The M3 Muscarinic Acetylcholine Receptor Expressed in HEK-293 Cells Signals to Phospholipase D via G12 but Not Gq-type G Proteins

Rümenapp, Ulrich; Asmus, Melanie; Schablowski, Helge; Woznicki, Markus; Han, Li; Jakobs, Karl H.; Fahimi-Vahid, Mercedeh; Michalek, Christina; Wieland, Thomas; Schmidt, Martina

doi:10.1074/jbc.m004957200

Cited by 79 publications

(35 citation statements)

References 47 publications

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“…In this report, we presented several lines of evidence that p63RhoGEF, a novel member of the Dbl family of GEFs, might represent the specific GEF, or at least one of the GEFs, mediating this response. Firstly, p63RhoGEF largely enhanced the transcriptional activity of the primarily G q/11 -coupled M 3 R and H 1 R in a manner sensitive to RGS2, a negative regulator of G q/11 activity (25,26), but insensitive to the G 12/13 -specific Lsc-RGS (22). Secondly, it largely enhanced the Rho-mediated gene transcription induced by the GTPase-deficient mutants of G␣ q and G␣ 11 but not of G␣ 12 and G␣ 13 .…”

Section: Discussionmentioning

confidence: 99%

“…1C). In contrast, the expression of the RGS domain of the mouse ortholog of p115RhoGEF Lsc (Lsc-RGS), which is a G␣ 12/13 -specific GAP and thereby suppresses G␣ 12/13 -mediated responses in HEK-293 cells (22), only slightly reduced (about 10 -20%) the M 3 Rinduced luciferase production. The transcriptional activity induced by the agonist-stimulated M 3 R coexpressed with p63RhoGEF was not affected by Lsc-RGS expression.…”

Section: Synergistic Activation Of Rho-mediated Gene Transcription Bymentioning

confidence: 99%

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The Guanine Nucleotide Exchange Factor p63RhoGEF, a Specific Link between Gq/11-coupled Receptor Signaling and RhoA

Lutz

Freichel-Blomquist

Yang

et al. 2005

Journal of Biological Chemistry

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182

160

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The monomeric GTPase RhoA, which is a key regulator of numerous cellular processes, is activated by a variety of G protein-coupled receptors, through either G 12 or G q family proteins. Here we report that p63RhoGEF, a recently identified RhoA-specific guanine nucleotide exchange factor, enhances the Rho-dependent gene transcription induced by agonist-stimulated G q/11 -coupled receptors (M 3 -cholinoceptor, histamine H 1 receptor) or GTPase-deficient mutants of G␣ q and G␣ 11 . We further demonstrate that active G␣ q or G␣ 11 , but not G␣ 12 or G␣ 13 , strongly enhances p63RhoGEF-induced RhoA activation by direct protein-protein interaction with p63RhoGEF at its C-terminal half. Moreover, the activation of p63RhoGEF by G␣ q/11 occurs independently of and in competition to the activation of the canonical G␣ q/11 effector phospholipase C ␤. Therefore, our results elucidate a new signaling pathway by which G q/11 -coupled receptors specifically induce Rho signaling through a direct interaction of activated G␣ q/11 subunits with p63RhoGEF.

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Section: Discussionmentioning

confidence: 99%

Section: Synergistic Activation Of Rho-mediated Gene Transcription Bymentioning

confidence: 99%

The Guanine Nucleotide Exchange Factor p63RhoGEF, a Specific Link between Gq/11-coupled Receptor Signaling and RhoA

Lutz

Freichel-Blomquist

Yang

et al. 2005

Journal of Biological Chemistry

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182

160

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“…1C). The expression of specific RGS domains is a well characterized method to selectively block signaling through different subfamilies of heterotrimeric G proteins (9)(10)(11), and the p115-RGS domain selectively binds and inactivates members of the G12 family (9,10). In addition, to inhibit Gq signaling in breast cancer cells, we expressed RGS2, an RGS that selectively binds and inactivates members of the Gq family (11) (Fig.…”

Section: G12 Signaling Does Not Promote the Growth Or Transformation Ofmentioning

confidence: 99%

The G12 family of heterotrimeric G proteins promotes breast cancer invasion and metastasis

Kelly

Moeller²,

Juneja

et al. 2006

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

159

191

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Although the prognosis for patients with early-stage breast cancer has improved, the therapeutic options for patients with locally advanced and metastatic disease are limited. To improve the treatment of these patients, the molecular mechanisms underlying breast cancer invasion and metastasis must be understood. In this study, we report that signaling through the G12 family of heterotrimeric G proteins (G␣12 and G␣13) promotes breast cancer cell invasion. Moreover, we demonstrate that inhibition of G12 signaling reduces the metastatic dissemination of breast cancer cells in vivo. Finally, we demonstrate that the expression of G␣12 is significantly up-regulated in the earliest stages of breast cancer, implying that amplification of G12 signaling may be an early event in breast cancer progression. Taken together, these observations identify the G12 family proteins as important regulators of breast cancer invasion and suggest that these proteins may be targeted to limit invasion-and metastasis-induced patient morbidity and mortality.cadherin ͉ Rho ͉ G protein-coupled receptor

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“…These receptors are members of a family of receptors that are coupled to heterotrimeric transducer G proteins. The M 1 , M 3 , and M 5 acetylcholine receptor subtypes are efficiently coupled to the pertussis toxin-insensitive G␣ q/11 and G 13 subtype G proteins, leading to activations of PLC and PLD, whereas the M 2 , and M 4 receptors are coupled to pertussis toxin-sensitive G i and G 0 subtype G proteins (27)(28)(29). The mechanism of PLD activation by the muscarinic receptor has been mainly studied for the M 3 receptor subtype.…”

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confidence: 99%

Inhibition of Muscarinic Receptor-linked Phospholipase D Activation by Association with Tubulin

Chae

Lee

et al. 2005

Journal of Biological Chemistry

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Mammalian phospholipase D (PLD) is considered a key enzyme in the transmission signals from various receptors including muscarinic receptors. PLD activation is a rapid and transient process, but a negative regulator has not been found that inhibits signal-dependent PLD activation. Here, for the first time, we report that tubulin binding to PLD 2 is an inhibition mechanism for muscarinic receptor-linked PLD 2 activation. Tubulin was identified in an immunoprecipitated PLD 2 complex from COS-7 cells by peptide mass fingerprinting. The direct interaction between PLD 2 and tubulin was found to be mediated by a specific region of PLD 2 (amino acids 476 -612). PLD 2 was potently inhibited (IC 50 <10 nM) by tubulin binding in vitro. In cells, the interaction between PLD 2 and tubulin was increased by the microtubule disrupting agent nocodazole and reduced by the microtubule stabilizing agent Taxol. Moreover, PLD 2 activity was found to be inversely correlated with the level of monomeric tubulin. In addition, we found that interaction with and the inhibition of PLD 2 by monomeric tubulin is important for the muscarinic receptor-linked PLD signaling pathway. Interaction between PLD 2 and tubulin was increased only after 1-2 min of carbachol stimulation when carbachol-stimulated PLD 2 activity was decreased. The expression of the tubulin binding region of PLD 2 blocked the later decrease in carbachol-induced PLD activity by masking tubulin binding. Taken together, these results indicate that an increase in local membrane monomeric tubulin concentration inhibits PLD 2 activity, and provides a novel mechanism for the inhibition of muscarinic receptor-induced PLD 2 activation by interaction with tubulin. Mammalian phospholipase D (PLD)1 hydrolyzes membrane phosphatidylcholine to generate phosphatidic acid and choline. PLD activity is dramatically activated in response to a variety of signals, including hormones, neurotransmitters, and growth factors (1). PLD products, phosphatidic acid itself or the hydrolyzed product diacylglycerol, have been known to act as intracellular lipid second messengers and to be involved in multiple physiological events, such as, the promotion of mitogenesis, stimulation of respiratory bursts, secretory processes, and actin cytoskeletal reorganization (2-7). Therefore, signal-dependent PLD activity must be tightly controlled in cells.Many reports have been issued about the mechanisms of receptor-mediated PLD activation. Although the mammalian PLD isozymes, PLD 1 and PLD 2 , have some different regulatory properties, in general, agonist-induced PLD is activated by various protein kinases, including protein kinase C, proteintyrosine kinase, and the MAP kinase family, in addition to small G proteins of the ARF, Rho, and Ras families (8 -13). The signal-dependent activation of PLD is rapid and transient. Although, the activation kinetics depend on the stimulus and cell type, the PLD signal is usually diminished within 10 min (14). However, the mechanisms involved in PLD signal inhibition remain unknow...

show abstract

The M3 Muscarinic Acetylcholine Receptor Expressed in HEK-293 Cells Signals to Phospholipase D via G12 but Not Gq-type G Proteins

Cited by 79 publications

References 47 publications

The Guanine Nucleotide Exchange Factor p63RhoGEF, a Specific Link between Gq/11-coupled Receptor Signaling and RhoA

The Guanine Nucleotide Exchange Factor p63RhoGEF, a Specific Link between Gq/11-coupled Receptor Signaling and RhoA

The G12 family of heterotrimeric G proteins promotes breast cancer invasion and metastasis

Inhibition of Muscarinic Receptor-linked Phospholipase D Activation by Association with Tubulin

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