Molecular and functional characterization of recombinant human metabotropic glutamate receptor subtype 5.
We have isolated and characterized overlapping cDNAs that encode two isoforms of the human metabotropic glutamate receptor subtype 5 (hmGluR5). The deduced amino acid sequences of human and rat mGluR5a are 94.5% identical. However, a region in the putative cytoplasmic domain (SER926-ALA1121) displays significant sequence divergence. Genomic analysis of this region showed that the sequence divergence results from species-specific differences in the genomic sequences, not from alternative splicing. The distribution of mGluR5 mRNA in human brain was most strongly detected throughout the hippocampus, with moderate levels in the caudate-putamen, cerebral cortex, thalamus, and deep cerebellar nuclei, and at low levels in the cerebellar cortex. Activation of both hmGluR5a and hmGluR5b transiently expressed in Xenopus oocytes and HEK293 cells was coupled to inositol phosphate (InsP) formation and elevation of the intracellular free calcium ([Ca2+]i). The agonist rank order of potency for activating recombinant hmGluR5a receptors in either system was quisqualate > L-glutamate > 1S,3R-ACPD. Both the quisqualate stimulated InsP and [Ca2+]i were inhibited by (+)-MCPG. Recombinant human mGluR5a was also stably expressed in mouse fibroblast Ltk- cells, in which the efficacy and potency of quisqualate were unchanged for more than 30 cell passages.
SIBIA, Inc., La Jolla, CA 92037-4641, USA.
L P
LP
Daggett
A I
AI
Sacaan
M
M
Akong
S P
SP
Rao
S D
SD
Hess
C
C
Liaw
A
A
Urrutia
C
C
Jachec
S B
SB
Ellis
J
J
Dreessen
D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism; D018076:DNA, Complementary / Q000096:biosynthesis; D004594:Electrophysiology; D005347:Fibroblasts; D018698:Glutamic Acid / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D017403:In Situ Hybridization; D007293:Inosine Triphosphate / Q000096:biosynthesis; D051379:Mice; D008969:Molecular Sequence Data; D009865:Oocytes / Q000378:metabolism; D011233:Precipitin Tests; D051381:Rats; D018094:Receptors, Metabotropic Glutamate / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D014982:Xenopus laevis
D018076:DNA, Complementary; D018094:Receptors, Metabotropic Glutamate; D011994:Recombinant Proteins; D007293:Inosine Triphosphate; D018698:Glutamic Acid; D002118:Calcium