Faculty Collaboration Database

M3 muscarinic acetylcholine receptors regulate cytoplasmic myosin by a process involving RhoA and requiring conventional protein kinase C isoforms. J Biol Chem 1999 Jun 25;274(26):18675-85

Date

06/22/1999

Pubmed ID

10373480

DOI

10.1074/jbc.274.26.18675

Scopus ID

2-s2.0-0033603052 (requires institutional sign-in at Scopus site)   43 Citations

Abstract

Although muscarinic acetylcholine receptors (mAChR) regulate the activity of smooth muscle myosin, the effects of mAChR activation on cytoplasmic myosin have not been characterized. We found that activation of transfected human M3 mAChR induces the phosphorylation of myosin light chains (MLC) and the formation of myosin-containing stress fibers in Chinese hamster ovary (CHO-m3) cells. Direct activation of protein kinase C (PKC) with phorbol 12-myristate 13-acetate (PMA) also induces myosin light chain phosphorylation and myosin reorganization in CHO-m3 cells. Conventional (alpha), novel (delta), and atypical (iota) PKC isoforms are activated by mAChR stimulation or PMA treatment in CHO-m3 cells, as indicated by PKC translocation or degradation. mAChR-mediated myosin reorganization is abolished by inhibiting conventional PKC isoforms with Go6976 (IC50 = 0.4 microM), calphostin C (IC50 = 2.4 microM), or chelerythrine (IC50 = 8.0 microM). Stable expression of dominant negative RhoAAsn-19 diminishes, but does not abolish, mAChR-mediated myosin reorganization in the CHO-m3 cells. Similarly, mAChR-mediated myosin reorganization is diminished, but not abolished, in CHO-m3 cells which are multi-nucleate due to inactivation of Rho with C3 exoenzyme. Expression of dominant negative RhoAAsn-19 or inactivation of RhoA with C3 exoenzyme does not affect PMA-induced myosin reorganization. These findings indicate that the PKC-mediated pathway of myosin reorganization (induced either by M3 mAChR activation or PMA treatment) can continue to operate even when RhoA activity is diminished in CHO-m3 cells. Conventional PKC isoforms and RhoA may participate in separate but parallel pathways induced by M3 mAChR activation to regulate cytoplasmic myosin. Changes in cytoplasmic myosin elicited by M3 mAChR activation may contribute to the unique ability of these receptors to regulate cell morphology, adhesion, and proliferation.

Author List

Strassheim D, May LG, Varker KA, Puhl HL, Phelps SH, Porter RA, Aronstam RS, Noti JD, Williams CL

Author

Carol L. Williams PhD Professor in the Pharmacology and Toxicology department at Medical College of Wisconsin

MESH terms used to index this publication - Major topics in bold

ADP Ribose Transferases
Animals
Botulinum Toxins
CHO Cells
Carbachol
Cricetinae
Cytoplasm
GTP-Binding Proteins
Humans
Isoenzymes
Myosins
Protein Kinase C
Receptor, Muscarinic M1
Receptor, Muscarinic M3
Receptors, Muscarinic
Tetradecanoylphorbol Acetate
rhoA GTP-Binding Protein