Abstract
Muscarinic acetylcholine receptors modulate the function of a variety of effectors through heterotrimeric G proteins. A prenylated peptide specific to the G protein gamma5 subunit type inhibits G protein activation by the M2 muscarinic receptor in a reconstitution assay. Scrambling the amino acid sequence of the peptide significantly reduces the efficacy of the peptide. The peptide does not disrupt the G protein heterotrimer. In cultured sympathetic neurons, the gamma5 peptide inhibits modulation of Ca(2+) current by the M4 receptor. Peptide activity is specific, the scrambled peptide and peptides specific to two other members of the G protein gamma subunit family are significantly less effective. The gamma5 peptide has no effect on Ca(2+) current modulation by the alpha2-adrenergic and somatostatin receptors. In addition, the gamma5 peptide inhibits muscarinic receptor signaling in spinal cord slices with specificity. These results support a specific role for G protein gamma subunit types in signal transduction, most likely at the receptor-G protein interface.
Highlights
Muscarinic acetylcholine receptors modulate the function of a variety of effectors through heterotrimeric G proteins
␥5 Subunit Type-specific Peptide Inhibits G Protein Activation by M2—Membranes from CHO cells expressing high levels of M2 receptor were depleted of endogenous G protein subunits as described in the methods section and assayed for activity by binding of antagonist
When M2 containing membranes were reconstituted with heterotrimeric Gi2, GTP␥S binding by the G protein was stimulated 5-fold by the agonist, carbachol, compared with the antagonist, atropine
Summary
Muscarinic acetylcholine receptors modulate the function of a variety of effectors through heterotrimeric G proteins. To examine the effect of the peptide in cells, we injected a peptide specific to the ␥5 COOH terminus into superior cervical ganglion (SCG)1 neurons and measured receptor modulation of N-type Ca2ϩ current (Ica).
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