We investigated this possibility for the growth pro moting role of Rac1. To do this we used PP1, a small molecule compound that has recently been shown in mammary epithelial cells and in PANC 1 cells to potently inhibit the kinase activity of TbRI/ALK5, selleck chemical to suppress TGF b1 induced phosphorylation Inhibitors,Modulators,Libraries of Smad2 and Smad3 and EMT. In addition, we have demon strated that PP1 dose dependently relieved the growth suppressive effect of TGF b1 in a Src unrelated fashion. To determine whether the autocrine TGF b growth inhibitory loop was subject to regulation by Rac1, we evaluated the effect of Rac1 depletion on pro liferative activity upon silenced autocrine TGF b signal ling under PP1 treatment. As shown in Figure 7A, PP1 increased the DNA synthesis in PANC 1 cells and, importantly, decreased the growth inhibitory Inhibitors,Modulators,Libraries effect of Rac1 siRNA when compared to vehicle controls.

We further reasoned that if TGF b autostimulation was permanently operating in cultures of PANC 1 cells then ectopic expression of a ca mutant of Rac1 should be able to stimulate p Smad2 even in the absence of exogenous TGF b1. This assumption was tested in transient cotransfection/immunoprecipita tion assays. Here, ca Rac1 was able to enhance the amount of Inhibitors,Modulators,Libraries p Smad2 over empty vector control samples in the absence of added TGF b1 and PP1, but was unable to do so in the presence of PP1. Together, these data strongly suggest that Rac1 modulates Smad signalling in response to both exogenous and autocrine TGF b signalling.

Discussion In this study we initially presented evidence that TGF b1 induced growth inhibition and cell migration in PDAC cells were differentially and selectively controlled by Smad3 and Smad2, respectively. Knockdown of Smad3 but not Smad2 Inhibitors,Modulators,Libraries relieved TGF b1 induced growth inhibition, indicating that this response was Smad3 dependent, an observation made previously in various other cell types including PANC 1 cells. In contrast, knockdown of Smad2 decreased the TGF b1 driven motility of PDAC cells revealing cell migration to be a Smad2 specific response. This is in line with the demonstration of a crucial role of Smad2 in regulating keratinocyte migra tion Inhibitors,Modulators,Libraries during wound healing. We went on to describe first time observations, namely that the effects of Smad2 depletion on TGF b1 mediated growth inhibition and cell migration were largely mimicked by inhibition of Rac1 expression or activity, or pharmaco logic inhibition, together suggesting a functional link between both proteins. We subsequently confirmed this assumption by showing that Rac1 inhibi tion abrogated TGF b1 induced Smad2 specific C term inal phosphorylation and transcriptional activity but increased TGF b1 mediated selleck inhibitor p21WAF1 expression.