Relationship between Internalization and mRNA Decay in Down-Regulation of Recombinant Type 1 Angiotensin II Receptor (AT1) Expression in Smooth Muscle Cells
- 1Department of Pharmacology (B.A., T.S.O., X.W., T.J.M.) and2Graduate Program in Molecular Therapeutics and Toxicology (B.A., X.W., T.J.M.), Graduate Division of Biological and Biomedical Sciences, Emory University School of Medicine, Atlanta, Georgia
Abstract
In vascular smooth muscle cells, the hormone angiotensin II is thought to cause internalization of the seven-transmembrane domain type 1 angiotensin II receptor (AT1-R) but it also suppresses expression of the receptor mRNA. As for similarly regulated members of this gene superfamily, the relative roles of these processes in receptor down-regulation are not well understood. In this study a recombinant AT1-R mRNA was synthesized in A7r5 vascular smooth muscle cells from a tetracycline-suppressible promoter using a retroviral vector system. Angiotensin II induces a profound internalization of the cell surface AT1-R protein but has no effect on steady-state AT1-R mRNA levels. Shortly after either bolus or prolonged dosing with angiotensin II, cell surface AT1-R expression recovers, indicating the existence of a significant restorative externalization pathway. The extent of this recovery is attenuated markedly when transcription of the recombinant AT1-R gene is suppressed by cotreatment of the cells with anhydrotetracycline. Although agonist-stimulated internalization appears to contribute directly to a loss of AT1-R protein, these observations provide direct evidence that a reduction in AT1-R mRNA content plays a significant role in sustained AT1-R down-regulation.
Footnotes
-
Send reprint requests to: Dr. T. J. Murphy, Department of Pharmacology, Emory University School of Medicine, Room 5031, O. W. Rollins Research Center, 1510 Clifton Rd., Atlanta, GA 30322. E-mail: tmurphy{at}pharm.emory.edu
-
This work was supported by Grants HL52180 and HL56107 from the National Heart Lung and Blood Institute. Further support was obtained from a Grant in Aid from the American Heart Association and Sanofi-Winthrop. T. J. M. is an Established Investigator of the American Heart Association.
- Abbreviations:
- AT1-R
- type 1 angiotensin II receptor
- VSMC
- vascular smooth muscle cells
- AnTet
- anhydrotetracycline
- AngII
- angiotensin II
- Sarile
- Sar1, Ile8-angiotensin II
- RNase
- ribonuclease
- tTA
- tetracycline transactivator
-
- Received October 7, 1998.
- Accepted March 16, 1999.
- The American Society for Pharmacology and Experimental Therapeutics
