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J Biol Chem, Vol. 273, Issue 52, 34687-34690, December 25, 1998
§,§,
,,,,, and
From the Departments of Physiology and
¶ Pharmacology, University of Texas Southwestern Medical Center,
Dallas, Texas 75235 and Department of Metabolic Diseases,
Hoffmann-La Roche, Inc., Nutley, New Jersey 07110
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ABSTRACT |
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Top
Abstract
Introduction
Materials & Methods
Results & Discussion
References
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Regulators of heterotrimeric G protein signaling
(RGS) proteins are GTPase-activating proteins (GAPs) that accelerate
GTP hydrolysis by Gq and Gi Heterotrimeric G proteins of the Gq class are
mediators of Ca2+ responses in animal cells. Signaling is
initiated by agonist binding to heptahelical transmembrane receptors
complexed with Gq RGS proteins are GTPase-activating proteins (GAPs) that accelerate GTP
hydrolysis by Gq and Gi In the present study, we used deletion mutations to identify two
domains in RGS4 that regulate agonist-dependent
Ca2+ signaling. The RGS box accelerates GTP hydrolysis by
G Expression and Purification of Recombinant RGS4
Proteins--
All recombinant RGS proteins were
His6-tagged at the N terminus. Protein expression and GAP
assays were performed as described (10).
Peptide Synthesis--
Synthetic
peptides3 were purified by
reverse-phase high pressure liquid chromatography and confirmed by
amino acid analysis and fast atom bombardment/mass spectroscopy.
Measurement of Ca2+ Release--
Measurement of
Ca2+-activated Cl GTPase Measurements--
The agonist-stimulated steady-state
GTPase activity of reconstituted phospholipid vesicles that contained
m1 muscarinic acetylcholine receptors and heterotrimeric Gq
was assayed in the presence or absence of RGS proteins or peptides as
described (14). Single-turnover measurements of the hydrolysis of
G The N Terminus of RGS4 Is Required for High Potency
Inhibition--
To determine the domains of RGS4 that conveyed high
potency and receptor-selective inhibition, rat pancreatic acinar cells were dialyzed with different recombinant RGS4 proteins through a patch
pipette and exposed to 100 µM carbachol, the minimal
concentration of carbachol needed to generate a maximal cellular
response (16, 17).2 Responses were detected as changes in
current carried by a Ca2+-activated Cl
We next tested the effect of the RGS domain of RGS4 (4Box) (Fig. 1,
d-f). Although full-length RGS4 and 4Box accelerated GTP hydrolysis by purified G
To identify the flanking structure in full-length RGS4 that conveys
high potency inhibition, we tested the activities of other deletion
mutants. C-terminal truncation of RGS4 (RGS4
Besides diminishing the effect and potency of RGS4 action, 4Box also
lost selectivity among different receptors. RGS4 inhibits signaling
preferentially through the m3 muscarinic receptor compared with the CCK
receptor, assayed either separately or sequentially within the same
cell (Fig. 2).2 By contrast,
4Box inhibited Ca2+ signaling by carbachol and CCK
equivalently over a wide range of concentrations (Fig. 2,
c-f). These data suggest that terminal regions of RGS4 both
enhance potency and mediate recognition of receptor.
Receptor-selective Inhibition by the RGS4 N-terminal
Domain--
Next, we tested whether the 33-amino acid N-terminal
peptide of RGS4 (P1-33) can itself alter
Gq-mediated signaling. P1-33 (100 nM) both blocked the initial spike and converted the
following sustained current to an oscillatory response (Fig. 2,
g-i). Higher concentrations of P1-33
completely blocked signaling by carbachol. In contrast, these
concentrations of P1-33 had no effect on CCK signaling.
Only at 1 µM P1-33 was the sustained
response evoked by CCK converted to an oscillatory current. These data
indicate both that P1-33 effectively inhibited Gq-mediated Ca2+ signaling and that inhibition
retained the selectivity for the m3 muscarinic receptor over the CCK
receptor that is characteristic of RGS4.
RGS4 Flanking Sequences Enhance G N Terminus and 4Box Act Synergistically to Inhibit Ca2+
Signaling--
The results in Figs. 1-3 clearly show that 4Box has
Gq-GAP activity but that the N-terminal domain of RGS4 both
conferred receptor selectivity upon RGS4 action and increased its
potency in intact cells. To address the mechanism of RGS4 action,
Ca2+ release from intracellular stores was measured in
streptolysin O-permeabilized cells. These cells sequester
Ca2+ from the incubation medium into cellular organelles
and retain Gq-coupled signaling in response to all agonists
that act on acinar cells (3, 20). This experimental approach
facilitated addition of GTP
As shown in Fig. 4a,
muscarinic stimulation released about 75-80% of the
IP3-sensitive Ca2+ pool. Addition of 125 nM RGS4 inhibited more than 95% of the normal
carbachol-stimulated Ca2+ release (Fig. 4f).
Permeabilized cells were less sensitive to RGS4 than patch-clamped
cells, probably because of a slower diffusion of proteins from the
extracellular media into the cytosol. RGS4 inhibition was fully
reversible by addition of GTP
Consistent with its effects in intact cells, P1-33
inhibited Ca2+ release in permeabilized cells in a
receptor-selective manner (Fig. 4, c-e). Carbachol-evoked
Ca2+ release was inhibited completely by 62 µM P1-33, with an IC50 of
25 ± 2 µM (n = 5). Similar
measurements with CCK showed an IC50 for P1-33
of 62 ± 5 µM (n = 3). Thus, in
permeabilized cells, P1-33 inhibited Ca2+
release evoked by carbachol 2.5-fold better than that evoked by CCK
(Fig. 4e). Several control experiments further indicated that inhibition by P1-33 was specific. First, cells
responded normally to addition of IP3, even in the presence
of 62 µM P1-33 (Fig. 4, c and
d). Second, HT-31, another amphipathic peptide that disrupts
protein kinase A anchoring to AKAPs (21), had no effect on signaling in
response to carbachol. Third, amino acid substitutions within
P1-33 dramatically lowered its inhibitory activity (see
legend to Fig. 4). Fourth, P1-33 preferentially inhibited
signaling via the muscarinic receptor compared with the CCK receptor.
An important distinction between P1-33 and RGS4 is that
the inhibitory effect of P1-33 was not reversed by GTP
A final and distinctive activity of P1-33 was that
exposure to both 4Box and P1-33 inhibited Ca2+
release with greater potency than either added alone (compare Fig. 4,
b, c, and h). Importantly, this
inhibition was reversed by GTP Conclusions--
The N-terminal domain and RGS box of RGS4
cooperate via at least two discrete mechanisms to convey
receptor-selective inhibition of G protein signaling. Inhibition of
Ca2+ signaling by RGS4 was receptor-selective whereas
inhibition by 4Box was not. Receptor selectivity of RGS4 inhibition was
not influenced by the identity of the Gq class
Interaction of the N terminus with components of the
receptor-Gq-PLC
Based on our results, we propose that receptor interactions with the N
terminus of RGS4 may help position RGS4 between effector and G protein,
where it is poised to inactivate the G protein 
subunits,
thus attenuating signaling. Mechanisms that provide more precise
regulatory specificity have been elusive. We report here that an
N-terminal domain of RGS4 discriminated among receptor signaling
complexes coupled via Gq. Accordingly, deletion of the
N-terminal domain of RGS4 eliminated receptor selectivity and reduced
potency by 104-fold. Receptor selectivity and potency of
inhibition were partially restored when the RGS4 box was added together
with an N-terminal peptide. In vitro reconstitution
experiments also indicated that sequences flanking the RGS4 box were
essential for high potency GAP activity. Thus, RGS4 regulates
Gq class signaling by the combined action of two domains:
1) the RGS box accelerates GTP hydrolysis by Gq and 2)
the N terminus conveys high affinity and receptor-selective inhibition.
These activities are each required for receptor selectivity and high
potency inhibition of receptor-coupled Gq signaling.
INTRODUCTION
Top
Abstract
Introduction
Materials & Methods
Results & Discussion
References
and phospholipase C-
(PLC)1 (1), which
generates IP3 to trigger Ca2+ release from
internal stores (2). Many cells express several Gq-coupled
receptors that regulate the location, intensity, and propagation of
intracellular Ca2+ waves. For example, pancreatic acini
respond to acetylcholine, bombesin, and cholecystokinin by activating
the same set of Gq class proteins and mobilizing the same
Ca2+ pool, but each receptor evokes distinct patterns of
Ca2+ waves (3). Ca2+ release may be regulated
by intracellular proteins that interact with guanine nucleotide binding
proteins, such as regulators of G protein signaling (RGS)
proteins.2
subunits, thus
attenuating signaling (5-8). Mammals express over 20 different RGS
proteins, of which RGS4 has received the most extensive biochemical
characterization (5, 7-12). RGS4 is composed of a central domain of
120 amino acids that is homologous to other RGS proteins, termed the
RGS box, flanked by less well conserved N- and C-terminal sequences (13). In rat pancreatic acinar cells, RGS4 preferentially inhibited Gq/11-mediated signaling evoked by carbachol relative to
bombesin and cholecystokinin regardless of the identity of the
Gq class subunit.2 Regulatory specificity
was apparently conferred by direct or indirect interaction between RGS4
and the receptor.
q whereas the N terminus conveys high affinity and
receptor-selective inhibition. These combined activities are required
for receptor selectivity and high potency inhibition of
receptor-coupled Gq signaling.
MATERIALS AND METHODS
Top
Abstract
Introduction
Materials & Methods
Results & Discussion
References
current and
Ca2+ release in permeabilized cells was exactly as
described.2
q-GTP were performed as described (15) using
[-32P]GTP bound to the R183C mutant of
Gq (chosen to slow hydrolysis and thus facilitate loading).
RESULTS AND DISCUSSION
Top
Abstract
Introduction
Materials & Methods
Results & Discussion
References
channel. In the control (Fig.
1a), carbachol evoked a
typical biphasic Ca2+ response consisting of an initial
spike caused by Ca2+ release from internal stores followed
by a plateau current. Infusion of full-length RGS4 (10 pM)
suppressed the initial Ca2+ release and caused subsequent
oscillations in response to maximal carbachol stimulation. Increasing
the RGS4 concentration to 100 pM further reduced the
Ca2+ response to a low frequency oscillation. A similar
transition from a sustained to a weak oscillatory response occurred
when the carbachol concentration was reduced from 100 to 1 µM (16, 17).2 This indicates that
carbachol-evoked signaling in intact cells dialyzed with 100 pM RGS4 is functionally equivalent to a 100-fold decrease
in the potency of carbachol stimulation. Exposing cells to higher
concentrations of carbachol did not change the inhibitory effect of
RGS4 on Ca2+ release. Two other proteins related to RGS4,
RGS1, and RGS16 (13, 18, 19) also inhibit carbachol-evoked
Ca2+ release, but RGS10, a GAP for the Gi class
(6, 10), has no effect on Gq-mediated signaling (data not
shown).
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Fig. 1.
Deletion of the RGS4 N terminus destroys high
potency inhibition of Ca2+ release. Cells were
dialyzed for at least 7 min with the pipette solution before the first
stimulation. a, standard response to carbachol
(Car). Inhibition of Ca2+ signaling by
recombinant proteins was assayed with: b and c,
RGS4; d-f, 4Box, 58-177; g and h,
RGS4 
C, 1-177; i and j, RGS4N, 58-205.
Data for RGS4 and 4Box are representative of at least 20 experiments
each (RGS4N, n 
6; RGS4C, n = 3).
i1 with equal potency in
vitro (10), 4Box was approximately 104-fold less
potent than full-length RGS4 in inhibiting in vivo Ca2+ signaling. For example, 10 pM RGS4
conferred partial inhibition of signaling whereas 100 nM
4Box was required before partial inhibition was apparent (Fig. 1,
b and d). Increasing the cellular concentration of full-length RGS4 gradually inhibited signaling, decreasing both the
amplitude of the initial current spike and the frequency and amplitude
of the subsequent current oscillations (Fig. 1, a-c). By
contrast, dialysis with as much as 100 nM 4Box did not significantly reduce the initial current spike but caused rapid termination of signaling and obliterated subsequent oscillations in the
continued presence of agonist (Fig. 1d). These data also indicated that the mechanisms of inhibition by RGS4 and 4Box were qualitatively different, suggesting that inhibition by intact RGS4
involves more than just Gq GAP activity.
C) reduced its potency
by about 100-fold (Fig. 1, g and h). For example, 10 nM RGS4C inhibited the initial current spike to the
same extent as did 100 pM full-length RGS4 and caused
similar subsequent oscillations in response to 100 µM
carbachol (compare Fig. 1, c and h). This effect
was not further characterized because deletion of the N-terminal sequence flanking the RGS box (RGS4N) caused an even more dramatic reduction in potency and altered the mechanism of RGS4 inhibition to
resemble that observed with 4Box (Fig. 1, i and
j). Hence, the N terminus of RGS4 is essential for high
potency inhibition and contributes significantly to RGS4 interaction
with the receptor-Gq protein complex.
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Fig. 2.
4Box is a non-selective inhibitor whereas
P1-33 confers preferential inhibition of carbachol
signaling. Cells were stimulated with 100 µM
carbachol (Car), inhibited with 10 µM atropine
(Atr), and then stimulated with 10 nM CCK8, as
indicated. a, cell dialyzed with buffer.2
b, addition of 100 pM RGS4 showed that
carbachol-dependent signaling was about 15 ± 3 (n = 17)-fold more sensitive to RGS4 than
CCK-dependent signaling.2 4Box inhibited the
response to carbachol and CCK equivalently: c, with 100 nM 4Box, the ratio of CCK to carbachol (CCK/Car) response
was 0.98 ± 0.07 (n = 33); d, 300 nM 4Box, CCK/Car = 1.03 ± 0.16 (n = 3); e, 1 µM 4Box,
CCK/Car = 0.93 ± 0.11 (n = 8); f,
10 µM 4Box, CCK/Car = 0.97 ± 0.15 (n = 5). g-i, P1-33 (100 nM, 300 nM, and 1 µM)
preferentially inhibited the response to carbachol. Similar effects
with increasing concentration of P1-33 were observed in
three experiments with separate cell preparations.
q-GAP Activity in
Vitro--
We then tested the relative GAP activity of RGS4 and 4Box
toward purified, recombinant Gq proteins in
vitro. In a solution phase, single-turnover Gq GAP
assay, RGS4 was about 10-fold more active than 4Box (Fig.
3, a and c), in
contrast to their equal potency when assayed with Gi1
(10). We then compared the GAP activity of full-length RGS4 and 4Box on
wild-type Gq in a steady-state, receptor-coupled in
vitro assay. Phospholipid vesicles were reconstituted with m1
muscarinic cholinergic receptor and trimeric Gq,
wherein steady-state GDP-GTP exchange on Gq was
agonist-dependent (14). In this assay, RGS4 was about
125-fold more potent than 4Box in stimulating steady-state
receptor-dependent GTPase activity (Fig. 3, b and
c). Phospholipid vesicles reconstituted with m2 muscarinic cholinergic receptor and trimeric Gi1 revealed an
even more pronounced difference between the relative activities of
full-length RGS4 and 4Box (data not shown). These findings provide
additional evidence that sequences flanking the RGS box are essential
for optimal GAP activity in receptor-G protein-coupled signaling.
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Fig. 3.
Activation of G q GTPase by
4Box and RGS4. a, single hydrolytic turnover.
GqR183C-[-32P]GTP was incubated in the
presence or absence of RGS4 or 4Box for 30 s at 20 °C to
approximate an initial rate. Data are the average of four
determinations (±S.D.) and are representative of three independent
experiments (full concentration range tested was 5-1000
nM). Basal hydrolysis was linear for up to 1 h, and
this rate was used to calculate the control value. b,
steady-state GTPase of Gq. m1 receptor and
Gq were co-reconstituted into phospholipid
vesicles as described (14). Steady-state GTPase activity was measured
in the presence of either 1 mM carbachol (black
bars) or 10 µM atropine (not shown; basal GTP
hydrolysis was below 0.12 fmol/min/fmol of Gq).
c, relative RGS4 and 4Box activities, shown as a ratio in
the two assay conditions, were calculated from the data in a
and b and additional experiments.
S to reverse inhibition by RGS4 as a test
of its Gq GAP activity.
S (Fig. 4f), suggesting that
inhibition of signaling reflects the GAP activity of RGS4 under these
conditions. By contrast, addition of up to 1.4 µM 4Box to
the incubation medium had no effect on carbachol-stimulated Ca2+ release (Fig. 4b) further demonstrating
that full-length RGS4 inhibits Gq-coupled signaling far
more potently than does 4Box.
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Fig. 4.
The RGS4 N-terminal peptide
P1-33 and 4Box act synergistically to inhibit
carbachol-stimulated release of intracellular Ca2+.
Pancreatic acini were incubated in medium (a) or different
concentrations of 4Box (b, h, i), P1-33
(c, d, e, g, h, i), and/or full-length RGS4 (f,
g). Cells were stimulated with 2 mM carbachol and then
treated sequentially with 2.5 µM GTP S and 2 µM IP3 to test for their ability to activate
Gq and to release stored Ca2+, respectively.
Similar results were obtained in at least three additional experiments.
In control experiments, the three Cys residues at positions 2, 12, and
33 of P1-33 were replaced with Ala
(P1-33CA2,12,33), which reduced the
effectiveness of the peptide by at least 100-fold (n = 6). The peptide P13-32 was 10-fold less effective than
P1-33 (n = 3). Secondary structure
analysis predicts P1-33 to have a hydrophobic N terminus
(residues 1-10) followed by an amphipathic helix. In additional
controls, the amphipathic peptide HT-31 (21) at 0.5 mM had
no effect on agonist- or IP3-induced Ca2+
release. P1-33 (1 µM) did not accelerate GTP
hydrolysis by Gq in the steady-state assay (Fig. 3), and
P1-33 (200 µM) only marginally stimulated
GTP hydrolysis by Gi1-bound GTP in the single-turnover
assay (10).
S (Fig. 4, c and d), indicating that its effect
does not depend on Gq-GAP activity. Sequential addition of
P1-33 and RGS4 blocked GTPS-insensitive inhibition
(Fig. 4g), indicating the peptide and full-length RGS4
compete for binding to a subsite of the RGS4 binding site in the
receptor-Gq complex. To further investigate
P1-33 inhibition of Ca2+ release, we changed
the order of addition of inhibitors and activators. If cells were
exposed to P1-33, then carbachol, subsequent addition of
either RGS4 or 4Box did not restore the ability of GTPS to stimulate
Ca2+ release (similar to Fig. 4d). RGS4 and 4Box
only worked when added prior to carbachol stimulation (Fig. 4,
f-h). P1-33 does not prevent
receptor-catalyzed GTP exchange on Gq nor does P1-33 act as a Gq GAP (data not shown). This
overall pattern of activities suggests that P1-33 binds at
a site normally occupied by the N terminus of RGS4, and when added
without 4Box, P1-33 blocks interaction of Gq
with its effector protein PLC.
S, indicating that the GAP activity of
4Box was functional in permeabilized cells (Fig. 4, h and
i). In cells dialyzed with P1-33 (10 nM) and 4Box (1 nM) through a patch pipette, at concentrations at which neither had activity on its own, the
combination fully inhibited carbachol-evoked Ca2+ signaling
(data not shown). Thus, the peptide P1-33 and 4Box
mutually influenced the activity of each other in cells. We propose the
P1-33 peptide facilitates 4Box interaction with the
receptor complex (reflected by enhanced potency) and the 4Box alters
the P1-33 binding site to relieve GTPS-resistant inhibition. The analysis in Fig. 4 suggests two separate regions of
RGS4 interact with the receptor complex to regulate Ca2+
signaling. A combination of the N-terminal peptide P1-33 and 4Box reconstituted the essential functions of RGS4, those of
receptor specificity and GAP activity. Sequence divergence in the N
terminus of different RGS proteins may convey regulatory specificity
toward other receptor complexes, and this specificity may be retained
in peptides analogous with P1-33.
subunit.2 Such selectivity suggests intact RGS4 interacts
directly or indirectly with receptors, most likely through its N
terminus. This is supported by the finding that RGS4 in patch-clamped
cells was 10,000-fold more potent than 4Box and RGS4 had 125-fold
higher Gq-GAP activity than did 4Box in an
agonist-dependent in vitro assay. Finally, partial inhibition by RGS4 simultaneously decreased both the amplitude of the initial Ca2+ signal and the frequency of subsequent
oscillations, suggesting recombinant RGS4 protein is active when
dialyzed into patch-clamped cells and can preassemble with the
receptor-Gq-PLC signaling complex. By contrast, 4Box
preferentially inhibited signaling following the initial
Ca2+ release evoked by carbachol and CCK. This suggests
that 4Box is recruited only to active receptor complexes when
production of the Gq-GTP substrate would be greatest, and
thereby 4Box GAP activity would be most pronounced.
signaling complex is further supported
by the ability of the N-terminal peptide P1-33 to inhibit
Ca2+ signaling in the absence of 4Box. Inhibition by
P1-33 displayed the same selectivity among receptors as
displayed by full-length RGS4. In contrast to RGS4, however, inhibition
by P1-33 was not overcome by GTPS, possibly because
P1-33 blocked access of Gq-GTPS to its
effector protein PLC. Cooperation between the N terminus and 4Box is
most apparent in that a combination of both P1-33 and 4Box
restored the regulatory activity of intact RGS4 in cells, including its
reversibility by GTPS. Thus, the N terminus provides anchorage (4,
22) and receptor selectivity whereas the RGS box acts as a GAP, and
both domains combine to yield the behavior of the intact protein.
subunit via the GAP
activity of the RGS box, even in the presence of persistent agonist
(Fig. 5). Subcellular co-localization of receptor, RGS proteins, and
downstream signaling proteins, including Gq and PLC,
could provide an additional level of regulatory specificity. Thus, the
multiple effects of RGS4 described here may contribute to the temporal
and spatial regulation that is a hallmark of intracellular Ca2+ signaling.
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Fig. 5.
Model of RGS4 interaction with the receptor
signaling complex. We propose that the N terminus of RGS4 mediates
high affinity interaction with Gq-coupled receptors. The
RGS4 box provides Gq-GAP activity. RGS4 is modeled to
reside within the complex on the path of G q between the
receptor, which catalyzes GTP binding in the presence of agonist, and
the effector protein PLC. We propose that active RGS4 accelerates
GTP hydrolysis on Gq before its interaction with PLC,
thus inhibiting the production of IP3 and subsequent
Ca2+ release.
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ACKNOWLEDGEMENTS |
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We thank Melanie Cobb, David Corey, Rama Ranganathan, Kai Zinn, and members of our laboratories for comments on the manuscript.
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FOOTNOTES |
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* This work was funded by a postdoctoral training fellowship and grants from the National Institutes of Health and by the R. A. Welch Foundation.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
§ These authors contributed equally to this work.
** Recipient of the American Heart Association Established Investigator Award. To whom correspondence should be addressed: Dept. of Pharmacology, UT-Southwestern, 5323 Harry Hines Blvd., Dallas, TX 75235-9041. E-mail: wilkie{at}utsw.swmed.edu.
The abbreviations used are:
PLC, phospholipase
C; IP3, inositol trisphosphate; RGS, regulators of G
protein signaling; GAP, GTPase-activating protein; 4Box, RGS domain of
RGS4; P1-33, N-terminal 33 amino acids of RGS4; Car, carbachol; CCK, cholecystokinin; GTPS, guanosine
5'-3-O-(thio)triphosphate.
2 Xu, X., Zeng, W., Popov, S., Berman, D. M., Davignon, I., Yu, K., Yowe, D., Offermanns, S., Muallem, S., and Wilkie, T. M. (1999) J. Biol. Chem., in press.
3 P1-33, MCKGLAGLPASCLRSAKDMKHRLGFLLQKSDSC (Ro27-3948); P1-33CA2,12,33, MAKGLAGLPASALRSAKDMKHRLGFLLQKSDSA (Ro27-3949); P13-32, LRSAKDMKHRLGFLLQKSDS (Ro27-3950); HT-31, DLIEEAASRIVDAVIEQVKAAY (Ro27-1970).
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REFERENCES |
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Top
Abstract
Introduction
Materials & Methods
Results & Discussion
References
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A. Benians, M. Nobles, S. Hosny, and A. Tinker Regulators of G-protein Signaling Form a Quaternary Complex with the Agonist, Receptor, and G-protein: A NOVEL EXPLANATION FOR THE ACCELERATION OF SIGNALING ACTIVATION KINETICS J. Biol. Chem., April 8, 2005; 280(14): 13383 - 13394. [Abstract] [Full Text] [PDF]
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 G.-x. Xie and P. P. Palmer RGS Proteins: New Players in the Field of Opioid Signaling and Tolerance Mechanisms Anesth. Analg., April 1, 2005; 100(4): 1034 - 1042. [Abstract] [Full Text] [PDF]
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S. C. Tovey and G. B. Willars Single-Cell Imaging of Intracellular Ca2+ and Phospholipase C Activity Reveals That RGS 2, 3, and 4 Differentially Regulate Signaling via the G{alpha}q/11-Linked Muscarinic M3 Receptor Mol. Pharmacol., December 1, 2004; 66(6): 1453 - 1464. [Abstract] [Full Text] [PDF]
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T. M. Cabrera-Vera, S. Hernandez, L. R. Earls, M. Medkova, A. K. Sundgren-Andersson, D. J. Surmeier, and H. E. Hamm RGS9-2 modulates D2 dopamine receptor-mediated Ca2+ channel inhibition in rat striatal cholinergic interneurons PNAS, November 16, 2004; 101(46): 16339 - 16344. [Abstract] [Full Text] [PDF]
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S. J. Bornheimer, M. R. Maurya, M. G. Farquhar, and S. Subramaniam Computational modeling reveals how interplay between components of a GTPase-cycle module regulates signal transduction PNAS, November 9, 2004; 101(45): 15899 - 15904. [Abstract] [Full Text] [PDF]
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X. Wang, G. Huang, X. Luo, J. M. Penninger, and S. Muallem Role of Regulator of G Protein Signaling 2 (RGS2) in Ca2+ Oscillations and Adaptation of Ca2+ Signaling to Reduce Excitability of RGS2-/- Cells J. Biol. Chem., October 1, 2004; 279(40): 41642 - 41649. [Abstract] [Full Text] [PDF]
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H. Mao, Q. Zhao, M. Daigle, M. H. Ghahremani, P. Chidiac, and P. R. Albert RGS17/RGSZ2, a Novel Regulator of Gi/o, Gz, and Gq Signaling J. Biol. Chem., June 18, 2004; 279(25): 26314 - 26322. [Abstract] [Full Text] [PDF]
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L. S. Bernstein, S. Ramineni, C. Hague, W. Cladman, P. Chidiac, A. I. Levey, and J. R. Hepler RGS2 Binds Directly and Selectively to the M1 Muscarinic Acetylcholine Receptor Third Intracellular Loop to Modulate Gq/11{alpha} Signaling J. Biol. Chem., May 14, 2004; 279(20): 21248 - 21256. [Abstract] [Full Text] [PDF]
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Z. Liu and R. A. Fisher RGS6 Interacts with DMAP1 and DNMT1 and Inhibits DMAP1 Transcriptional Repressor Activity J. Biol. Chem., April 2, 2004; 279(14): 14120 - 14128. [Abstract] [Full Text] [PDF]
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A. M. Krumins, S. A. Barker, C. Huang, R. K. Sunahara, K. Yu, T. M. Wilkie, S. J. Gold, and S. M. Mumby Differentially Regulated Expression of Endogenous RGS4 and RGS7 J. Biol. Chem., January 23, 2004; 279(4): 2593 - 2599. [Abstract] [Full Text] [PDF]
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A. A. Roy, K. E. Lemberg, and P. Chidiac Recruitment of RGS2 and RGS4 to the Plasma Membrane by G Proteins and Receptors Reflects Functional Interactions Mol. Pharmacol., September 1, 2003; 64(3): 587 - 593. [Abstract] [Full Text] [PDF]
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T. K. Chatterjee, Z. Liu, and R. A. Fisher Human RGS6 Gene Structure, Complex Alternative Splicing, and Role of N Terminus and G Protein {gamma}-Subunit-like (GGL) Domain in Subcellular Localization of RGS6 Splice Variants J. Biol. Chem., August 8, 2003; 278(32): 30261 - 30271. [Abstract] [Full Text] [PDF]
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T. K. Chatterjee and R. A. Fisher Mild Heat and Proteotoxic Stress Promote Unique Subcellular Trafficking and Nucleolar Accumulation of RGS6 and Other RGS Proteins: ROLE OF THE RGS DOMAIN IN STRESS-INDUCED TRAFFICKING OF RGS PROTEINS J. Biol. Chem., August 8, 2003; 278(32): 30272 - 30282. [Abstract] [Full Text] [PDF]
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 D. M. Shin, M. Dehoff, X. Luo, S. H. Kang, J. Tu, S. K. Nayak, E. M. Ross, P. F. Worley, and S. Muallem Homer 2 tunes G protein-coupled receptors stimulus intensity by regulating RGS proteins and PLC{beta} GAP activities J. Cell Biol., July 21, 2003; 162(2): 293 - 303. [Abstract] [Full Text] [PDF]
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P. Tosetti, N. Pathak, M. H. Jacob, and K. Dunlap RGS3 mediates a calcium-dependent termination of G protein signaling in sensory neurons PNAS, June 10, 2003; 100(12): 7337 - 7342. [Abstract] [Full Text] [PDF]
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J. L. Osterhout, A. A. Waheed, A. Hiol, R. J. Ward, P. C. Davey, L. Nini, J. Wang, G. Milligan, T. L. Z. Jones, and K. M. Druey Palmitoylation Regulates Regulator of G-protein Signaling (RGS) 16 Function: II. PALMITOYLATION OF A CYSTEINE RESIDUE IN THE RGS BOX IS CRITICAL FOR RGS16 GTPase ACCELERATING ACTIVITY AND REGULATION OF Gi-COUPLED SIGNALING J. Biol. Chem., May 23, 2003; 278(21): 19309 - 19316. [Abstract] [Full Text] [PDF]
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S. Salim, S. Sinnarajah, J. H. Kehrl, and C. W. Dessauer Identification of RGS2 and Type V Adenylyl Cyclase Interaction Sites J. Biol. Chem., April 25, 2003; 278(18): 15842 - 15849. [Abstract] [Full Text] [PDF]
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