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.
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.
Heng
H
Xu
Xiaoying
X
Wang
Darin
D
Zimmerman
Emily S
ES
Boja
Jiabei
J
Wang
Edward J
EJ
Bilsky
Richard B
RB
Rothman
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