PI3K mTORC1 pathway activation needs JAK task but perhaps not GP130 tyrosine phosphorylation. This coincided with reduced expression of angiopoietin 2, which is typically produced by endothelial cells during tumor order OSI-420 vascularization. But, as previously noted, RAD001 therapy avoided induction of hypoxia inducible factor 1?? at both the transcript and protein level. Expression of Vegfa, a transcriptional target for STAT3 well as Hif1??as, also remained unchanged following RAD001 treatment. GP130 activates mTORC1 via PI3K/AKT in a STAT1 independent manner and STAT3. To explore whether GP130 stimulates the mTORC1 pathway through PI3K activation, we supervised subcellular relocalization of the PI3K product PIP3, using a glutathione S transferase tagged pleckstrin homology domain from the phosphoinositides 1 receptor GRP1 like a probe. Compared with the diffuse background staining observed in unstimulated 293T cells, experience of the custom cytokine hyper IL 6 resulted in temporary accumulation of PIP3 in the plasma membrane within 3 minutes. We observed similar kinetics of PIP3 deposition after erythropoietin stimulation of cells transfected with a chimeric receptor comprising the extra-cellular Digestion domain of the Epo receptor fused to the intracellular domain of human wild-type GP130. By comparison, stimulation of the EpoR/ gp130F2 mutant, which encodes the human equivalent of the murine gp130Y757F substitution, triggered excessive and prolonged PIP3 accumulation at the plasma membrane, while untransfected 293T cells did not answer Epo. We interfered with endogenous STAT3 action in 293T cells using either STAT3 siRNA or perhaps a dominant negative variant of STAT3, to ensure that PI3K activation was STAT3 p53 ubiquitination independent. Successful STAT3 withdrawal was verified by immunoblot and by measuring the experience of a STAT3 responsive luciferase reporter construct. Significantly, STAT3 inhibition did not affect subcellular relocalization of PIP3 in cells harboring either the wild type or the EpoR/gp130F2 receptor. Collectively, these results claim that GP130 dependent PI3K/mTORC1 activation occurs independently of STAT3 and STAT1.