Transforming Growth Factor-β1 Inhibits Type I Inositol 1,4,5-Trisphosphate Receptor Expression and Enhances Its Phosphorylation in Mesangial Cells*
- From the ‡Department of Medicine, Nephrology Division and the ¶Department of Anatomy, Pathology, and Cell Biology, Thomas Jefferson University School of Medicine, Philadelphia, Pennsylvania 19107
Abstract
A potentially important cross-talk characteristic of transforming growth factor-β (TGF-β) is to inhibit platelet-derived growth factor-induced intracellular calcium rise (Baffy, G., Sharma, K., Shi, W., Ziyadeh, F. N., and Williamson, J. R. (1995) Biochem. Biophys. Res. Commun. 210, 378–383) in murine mesangial cells. The present study examined the possible basis for this effect by evaluating the regulation of the type I inositol 1,4,5-trisphosphate receptor (IP3R) by TGF-β. TGF-β1 down-regulates IP3R protein expression by >90% with maximal and half-maximal effects after 8 and 2 h, respectively. TGF-β1 also decreased IP3R mRNA expression by 59% after 1 h. Phosphorylation of the IP3R was also demonstrated as early as 15 min after TGF-β1 exposure. Back phosphorylation assays of IP3R from TGF-β1-treated mesangial cells with protein kinase A (PKA), indicated that TGF-β1-induced phosphorylation of the IP3R occurs at similar sites as for PKA. In vitro kinase assays using the known IP3R peptide substrates for PKA, RPSGRRESLTSFGNP and ARRDSVLAAS, demonstrated that TGF-β1 induces phosphorylation of both peptides (158 and 123% of control values, respectively). TGF-β1-induced phosphorylation was prevented by the addition of the PKA inhibitor peptide in the in vitro kinase assay. It is proposed that TGF-β-mediated effects on the IP3R may be an important characteristic of its ability to modulate the response of cells to factors that employ IP3R-mediated calcium release.
Footnotes
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↵* This work was supported by National Institutes of Health Grant KO8 DK02308 (to K. S.), a National Kidney Foundation Young Investigator Award (to K. S.), and by National Institutes of Health Post-doctoral Training Grant T32-AA07463 (to S. B.) and Grant R01-AA10971 (to S. K. J.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵§ To whom correspondence should be addressed: Div. of Nephrology, Dept. of Medicine, Thomas Jefferson University, Suite 353, JAH, 1020 Locust St., Philadelphia, PA 19107. Tel.: 215-503-6950; Fax: 215-923-7212; E-mail: sharma1{at}jeflin.tju.edu.
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↵1 The abbreviations used are: TGF, transforming growth factor; IP3, inositol 1,4,5-trisphosphate; IP3R, IP3 receptor; PDGF, platelet-derived growth factor; MMC, murine mesangial cells; PBS, phosphate-buffered saline; DMEM, Dulbecco’s modified Eagle’s medium; PKA, protein kinase A; PKI, protein kinase A inhibitor; PMSF, phenylmethylsulfonyl fluoride; PAGE, polyacrylamide gel electrophoresis; PCR, polymerase chain reaction; bp, base pair.
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↵2 K. Sharma, unpublished observations.
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- Received January 10, 1997.
- Revision received March 18, 1997.
