RGS8 Protein Is Distributed in Dendrites and Cell Body of Cerebellar Purkinje Cell

Itoh, Masayuki; Odagiri, Megumi; Abe, Hideki; Saitoh, Osamu

doi:10.1006/bbrc.2001.5489

Cited by 18 publications

(16 citation statements)

References 24 publications

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“…As the sequence of the Nterminal end differs between RGS8 and RGS8S, we compared the subcellular distribution and regulation of distribution patterns in transfected DDT1MF2 cells immunocytochemically. RGS8 protein was localized most abundantly in the nucleus in most of the transfected cells, and cotransfection of RGS8 and constitutively active G␣o (G␣oQL) cDNA resulted in the translocation of RGS8 protein to the plasma membrane as described (6,15). RGS8S protein showed similar nuclear localization and translocation to the plasma membrane induced by G␣oQL (Fig.…”

Section: Cellular Distribution Of Rgs8 and Rgs8smentioning

confidence: 76%

“…Immunofluorescence staining was performed with anti-RGS8 antibody, which was raised against a peptide corresponding to residues 27-42 of rat RGS8. Details of antibody specificity and microscopy methods have been described (15). Briefly, cells were cultured on glass coverslips and fixed with 4% paraformaldehyde in PBS for 30 min at room temperature.…”

Section: Materials and Methods Reverse Transcriptase-pcr (Rt-pcr)mentioning

confidence: 99%

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Alternative splicing of RGS8 gene determines inhibitory function of receptor type-specific Gq signaling

Saitoh

Murata

Odagiri

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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The regulators of G protein signaling (RGS) proteins modulate heterotrimeric G protein signaling. RGS8 is a brain-specific RGS protein of 180 aa. Here we identified a short isoform of RGS8, RGS8S, that arises by alternative splicing. RGS8S cDNA encodes a N terminus of 7 aa instead of amino acids 1-9 of RGS8 and 10 -180 of RGS8. The subcellular distribution of RGS8 and RGS8S did not differ significantly in transfected cells. RGS8S accelerated, not as efficiently as RGS8, the turning on and off of Gi͞o-mediated modulation of G protein-gated inwardly rectifying K ؉ channels in Xenopus oocytes. We next examined the effects of RGS8 and RGS8S on Gq-mediated signaling. RGS8 decreased the amplitude of the response upon activation of m1 muscarinic or substance P receptors, but did not remarkably inhibit signaling from m3 muscarinic receptors. In contrast, RGS8S showed much less inhibition of the response of either of these Gq-coupled receptors. By quantitative analysis of the inhibitory effect and the protein expression level, we confirmed that the difference of inhibitory effect is caused by both the qualitative difference between RGS8 and RGS8S and the quantitative difference of the protein expression level. We also confirmed that the receptor-type specificity of inhibition is not caused by the difference of the expression level of the receptors. In summary, we showed that 9 aa in the N terminus of RGS8 contribute to the function to inhibit Gq-coupled signaling in a receptor type-specific manner and that the regulatory function of RGS8S is especially diminished on Gq-coupled responses. R egulators of G protein signaling (RGS) proteins comprise a large family of more than 20 members, which modulate heterotrimeric G protein signaling. They share a homologous domain, the RGS domain, which is flanked by diverse N and C termini (1-3). The RGS domain alone is sufficient for activating the GTPase of G␣, whereas the flanking domains confer various regulatory properties (3). RGS8 was identified in rat brain and is a small RGS protein along with RGS4, RGS5, and RGS16 (4, 5). We recently showed that RGS8 protein was concentrated in nuclei of cells transfected with cDNA for RGS8 expression and that coexpression of a constitutively active G␣o resulted in the translocation of RGS8 protein to the plasma membrane. The deletion of the N-terminal region (35 aa) of RGS8 abolished its nuclear localization and active G␣o-induced redistribution. This truncated mutant of RGS8, however, is still functional in inhibiting pheromone signaling in yeast to some extent. When coexpressed with G protein-gated inwardly rectifying K ϩ (GIRK) channels, the truncated RGS8 accelerated both turning on and off similar to RGS8. Acute desensitization of GIRK current observed in the presence of RGS8, however, was not induced. Thus, we clarified that RGS8 requires its N terminus for subcellular localization and full regulatory function (6). On the other hand, Zeng et al. (7) reported that the N-terminal domain (1-33 aa) of RGS4 confers receptor-selective inhibi...

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Section: Cellular Distribution Of Rgs8 and Rgs8smentioning

confidence: 76%

Section: Materials and Methods Reverse Transcriptase-pcr (Rt-pcr)mentioning

confidence: 99%

Alternative splicing of RGS8 gene determines inhibitory function of receptor type-specific Gq signaling

Saitoh

Murata

Odagiri

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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“…Protein eluates were subjected to SDS‐PAGE and electrotransferred to a nitrocellulose membrane. The membranes were blocked with 5% skim milk in Tris‐buffered saline containing 0.1% Tween 20 (T‐TBS) for 2 h. Then, the membranes were probed for 90 min with anti‐RGS8 antibody, which was previously described (Itoh et al . 2001), followed by washing five times with T‐TBS.…”

Section: Methodsmentioning

confidence: 99%

“…Both RGS8 and RGS8S are specifically expressed in the brain, especially in cerebellar Purkinje cells (Saitoh et al . 1997; Itoh et al . 2001).…”

mentioning

confidence: 99%

Alternative splicing of RGS8 gene changes the binding property to the M1 muscarinic receptor to confer receptor type‐specific Gq regulation

Itoh¹,

Nagatomo²,

Kubo³

et al. 2006

Journal of Neurochemistry

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RGS proteins constitute a large family that modulates heterotrimeric G-protein signaling. We previously showed that RGS8 suppressed Gq signaling in a receptor type-specific manner. To elucidate molecular mechanisms underlying receptor-specific attenuation by RGS8, we examined whether RGS8 can interact with certain G-protein-coupled receptors. By pull-down assay, we showed that RGS8 directly binds to the third intracellular (i3) loop of M1 and M3 muscarinic acetylcholine receptors (mAChRs). The binding of RGS8S, a splice variant with a different N-terminus, was weaker. RGS8 could bind specifically to the C-terminal part of M1i3 (containing amino acids of 304-353 of i3 of human M1-mAChR), but RGS8S could not. Moreover, deletion of the N-terminal 9 amino acids and substitution of both Arg-8 and Arg-9 of RGS8 with Ala resulted in reduced binding to M1i3. BRET experiments revealed that RGS8 actually interacts with M1-mAChR, but RGS8S does not interact in the living cells. The RGS8 mutant, which had less binding ability to M1i3, showed a reduced inhibitory function of Gq signaling through M1-mAChR. These results demonstrated that RGS8 can directly interact with M1-mAChR via its N-terminus and the i3 loop of the receptor, and this binding must play an essential role in receptor-specific suppression by RGS8.

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“…Here, RGS8 is expressed exclusively in the soma and dendrites of Purkinje cells (Itoh et al, 2001;, where it increases both the on and off rate of Gbg-mediated GIRK channel current following GPCR activation (Saitoh et al, 1997). Interestingly, rapid activation of the GIRK channel appears to be mediated by the N-terminal region of RGS8 as opposed to the RGS domain (Jeong and Ikeda, 2001), demonstrating a function for RGS8 independent of GAP activity.…”

Section: Other Rgs Proteinsmentioning

confidence: 99%

Roles for Regulator of G Protein Signaling Proteins in Synaptic Signaling and Plasticity

2015

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The regulator of G protein signaling (RGS) family of proteins serves critical roles in G protein-coupled receptor (GPCR) and heterotrimeric G protein signal transduction. RGS proteins are best understood as negative regulators of GPCR/G protein signaling. They achieve this by acting as GTPase activating proteins (GAPs) for Ga subunits and accelerating the turnoff of G protein signaling. Many RGS proteins also bind additional signaling partners that either regulate their functions or enable them to regulate other important signaling events. At neuronal synapses, GPCRs, G proteins, and RGS proteins work in coordination to regulate key aspects of neurotransmitter release, synaptic transmission, and synaptic plasticity, which are necessary for central nervous system physiology and behavior. Accumulating evidence has revealed key roles for specific RGS proteins in multiple signaling pathways at neuronal synapses, regulating both pre-and postsynaptic signaling events and synaptic plasticity. Here, we review and highlight the current knowledge of specific RGS proteins (RGS2, RGS4, RGS7, RGS9-2, and RGS14) that have been clearly demonstrated to serve critical roles in modulating synaptic signaling and plasticity throughout the brain, and we consider their potential as future therapeutic targets.

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RGS8 Protein Is Distributed in Dendrites and Cell Body of Cerebellar Purkinje Cell

Cited by 18 publications

References 24 publications

Alternative splicing of RGS8 gene determines inhibitory function of receptor type-specific Gq signaling

Alternative splicing of RGS8 gene determines inhibitory function of receptor type-specific Gq signaling

Alternative splicing of RGS8 gene changes the binding property to the M1 muscarinic receptor to confer receptor type‐specific Gq regulation

Roles for Regulator of G Protein Signaling Proteins in Synaptic Signaling and Plasticity

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