Marlin-1, a Novel RNA-binding Protein Associates with GABA Receptors

Abstract

GABAB receptors are heterodimeric G protein-coupled receptors that mediate slow synaptic inhibition in the central nervous system. Whereas heterodimerization between GABAB receptor GABABR1 and GABABR2 subunits is essential for functional expression, how neurons coordinate the assembly of these critical receptors remains to be established. Here we have identified Marlin-1, a novel GABAB receptor-binding protein that associates specifically with the GABABR1 subunit in yeast, tissue culture cells, and neurons. Marlin-1 is expressed in the brain and exhibits a granular distribution in cultured hippocampal neurons. Marlin-1 binds different RNA species including the 3′-untranslated regions of both the GABABR1 and GABABR2 mRNAs in vitro and also associates with RNA in cultured neurons. Inhibition of Marlin-1 expression via small RNA interference technology results in enhanced intracellular levels of the GABABR2 receptor subunit without affecting the level of GABABR1. Together our results suggest that Marlin-1 functions to regulate the cellular levels of GABAB R2 subunits, which may have significant effects on the production of functional GABAB receptor heterodimers. Therefore, our observations provide an added level of regulation for the control of GABAB receptor expression and for the efficacy of inhibitory synaptic transmission. GABAB receptors are heterodimeric G protein-coupled receptors that mediate slow synaptic inhibition in the central nervous system. Whereas heterodimerization between GABAB receptor GABABR1 and GABABR2 subunits is essential for functional expression, how neurons coordinate the assembly of these critical receptors remains to be established. Here we have identified Marlin-1, a novel GABAB receptor-binding protein that associates specifically with the GABABR1 subunit in yeast, tissue culture cells, and neurons. Marlin-1 is expressed in the brain and exhibits a granular distribution in cultured hippocampal neurons. Marlin-1 binds different RNA species including the 3′-untranslated regions of both the GABABR1 and GABABR2 mRNAs in vitro and also associates with RNA in cultured neurons. Inhibition of Marlin-1 expression via small RNA interference technology results in enhanced intracellular levels of the GABABR2 receptor subunit without affecting the level of GABABR1. Together our results suggest that Marlin-1 functions to regulate the cellular levels of GABAB R2 subunits, which may have significant effects on the production of functional GABAB receptor heterodimers. Therefore, our observations provide an added level of regulation for the control of GABAB receptor expression and for the efficacy of inhibitory synaptic transmission. It has become widely accepted that protein-protein interactions are responsible for the formation and maintenance of the signaling platforms that organize local transduction units (1Alberts B. Cell. 1998; 92: 291-294Google Scholar). Protein associations have also been shown to modulate the synthesis, targeting, and stabilization of membrane receptors (2Grant S.G. Husi H. Trends Biotechnol. 2001; 19: S49-S54Abstract Full Text Full Text PDF Google Scholar). The study of the spatial and temporal compartmentalization of G protein-coupled receptors (GPCRs) 1The abbreviations used are: GPCR, G protein-coupled receptor; CCR, coiled-coil region; GST, glutathione S-transferase; UTR, untranslated region; FITC, fluorescein isothiocyanate; PBS, phosphate-buffered saline; contig, group of overlapping clones; EGFP, enhanced green fluorescent protein; SCG, superior cervical ganglion; sRNAi, small RNA interference. is essential to understand the mechanisms for neuronal integration of multiple stimuli. The identification of signaling partners has been well documented for ion channels (3Husi H. Ward M.A. Choudhary J.S. Blackstock W.P. Grant S.G. Nat. Neurosci. 2000; 3: 661-669Google Scholar, 4Craig A.M. Boudin H. Nat. Neurosci. 2001; 4: 569-578Google Scholar, 5Moss S.J. Smart T.G. Nat. Rev. Neurosci. 2001; 4: 240-250Google Scholar), but less is known about the proteins that assist GPCRs (6Davare M.A. Avdonin V. Hall D.D. Peden E.M. Burette A. Weinberg R.J. Horne M.C. Hoshi T. Hell J.W. Science. 2001; 293: 98-101Google Scholar, 7El Far O. Betz H. Biochem. J. 2002; 365: 329-336Google Scholar). GABAB receptors mediate the slow and prolonged phase of synaptic inhibition (8Mott D.D. Lewis D.V. Int. Rev. Neurobiol. 1994; 36: 97-223Google Scholar) and, unlike other GPCRs, they require the formation of a heterodimer between GABABR1 and GABABR2 (9Bouvier M. Nat. Rev. Neurosci. 2001; 4: 274-286Google Scholar). These two subunits display high homology to metabotropic glutamate, Ca2+-sensing, vomeronasal, and putative pheromone receptors, and recombinant GABABR1/GABABR2 receptors mimic the effector-coupling and pharmacological properties of native receptors (10Couve A. Moss S.J. Pangalos M.N. Mol. Cell Neurosci. 2000; 16: 296-312Google Scholar). Clinically, GABAB receptors have been implicated in depression, neuroprotection, and cognition, and the production of GABABR1 knock-out mice has confirmed their role in epilepsy and pain (11Prosser H.M. Gill C.H. Hirst W.D. Grau E. Robbins M. Calver A. Soffin E.M. Farmer C.E. Lanneau C. Gray J. Schneck E. Warmerdam B.S. Clapham C. Reavill C. Rogers D.C. Stean T. Upton N. Humphreys K. Randall A. Geppert M. Davies C.H. Pangalos M.N. Mol. Cell Neurosci. 2001; 17: 1059-1070Google Scholar, 12Schuler V. Lüscher C. Blanchet C. Klix N. Sansig G. Klebs K. Schmutz M. Heid J. Gentry C. Urban L. Fox A. Spooren W. Jaton A. Vigouret J.M. Pozza M. Kelly P.H. Mosbacher J. Froestl W. Käslin E. Korn R. Bischoff S. Kaupmann K. van der Putten H. Bettler B. Neuron. 2001; 31: 47-58Google Scholar). Furthermore, recent studies indicate that their activation may be beneficial in the treatment of withdrawal symptoms from addictive drugs (13Roberts D.C. Andrews M.M. Psychopharmacology (Berl.). 1997; 131: 271-277Google Scholar). Given their critical role in synaptic transmission and their potential therapeutic implications, it is fundamental to understand all aspects of GABAB receptor signaling. The modulation of neuronal GABAB receptors is likely to occur at multiple levels ranging from transcriptional/translational control and RNA metabolism to receptor targeting, post-translational modifications, and membrane stabilization. The roles of heterodimer assembly and GABABR2 phosphorylation have been documented previously (14Couve A. Filippov A.K. Connolly C.N. Bettler B. Brown D.A. Moss S.J. J. Biol. Chem. 1998; 273: 26361-26367Google Scholar, 15Jones K.A. Borowski B. Tamm J.A. Craig D.A. Durkin M.M. Dai M. Yao W. Johnson M. Gunwaldsen C. Huang L. Tang C. Shen Q. Salon J.A. Morse K. Laz T. Smith K.E. Nagarathnam D. Noble S.A. Branchek T.A. Gerald C. Nature. 1998; 396: 674-679Google Scholar, 16Kaupmann K. Malitschek B. Schuler V. Heid J. Froestl W. Beck P. Mosbacher J. Bischoff S. Kulik A. Shigemoto R. Karschin A. Bettler B. Nature. 1998; 396: 683-687Google Scholar, 17Kuner R. Kohr G. Grunewald S. Eisenhartdt G. Bach A. Kornau H.C. Science. 1999; 283: 74-77Google Scholar, 18White J.H. Wise A. Main M.J. Green A. Fraser N.J. Disney G.H. Barnes A.A. Emson P. Foord S.M. Marshall F. Nature. 1998; 396: 679-682Google Scholar, 19Couve A. Thomas P. Calver A.R. Hirst W.D. Pangalos M.N. Walsh F.S. Moss S.J. Nat. Neurosci. 2002; 5: 415-424Google Scholar). In addition, the identification of 14-3-3, ATF4/CREB2, and Mupp1 as GABAB-associated proteins has started to reveal the macromolecular complexity of GABAB receptor signaling (20Couve A. Kittler J.T. Uren J.M. Calver A.R. Pangalos M.N. Moss S.J. Mol. Cell Neurosci. 2001; 17: 317-328Google Scholar, 21Nehring R.B. Horikawa H.P. El Far O. Kneussel M. Brandstatter J.H. Stamm S. Wischmeyer E. Betz H. Karschin A. J. Biol. Chem. 2000; 275: 35185-35191Google Scholar, 22Vernon E. Meyer G. Pickard L. Dev K. Molnar E. Collingridge G.L. Henley J.M. Mol. Cell Neurosci. 2001; 17: 637-645Google Scholar, 23White J.H. McIllhinney R.A. Wise A. Ciruela F. Chan W.Y. Emson P.C. Billinton A. Marshall F.H. Proc. Natl. Acad. Sci. U. S. A. 2000; 97: 13967-13972Google Scholar, 24Milligan G. White J.H. Trends Pharmacol. Sci. 2001; 22: 513-518Google Scholar). Recently, the local metabolism of mRNAs has been identified as a key element in the organization of synaptic sites (25Palacios I.M. St. Johnston D. Annu. Rev. Cell Dev. Biol. 2001; 17: 569-614Google Scholar). However, there is no evidence yet for the role of RNA metabolism in the regulation of GABAB receptor targeting and stability. Here we report the identification of Marlin-1, a novel brain-specific GABAB-binding protein that associates specifically with the GABABR1 subunit. We demonstrate that Marlin-1 also associates with RNA and that its interaction with the receptor and RNA occurs selectively through discrete coiled-coil regions. Moreover, we show that a decrease in Marlin-1 expression results in an abnormal accumulation of the GABABR2 receptor protein consistent with a role in RNA metabolism. The regulation of receptor biology through RNA metabolism widens significantly the modulatory levels of GABAB receptor function. Materials—Syto14 was purchased from Molecular Probes, Inc. (Eugene, OR). RNA polymers (poly(I)-poly(C) and poly(C)) were from was purchased from The the of GABABR1 in the and the and expression have been previously (14Couve A. Filippov A.K. Connolly C.N. Bettler B. Brown D.A. Moss S.J. J. Biol. Chem. 1998; 273: 26361-26367Google Scholar, 19Couve A. Thomas P. Calver A.R. Hirst W.D. Pangalos M.N. Walsh F.S. Moss S.J. Nat. Neurosci. 2002; 5: 415-424Google Scholar, A. Kittler J.T. Uren J.M. Calver A.R. Pangalos M.N. Moss S.J. Mol. Cell Neurosci. 2001; 17: 317-328Google Scholar). The of in the GABAB have also been (20Couve A. Kittler J.T. Uren J.M. Calver A.R. Pangalos M.N. Moss S.J. Mol. Cell Neurosci. 2001; 17: 317-328Google Scholar). Marlin-1 was the and the was at The of the of the and receptors were from and J. and to the glutathione protein and The of different of Marlin-1 were and in in the to and and of were were as previously (20Couve A. Kittler J.T. Uren J.M. Calver A.R. Pangalos M.N. Moss S.J. Mol. Cell Neurosci. 2001; 17: 317-328Google Scholar). The the to the of GABABR1 was the in the was used to for interactions were on and and In addition, the were interactions were confirmed the and were from and a The was purchased from Inc. and were purchased from The proteins and were used as to and to and were as of the The and to fluorescein and a were purchased from The to was purchased from membrane was with the from the The membrane was and with a control Cell and were and as previously (14Couve A. Filippov A.K. Connolly C.N. Bettler B. Brown D.A. Moss S.J. J. Biol. Chem. 1998; 273: 26361-26367Google Scholar). and and and hippocampal neurons were and as previously (14Couve A. Filippov A.K. Connolly C.N. Bettler B. Brown D.A. Moss S.J. J. Biol. Chem. 1998; 273: 26361-26367Google Scholar, A.K. A. Pangalos M.N. Walsh F.S. Brown D.A. Moss S.J. J. Neurosci. 2000; Scholar, J.T. P. Brown D.A. Smart T.G. Moss S.J. J. Neurosci. 2000; Scholar). neurons were and hippocampal neurons were from and and hippocampal neurons were the of the and for in from were in of in a The was at for and the was The was in the and at for The were and at for to a membrane and a The membrane was in and at a of The was at for The was and the was in an of of brain cells, and cultured neurons were as previously A. Thomas P. Calver A.R. Hirst W.D. Pangalos M.N. Walsh F.S. Moss S.J. Nat. Neurosci. 2002; 5: 415-424Google Scholar). were an was the were in a and were of were with and with in as J. Davies C.H. Pangalos M.N. Mol. Cell Neurosci. Scholar). were the were and the tissue was in was at were a tissue were with in as previously J. Davies C.H. Pangalos M.N. Mol. Cell Neurosci. Scholar). was with with the from were were were with PBS, for in and for in in were in for at and for in were with and for at in were with in culture for at and in to were a and and were as previously A. Thomas P. Calver A.R. Hirst W.D. Pangalos M.N. Walsh F.S. Moss S.J. Nat. Neurosci. 2002; 5: 415-424Google Scholar, A. Kittler J.T. Uren J.M. Calver A.R. Pangalos M.N. Moss S.J. Mol. Cell Neurosci. 2001; 17: 317-328Google Scholar). proteins were with the in vitro were with of proteins for at in were in of for and were and to a The were in a and of was the RNA of proteins were in were in RNA for at and in RNA for at The RNA was and the The to the regions of GABABR1 and GABABR2 were and the RNA was the in vitro were in RNA and to a The were in a and of and sites were in vitro The were to to a different of the Marlin-1 The regions to the of an the for and the which in its The for was also The were a the RNA S. and P. from these a which to the Marlin-1 through the and its of were hippocampal neurons with of to and of Marlin-1, a novel partners of the GABAB the intracellular of the GABABR1 subunit was used to a in a of were identified on the of the a of group was the different the and we overlapping of the that was to a group of putative proteins in The assembly of the from was to the of the and the of Therefore, the of the a the was from a brain The of with a of of the on the that and novel protein has been Marlin-1 and that Marlin-1 coiled-coil regions two of which the in the overlapping of Marlin-1 and the distribution of the Marlin-1 in different a was and Marlin-1 species was expressed in the brain In addition, levels of species were in The of these was the protein distribution of Marlin-1, two proteins the and of Marlin-1 were used as to and from brain the of a of the of the the Marlin-1 was a expression and at the with the was cells, and were for of was with in and was from from and The was with Marlin-1 and The of the recombinant species be the of the the Marlin-1 The of the in the brain has been The distribution of the Marlin-1 protein was in from levels of the were in the brain These results indicate that Marlin-1 and protein are expressed in the central nervous consistent with a role in GABAB receptor of GABAB and interaction of GABAB receptors and Marlin-1 was a of the interaction was through a Marlin-1 expressed as a of were and to a The membrane was with the of GABABR1 with interaction was between GABABR1 and Marlin-1 but between GABABR1 and control the was was in vitro and in the of control the of the GABABR1 GABABR2 metabotropic receptor and receptors Marlin-1 with GABABR1 but with the other receptor used as These results the and indicate that the interaction between the GABABR1 receptor and Marlin-1 is and The between GABAB receptors and Marlin-1 was of recombinant proteins in tissue culture were with both of that and were expressed and were with a and with with and was from control These results demonstrate that GABABR1 and Marlin-1 in a cellular the was used to the of interaction of GABABR1 on regions of Marlin-1 were a expression and in vitro a in the of The proteins were in the of control The were and the were and in a The results of multiple indicate that Marlin-1 the and the coiled-coil but the coiled-coil the of Marlin-1 with the GABABR1 subunit. These the observations and demonstrate that the different coiled-coil of Marlin-1 display a high of of Marlin-1 and GABAB cellular distribution of GABAB receptors and Marlin-1 was in from and significant in the expression of the two proteins was in different brain regions. a of GABABR1 and Marlin-1 was in the of a of neurons in the and the their spatial the distribution of GABAB receptors and Marlin-1 was in were with and and was with and an the for subunit (14Couve A. Filippov A.K. Connolly C.N. Bettler B. Brown D.A. Moss S.J. J. Biol. Chem. 1998; 273: 26361-26367Google Scholar) The distribution of was of Marlin-1, and both proteins in a a the distribution of display significant with Marlin-1 the distribution of Marlin-1 was in hippocampal neurons in The with both Marlin-1 was and levels of Marlin-1 in the and of hippocampal neurons an and granular of expression that was in the central of the and in the of neuronal The shown in and was in no significant the of the of hippocampal neurons in a granular and in the of that the and neuronal and The of GABAB receptors and Marlin-1 was in cultured hippocampal neurons. The of distribution of Marlin-1 with both receptor the in the was with GABABR2 with GABABR1 and These results indicate that a of GABAB receptors and Marlin-1 in Moreover, the with GABABR2 that the of may to that of receptor In to the distribution of Marlin-1 and GABAB receptors in a neuronal and receptors were in hippocampal neurons via of neurons that the of Marlin-1 in that the and through the and were a significant of GABAB receptors in these Marlin-1 receptors the were to these In addition, the that the distribution of the was the expression of GABAB in the of GABAB receptors that the Marlin-1 were less and These results indicate that GABAB receptors and Marlin-1 in a neuronal and suggest that Marlin-1 associations that are GABABR1 in hippocampal neurons. Inhibition of Marlin-1 and of GABAB on provide on the role of Therefore, an on the inhibition of expression of Marlin-1 via technology was in to its function. different were a the and for their to Marlin-1 expression in were with Marlin-1, EGFP, and control The control and two of the no in the of Marlin-1 In of the the expression of Marlin-1 with effects on These observations indicate that a to the inhibition of Marlin-1 expression in a neuronal the was of and the of Marlin-1 was Marlin-1 expression significantly in neurons with and Marlin-1 in These results demonstrate that Marlin-1 expression levels are and specifically through in cultured neurons. In of these of in vitro hippocampal neurons that the levels of Marlin-1 were in control proteins study the of Marlin-1 in GABAB receptor neurons were with and the levels of and were in neurons with levels of and an in the of but of was in neurons of the levels of GABABR2 a significant in the subunit levels in neurons GABABR2 in In the levels of the control protein that Marlin-1 expression is likely as a of these results suggest that the of Marlin-1 a in the levels of GABABR2 These observations suggest that in the Marlin-1 protein levels with the of GABAB receptors the of neurons. Marlin-1 with RNA in and of the Marlin-1 previously (25Palacios I.M. St. Johnston D. Annu. Rev. Cell Dev. Biol. 2001; 17: 569-614Google Scholar, A.M. Neuron. 2001; Scholar, S.J. D. L. N. E. Neuron. 2001; Scholar). We that the of Marlin-1 to with RNA the of GABABR2 expression at a RNA metabolism the of Marlin-1 to RNA was with RNA RNA was with and used as a to study the of RNA to the control protein GST, to the protein Marlin-1, to the proteins and the and coiled-coil regions of Marlin-1, with its granular Marlin-1, but the control with the RNA was to mediate the the GABAB was at RNA It is well known that in the targeting of mRNAs in the of the Johnston D. Cell. Scholar). Therefore, in to with Marlin-1, the regions of the GABABR1 and GABABR2 were in the of and used as in the RNA both In addition, was to mediate the interaction and The GABAB were less at Marlin-1 the but all were at The is and remains to be in It is that proteins as a known protein that binds in have a RNA in vitro L. T. M. L. Mol. Cell Biol. 1999; Scholar, F. Trends Cell Biol. 2000; Scholar). However, as a to the of Marlin-1 RNA the RNA was in the of RNA RNA was at the the RNA the RNA remains these results suggest that Marlin-1 is likely to a Marlin-1 with RNA in a the in vitro RNA results the distribution of Marlin-1 was brain and with the distribution of proteins in that Marlin-1 in two in the of Marlin-1 was in a significant levels were also in a membrane with the GABAB receptor and of the membrane with in the of Marlin-1 from the membrane that it to a protein of the that Marlin-1 was a protein but that it a the and protein A.M. Neuron. 2001; Scholar) but the and These results suggest that a of Marlin-1 may with to the of Marlin-1 with RNA in a cellular neurons hippocampal neurons were with Marlin-1 and for with and a that selectively associates with RNA and R.B. J.H. J. Neurosci. 16: Scholar, M.A. P. M. P. J. J. Neurosci. 1999; 19: Scholar). Marlin-1 the and, these of RNA results were with hippocampal neurons The of the granular was with for other proteins known to with the Marlin-1 with for the and and these results indicate that Marlin-1 associates with RNA in vitro and in cultured neurons and suggest that the effects on GABABR2 may be through the of RNA metabolism RNA The results provide evidence to suggest that the GABAB receptor is a group of The observations in study indicate that Marlin-1 is a novel brain-specific which is of with the GABAB receptor and with different species of RNA including both GABABR1 and Marlin-1 coiled-coil and, in with a recent report coiled-coil Science. Scholar). The coiled-coil in the in the of receptor RNA but the associates with the membrane in tissue culture and neurons. and S. J. The central coiled-coil associates with but it is at GABAB The RNA has been but the coiled-coil a a of of proteins and L. J. Biol. Chem. 1998; 273: Scholar, K. K. R. M. 1999; Scholar). We have been to significant between the RNA of Marlin-1 and other RNA to a that the of Marlin-1 but with of the a group of proteins in regulation M. van R. M.A. 2000; Scholar). The coiled-coil has the and associates with GABABR1 but with The of these associations are they are in with a in which Marlin-1 functions as an between a membrane receptor and RNA The of Marlin-1 in the neuronal and a role in of the control of expression as targeting of RNA for receptor other aspects of metabolism of in the of inhibitory have of Marlin-1 in that Marlin-1 in the in aspects of neuronal that are to synaptic transmission. have also that Marlin-1 is of protein in the and and, that they a small of proteins yet the proteins of their in the GABAB that their to GABAB receptors may be these their may be their These aspects of Marlin-1 an of The in the levels of GABABR2 in the to the that the of Marlin-1 with the receptor heterodimer occurs via the GABABR1 subunit. However, it is that GABABR1 the targeting, protein of GABABR2 through proteins In the of Marlin-1, the of GABABR2 may a the of local control may to The of the GABABR2 protein but of the GABABR2 in the GABABR1 knock-out mice that the protein levels of GABABR2 and heterodimer assembly are the control of the GABABR1 subunit (11Prosser H.M. Gill C.H. Hirst W.D. Grau E. Robbins M. Calver A. Soffin E.M. Farmer C.E. Lanneau C. Gray J. Schneck E. Warmerdam B.S. Clapham C. Reavill C. Rogers D.C. Stean T. Upton N. Humphreys K. Randall A. Geppert M. Davies C.H. Pangalos M.N. Mol. Cell Neurosci. 2001; 17: 1059-1070Google Scholar, 12Schuler V. Lüscher C. Blanchet C. Klix N. Sansig G. Klebs K. Schmutz M. Heid J. Gentry C. Urban L. Fox A. Spooren W. Jaton A. Vigouret J.M. Pozza M. Kelly P.H. Mosbacher J. Froestl W. Käslin E. Korn R. Bischoff S. Kaupmann K. van der Putten H. Bettler B. Neuron. 2001; 31: 47-58Google Scholar). These observations are consistent with our It is well that and proteins as control the expression levels of a of neuronal receptors as and receptors P.H. J. Scholar, W. M. D. J. Biol. Chem. Scholar, G. D.D. J.W. J. Biol. Chem. 1998; 273: Scholar, P.C. 2002; Scholar, C.N. Mol. Scholar, S. H. A. A. J. Neurosci. Scholar). consistent with our of regulation is to be the of the protein levels for from we have been to Marlin-1 in the Marlin-1 is to between the and the remains a the evidence that Marlin-1 the of GABAB receptors via the control of RNA metabolism. The distribution of the GABABR1 and GABABR1 mRNAs has been in cultured neurons. However, the of other RNA species from the to the has been are a of of mRNAs in to and these the the the and (25Palacios I.M. St. Johnston D. Annu. Rev. Cell Dev. Biol. 2001; 17: 569-614Google Scholar, F. Trends Cell Biol. 2000; Scholar, O. Proc. Natl. Acad. Sci. U. S. A. 2001; Scholar, J. Cell Biol. 1999; Scholar, O. E.M. Annu. Rev. Neurosci. 2001; Scholar). The of RNA is to F. Trends Cell Biol. 2000; Scholar, R.B. J.H. J. Neurosci. 16: Scholar) and has been to in protein in the of synaptic it may modulate synaptic A.M. Neuron. 2001; Scholar). protein and have been identified as of the they multiple species and are in the and neuronal M.A. P. M. P. J. J. Neurosci. 1999; 19: Scholar, A. T. L. S. J. Neuron. Scholar). studies between proteins in the targeting of and Marlin-1 provide on the of the Marlin-1 and their role in neuronal function. protein and of the as the the to the and have also been in the of mRNAs to proteins are likely to be targeting O. E.M. Annu. Rev. Neurosci. 2001; Scholar, T. Biol. 2001; Scholar, C. J. Nat. Neurosci. Rev. 2001; Scholar). GABAB receptors are to their local protein through proteins remains to be in and of with Marlin-1 from brain and neuronal provide these key aspects of GABAB receptor function.