Title : The identification of N-glycosylated
residues of the human
5-HT3B receptor subunit : importance for cell membrane expression
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