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Co-expression of ?2 subunits hinders processing of N-linked glycans attached to the N104 glycosylation sites of GABAA receptor ?2 subunits.

ABSTRACT: GABAA receptors, the major mediators of fast inhibitory neuronal transmission, are heteropentameric glycoproteins assembled from a panel of subunits, usually including ? and ? subunits with or without a ?2 subunit. The ?1?2?2 receptor is the most abundant GABAA receptor in brain. Co-expression of ?2 with ?1 and ?2 subunits causes conformational changes, increases GABAA receptor channel conductance, and prolongs channel open times. We reported previously that glycosylation of the three ?2 subunit glycosylation sites, N32, N104 and N173, was important for ?1?2 receptor channel gating. Here, we examined the hypothesis that steric effects or conformational changes caused by ?2 subunit co-expression alter the glycosylation of partnering ?2 subunits. We found that co-expression of ?2 subunits hindered processing of ?2 subunit N104 N-glycans in HEK293T cells. This ?2 subunit-dependent effect was strong enough that a decrease of ?2 subunit expression in heterozygous GABRG2 knockout (?2(+/-)) mice led to appreciable changes in the endoglycosidase H digestion pattern of neuronal ?2 subunits. Interestingly, as measured by flow cytometry, ?2 subunit surface levels were decreased by mutating each of the ?2 subunit glycosylation sites. The ?2 subunit mutation N104Q also decreased GABA potency to evoke macroscopic currents and reduced conductance, mean open time and open probability of single channel currents. Collectively, our data suggested that ?2 subunits interacted with ?2 subunit N-glycans and/or subdomains containing the glycosylation sites, and that ?2 subunit co-expression-dependent alterations in the processing of the ?2 subunit N104 N-glycans were involved in altering the function of surface GABAA receptors.

SUBMITTER: Lo WY

PROVIDER: S-EPMC4018433 | biostudies-literature | 2014 Jun

REPOSITORIES: biostudies-literature

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Co-expression of γ2 subunits hinders processing of N-linked glycans attached to the N104 glycosylation sites of GABAA receptor β2 subunits.

Lo Wen-Yi WY Lagrange Andre H AH Hernandez Ciria C CC Gurba Katharine N KN Macdonald Robert L RL

Neurochemical research 20131112 6

GABAA receptors, the major mediators of fast inhibitory neuronal transmission, are heteropentameric glycoproteins assembled from a panel of subunits, usually including α and β subunits with or without a γ2 subunit. The α1β2γ2 receptor is the most abundant GABAA receptor in brain. Co-expression of γ2 with α1 and β2 subunits causes conformational changes, increases GABAA receptor channel conductance, and prolongs channel open times. We reported previously that glycosylation of the three β2 subunit ...[more]

PMID: 24213971

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Project description:The addition of asparagine (N)-linked polysaccharide chains (i.e., glycans) to the gp120 and gp41 glycoproteins of human immunodeficiency virus type 1 (HIV-1) envelope is not only required for correct protein folding, but also may provide protection against neutralizing antibodies as a "glycan shield." As a result, strong host-specific selection is frequently associated with codon positions where nonsynonymous substitutions can create or disrupt potential N-linked glycosylation sites (PNGSs). Moreover, empirical data suggest that the individual contribution of PNGSs to the neutralization sensitivity or infectivity of HIV-1 may be critically dependent on the presence or absence of other PNGSs in the envelope sequence. Here we evaluate how glycan-glycan interactions have shaped the evolution of HIV-1 envelope sequences by analyzing the distribution of PNGSs in a large-sequence alignment. Using a "covarion"-type phylogenetic model, we find that the rates at which individual PNGSs are gained or lost vary significantly over time, suggesting that the selective advantage of having a PNGS may depend on the presence or absence of other PNGSs in the sequence. Consequently, we identify specific interactions between PNGSs in the alignment using a new paired-character phylogenetic model of evolution, and a Bayesian graphical model. Despite the fundamental differences between these two methods, several interactions are jointly identified by both. Mapping these interactions onto a structural model of HIV-1 gp120 reveals that negative (exclusive) interactions occur significantly more often between colocalized glycans, while positive (inclusive) interactions are restricted to more distant glycans. Our results imply that the adaptive repertoire of alternative configurations in the HIV-1 glycan shield is limited by functional interactions between the N-linked glycans. This represents a potential vulnerability of rapidly evolving HIV-1 populations that may provide useful glycan-based targets for neutralizing antibodies.

Dataset Information

Co-expression of ?2 subunits hinders processing of N-linked glycans attached to the N104 glycosylation sites of GABAA receptor ?2 subunits.

Publications

Co-expression of γ2 subunits hinders processing of N-linked glycans attached to the N104 glycosylation sites of GABAA receptor β2 subunits.

Similar Datasets

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