Chronic morphine up-regulates G alpha12 and cytoskeletal proteins in Chinese hamster ovary cells expressing the cloned mu opioid receptor.
A growing body of literature indicates that chronic morphine exposure alters the expression and function of cytoskeletal proteins in addition to the well established interactions between mu opioid receptors and G proteins. In the present study, we hypothesized that chronic morphine alters the expression and functional effects of G alpha12, a G protein that regulates downstream cytoskeletal proteins via its control of RhoA. Our results showed that chronic morphine treatment decreased the expression of G alpha i2 (64%) and G alpha i3 (60%), had no effect of G alpha o, and increased G alpha12 (66%) expression in Chinese hamster ovary (CHO) cells expressing the cloned human mu opioid receptors (hMOR-CHO cells) but not in cells expressing a mutant mu opioid receptor that do not develop morphine tolerance and dependence (T394A-CHO cells). Morphine treatment had no significant effect on PAR-1 thrombin receptor-activated G protein activity, as measured by thrombin-stimulated guanosine 5'-O-(3-[35S]thio)triphosphate binding. Chronic morphine treatment significantly enhanced thrombin-stimulated RhoA activity and thrombin-stimulated expression of alpha-actinin, a cytoskeletal anchoring protein, in hMOR-CHO cells. Proteomic analysis of two-dimensional gel spots prepared from hMOR-CHO cells showed that morphine treatment affected the expression of a number of proteins associated with morphological changes. Up-regulation of G alpha12 and alpha-actinin by chronic morphine was also observed in mouse brain. Viewed collectively, these findings indicate, for the first time, that chronic morphine enhances the G alpha12-associated signaling system, which is involved in regulating cellular morphology and growth, supporting other findings that chronic morphine may alter cellular morphology, in addition to cellular function.
Xu
Heng
H
Clinical Psychopharmacology Section, Intramural Research Program, National Institute on Drug Abuse, National Institutes of Health, Department of Health and Human Services, Baltimore, MD 21224-2735, USA.
Wang
Xiaoying
X
Zimmerman
Darin
D
Boja
Emily S
ES
Wang
Jiabei
J
Bilsky
Edward J
EJ
Rothman
Richard B
RB
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003598:Cytoskeletal Proteins / Q000096:biosynthesis*; D044365:GTP-Binding Protein alpha Subunits, G12-G13 / Q000096:biosynthesis*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D008969:Molecular Sequence Data; D009020:Morphine / Q000494:pharmacology*; D040901:Proteomics; D017450:Receptors, Opioid, mu / Q000502:physiology*; D018179:Receptors, Thrombin / Q000502:physiology; D015854:Up-Regulation; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
D003598:Cytoskeletal Proteins; D017450:Receptors, Opioid, mu; D018179:Receptors, Thrombin; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D009020:Morphine; D044365:GTP-Binding Protein alpha Subunits, G12-G13; D020742:rhoA GTP-Binding Protein