Molecular and functional interaction between protocadherin-γC5 and GABAA receptors.
We have found that the γ2 subunit of the GABA(A) receptor (γ2-GABA(A)R) specifically interacts with protocadherin-γC5 (Pcdh-γC5) in the rat brain. The interaction occurs between the large intracellular loop of the γ2-GABA(A)R and the cytoplasmic domain of Pcdh-γC5. In brain extracts, Pcdh-γC5 coimmunoprecipitates with GABA(A)Rs. In cotransfected HEK293 cells, Pcdh-γC5 promotes the transfer of γ2-GABA(A)R to the cell surface. We have previously shown that, in cultured hippocampal neurons, endogenous Pcdh-γC5 forms clusters, some of which associate with GABAergic synapses. Overexpression of Pcdh-γC5 in hippocampal neurons increases the density of γ2-GABA(A)R clusters but has no significant effect on the number of GABAergic contacts that these neurons receive, indicating that Pcdh-γC5 is not synaptogenic. Deletion of the cytoplasmic domain of Pcdh-γC5 enhanced its surface expression but decreased the association with both γ2-GABA(A)R clusters and presynaptic GABAergic contacts. Cultured hippocampal neurons from the Pcdh-γ triple C-type isoform knock-out (TCKO) mouse (Pcdhg(tcko/tcko)) showed plenty of GABAergic synaptic contacts, although their density was reduced compared with sister cultures from wild-type and heterozygous mice. Knocking down Pcdh-γC5 expression with shRNA decreased γ2-GABA(A)R cluster density and GABAergic innervation. The results indicate that, although Pcdh-γC5 is not essential for GABAergic synapse formation or GABA(A)R clustering, (1) Pcdh-γC5 regulates the surface expression of GABA(A)Rs via cis-cytoplasmic interaction with γ2-GABA(A)R, and (2) Pcdh-γC5 plays a role in the stabilization and maintenance of some GABAergic synapses.
Department of Physiology and Neurobiology, University of Connecticut, Storrs, Connecticut 06269, USA.
Yanfang
Y
Li
Haiyan
H
Xiao
Tzu-Ting
TT
Chiou
Hongbing
H
Jin
Bevan
B
Bonhomme
Celia P
CP
Miralles
Noelia
N
Pinal
Rashid
R
Ali
Weisheng V
WV
Chen
Tom
T
Maniatis
Angel L
AL
De Blas
D000818:Animals; D019921:Biotinylation; D000070585:Cadherin Related Proteins; D015820:Cadherins / Q000235:genetics / Q000378:metabolism*; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D000076122:Disks Large Homolog 4 Protein; D004622:Embryo, Mammalian; D005260:Female; D005786:Gene Expression Regulation / Q000235:genetics; D005968:Glutamate Decarboxylase / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics; D051528:Guanylate Kinases / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D047468:Immunoprecipitation; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011963:Receptors, GABA-A / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D050495:Vesicular Inhibitory Amino Acid Transport Proteins / Q000378:metabolism
D000070585:Cadherin Related Proteins; D015820:Cadherins; D000076122:Disks Large Homolog 4 Protein; C501447:Dlg4 protein, mouse; C464785:Gamma-protocadherins; D008565:Membrane Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D011963:Receptors, GABA-A; D050495:Vesicular Inhibitory Amino Acid Transport Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D051528:Guanylate Kinases; D005968:Glutamate Decarboxylase