The Human Glycine Receptor Subunit α3

Abstract

The neuronal glycine receptor is a ligand-gated chlo-ride channel composed of ligand binding ␣ and struc-tural ␤ polypeptides. Homology screening of a human fetal brain cDNA library resulted in the identification of two alternative splice variants of the glycine recep-tor ␣3 subunit. The amino acid sequence predicted for the ␣3L variant was largely identical to the corre-sponding rat subunit. In contrast, the novel splice var-iant ␣3K lacked the coding sequence for 15 amino acids located within the cytoplasmic loop connecting trans-membrane spanning region 3 (TM3) and TM4. Using P1 artificial chromosome (PAC) clones, the structure of the GLRA3 gene was elucidated and its locus assigned to human chromosomal bands 4q33-q34 by fluores-cence in situ hybridization. Two transcripts of 2.4 and 9 kilobases, corresponding to ␣3L and ␣3K, respec-tively, were identified and found to be widely distrib-uted throughout the human central nervous system. Structural analysis of the GLRA3 gene revealed that the ␣3K transcript resulted from a complex splice event where excision of the novel exon 8A comprising the alternative sequence of 45 base pairs coincides with the persistence of a large intronic sequence in the 3؅-untranslated region. Functional expression in HEK 293 cells of ␣3L and ␣3K subunits resulted in the for-mation of glycine-gated chloride channels that dif-fered significantly in desensitization behavior, thus defining the cytoplasmic loop as an important deter-minant of channel inactivation kinetics.