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The identification of N ‐glycosylated residues of the human 5‐HT3B receptor subunit: importance for cell membrane expression
Author(s) -
Massoura Andrew N.,
Dover Terri J.,
Newman Amy S.,
Barnes Nicholas M.
Publication year - 2011
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.2010.07129.x
Subject(s) - protein subunit , identification (biology) , cell , receptor , expression (computer science) , cell membrane , microbiology and biotechnology , chemistry , biology , biochemistry , computational biology , gene , computer science , botany , programming language
J. Neurochem. (2011) 116 , 975–983. Abstract The 5‐hydroxytryptamine 3 (5‐HT 3 ) receptor is a pentameric ligand‐gated ion channel with potential molecular isoforms arising from different subunit combinations and/or different post‐translational modifications of the individual subunits. Since N ‐glycosylation of the 5‐HT3A subunit impacts cell surface trafficking, the presence of N ‐glycosylation of the human (h) 5‐HT3B subunit and the influence upon cell membrane expression was investigated. Following transient expression of the h5‐HT3B subunit by human embryonic kidney cells (HEK293 cells) stably expressing the h5‐HT3A subunit, the N ‐glycosylation inhibitor tunicamycin reduced the size of the predominant h5‐HT3B‐immunoreactive protein (∼ 55 kDa reduced to ∼ 40 kDa). Disruption of each consensus N ‐glycosylation sequences in the h5‐HT3B subunit (N31S, N75S, N117S, N147S and N182S) resulted in a reduced molecular weight (by ∼ 2–4 kDa) of each mutant when expressed by HEK293 cells stably expressing the h5‐HT3A subunit. Immunocytochemical studies demonstrated that disruption of each of the N ‐glycosylation sequences (individually or combined) reduced the expression of the mutant h5‐HT3B subunit protein in the cell membrane when co‐expressed with the h5‐HT3A subunit. The present study has identified utilised N ‐glycosylation sites of the h5‐HT3B subunit and demonstrated that they promote subunit expression in the cell membrane; a prerequisite for 5‐HT 3 receptor function.