Alpha subunit variants of the human glycine receptor: primary structures, functional expression and chromosomal localization of the corresponding genes.

Abstract

Two cDNAs encoding variants (alpha 1 and alpha 2) of the strychnine binding subunit of the inhibitory glycine receptor (GlyR) were isolated from a human fetal brain cDNA library. The predicted amino acid sequences exhibit approximately 99% and approximately 76% identity to the previously characterized rat 48 kd polypeptide. Heterologous expression of the human alpha 1 and alpha 2 subunits in Xenopus oocytes resulted in the formation of glycine‐gated strychnine‐sensitive chloride channels, indicating that both polypeptides can form functional GlyRs. Using a panel of rodent‐human hybrid cell lines, the gene encoding alpha 2 was mapped to the short arm (Xp21.2‐p22.1) of the human X chromosome. In contrast, the alpha 1 subunit gene is autosomally located. These data indicate molecular heterogeneity of the human GlyR at the level of alpha subunit genes.

Coexpression of the receptor‐associated protein gephyrin changes the ligand binding affinities of α2 glycine receptors

“…Previous studies have demonstrated that the human GlyR CL? subunit forms strychnine-sensitive chloride channels upon heterologous expression in Xenopus oocytes [16]. Accordingly, large glycine-gated currents were revealed here in cotransfected cells by voltage-clamp recording (Fig.…”

“…The human CilyR a: [16] and the rat GlyR a, [18] and/3 [5] subunit cDNAs as well as the rat gephyrin pl cDNA [S]. each subcloned into the eukarydtic expression vector pCIS, were used to transfcct the human ombyronic kidney cell line 293 (ATCC CRL1573) by calcium phosphate precipitation as described [20].…”

“…Presently three a subunit variants, c+a3, have been characterized in detail [15-181, which all form glycine-gated chloride channels upon heterologous expression in Xerropus oocytes [16][17][18][19] or mammalian cell lines [20]. The pharmacology of these homooligomeric GlyRs is very similar to that found for in vivo receptors; however, higher agonist *Present address: Department of Physiology, Kanagawa Dental College, Yokosuka, Japan.…”

“…concentrations are required for channel gating. Moreover, in case of neonatal GlyRs, which contain cr, as the ligand binding subunit [17], significant differences in sensitivity to the glycinergic antagonist strychnine have been revealed by biochemical, electrophysiological and heterologous expression experiments [2,14,16,17]. These discrepancies may relate to the lack of additional subunits [3,5].…”

Agonist binding to purified glycine receptor reconstituted into giant liposomes elicits two types of chloride currents

“…For instance, the 48 kDa polypeptide is able to form homomerit channels when expressed in oocytes or mammalian cells [IS,191, and has also been shown to be phosphorylated in vitro by protein kinase C [20]. Furthermore, recent reports indicate the existence of several different mRNAs encoding subtypes of the 48 kDa GlyR subunit in both the neonatal and the adult rat spinal cord [l&21,22] and two different 48 kDa subunit genes located in different chromosomes have been recently detected in humans [23]. This expected molecular heterogeneity would be similar to the receptor subtype diversity found for other ion channel-linked neuroreceptors homologous to the GlyR, such as the GABAA and the neuronal nicotinic acetylcholine receptors (see [24] for a review), Further experiments are needed to ascertain the molecular basis of the existence of functionally different forms of the glycine receptor and to fully characterize the electrophysiology of the ionic channels involved.…”