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. 2008 Aug 11;182(3):603–614. doi: 10.1083/jcb.200705102

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© 2008 Veracini et al.

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.jcb.org/misc/terms.shtml). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).

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Figure 5. — PAG-N inhibits SFK mitogenic signaling. (A) PDGFR–SFK complex formation is reduced in CEF of NIH 3T3 cells infected with PAG-N. Quiescent cells infected with the indicated retroviruses were stimulated with PDGF for 10 min. SFK association with caveolin and active PDGFR in CEF were evaluated by Western blotting of immunoprecipitated kinases with the indicated antibodies. Level of immunoprecipitated SFK is also shown. (B) Src mitogenic signaling is affected by PAG-N. SFK activity was measured by Western blotting with anti-pY416 antibody (left) or by in vitro kinase assay using denatured enolase (right) from immunoprecipitated kinases of the indicated cells stimulated or not with PDGF. Quantification (mean ± SD) of SFK activity was expressed as the ratio between active SFK and total SFK levels. (C) PAG-N inhibits phosphorylation of SFK mitogenic substrates. Levels of Stat3 and Shc phosphorylated on Tyr705 (left) or on Tyr239 and Tyr240 (right) are shown for cells infected with the indicated retroviruses. (D) PAG-N does not affect PDGF-induced Erk1-2 activation. Lysates of the indicated cells stimulated, or not, with PDGF for the indicated times were subjected to Western blotting with anti–phospho-Erk and anti-Erk–specific antibodies. Molecular masses are indicated in kilodaltons. *, P < 0.05 (using Student's t test).