Nonmuscle Myosin Type IIA/genetics; Nonmuscle Myosin Type IIA/metabolism; Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase/metabolism; Phosphorylation; Pregnancy; Protein Conformation; Rats; Serotonin/metabolism; Serotonin Plasma Membrane Transport Proteins; Structure-Activity Relationship; Transfection; Tumor Cells, Cultured; Uterine Neoplasms
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
RAT:SC6A4	enables	GO:0042803: protein homodimerization activity	ECO:0000315: mutant phenotype evidence used in manual assertion		F		Seeded From UniProt	complete
RAT:SC6A4	involved_in	GO:0051610: serotonin uptake	ECO:0000315: mutant phenotype evidence used in manual assertion		P		Seeded From UniProt	complete
RAT:SC6A4	enables	GO:0017022: myosin binding	ECO:0000353: physical interaction evidence used in manual assertion	UniProtKB:Q62812	F		Seeded From UniProt	complete
RAT:SC6A4	involved_in	GO:0051260: protein homooligomerization	ECO:0000314: direct assay evidence used in manual assertion		P		Seeded From UniProt	complete

References

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