Disruption of Gap Junctional Communication by the Platelet-derived Growth Factor Is Mediated via Multiple Signaling Pathways*
- From ‡Molecular Medicine, Northwest Hospital, Seattle, Washington 98125 and the ¶Schepens Eye Research Institute, Harvard Medical School, Boston, Massachusetts 02114
Abstract
The platelet-derived growth factor (PDGF) mediates its cellular functions via activation of its receptor tyrosine kinase followed by the recruitment and activation of several signaling molecules. These signaling molecules then initiate specific signaling cascades, finally resulting in distinct physiological effects. To delineate the PDGF signaling pathway responsible for the disruption of gap junctional communication (GJC), wild-type PDGF receptor β (PDGFRβ) and a series of PDGFRβ mutants were expressed in T51B rat liver epithelial cells. In cells expressing wild-type PDGFRβ, PDGF induced disruption of GJC and phosphorylation of a gap junctional protein, connexin-43 (Cx43), which required activation of mitogen-activated protein kinase, although involvement of additional factors was also evident. In the F5 mutant lacking binding sites for phosphatidylinositol 3-kinase, GTPase-activating protein, SHP-2, and phospholipase Cγ1 (PLCγ1), PDGF induced mitogen-activated protein kinase, but failed to affect GJC or Cx43, indicating involvement of additional signals presumably initiated by one or more of the mutated binding sites. Examination of the single-site mutants revealed that PDGF effects were not mediated via a single signaling component. This was confirmed by the “add-back” mutants, which showed that restoration of either SHP-2 or PLCγ1 binding was sufficient to propagate the GJC inhibitory actions of PDGF. Further analysis showed that activation of PLCγ1 is involved in Cx43 phosphorylation, which surprisingly failed to correlate with GJC blockade. The results of our study demonstrate that PDGF-induced disruption of GJC can be mediated by multiple signaling pathways and requires participation of multiple components.
Footnotes
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↵* This work was supported by National Institutes of Heath Grant CA 57064.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: Molecular Medicine, Northwest Hospital, 120 Northgate Plaza, Suite 230, Seattle, WA 98125. Tel.: 206-368-3042; Fax: 206-368-3009; E-mail:mhossain{at}nwhsea.org.
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↵2 Hossain, M. Z., Jagdale, A. B., Ao, P., and Boynton, A. L., (1999) J. Cell. Physiol., in press.
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↵3 A. Golard, M. Z. Hossain, R. P. Huang, A. Kazlauskas, and A. L. Boynton, unpublished data.
- Abbreviations:
- PDGF
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platelet-derived growth factor
- PDGFR
-
platelet-derived growth factor receptor
- PI3K
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phosphatidylinositol 3-kinase
- GAP
-
GTPase-activating protein
- PLCγ
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phospholipase Cγ
- GJC
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gap junctional communication
- Cx43
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connexin-43
- PKC
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protein kinase C
- MAPK
-
mitogen-activated protein kinase
- PMAPK
-
phosphorylated mitogen-activated protein kinase
- PAGE
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polyacrylamide gel electrophoresis
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- Received October 23, 1998.
- Revision received January 6, 1999.
- The American Society for Biochemistry and Molecular Biology, Inc.
