Overex pression of AGK leads to activation of EGFR and promotes the proliferation and migration of prostate cancer cells, suggesting that AGK might act as a potent oncogene. Even so, the clinical significance of AGK and its associated signaling path ways stay unclear. Herein, we report that AGK is markedly overexpressed in esophageal squamous cell carcinoma and correlates with poorer illness zero cost survival and shorter overall sur vival in principal ESCC. Additionally, we discovered that AGK straight binds on the JH2 domain of JAK2 and blocks JH2 mediated inhibi tion of JAK2, resulting in constitutive activation of JAK2/STAT3 signaling and propagation with the CSC population in ESCC in vitro and in vivo. Much more importantly, AGK expression was proven to cor relate appreciably with STAT3 regulated signatures in ESCC, lung cancer, and breast cancer patient gene expression profiles. These findings uncover a mutation independent mechanism of JH2 inhibition that sustains activation of JAK2 in strong tumors.
Identification of AGK being a JH2 domain interacting protein that activates the JAK2/STAT3 pathway. To discover the mechanism by which solid selleckchem tumor cells override the autoinhibitory impact of JH2 to maintain activation of JAK2/STAT3 signaling, affinity purification and mass spectrometry had been utilized to determine JH2 interacting proteins in ECa109 ESCC cells. As proven in figure 1, A and B, and Supplemental figure 1A, AGK and seven other proteins were identified as potent JH2 interacting proteins. Importantly, reciprocal coimmunoprecipitation and Western blot assays more demonstrated that AGK could type a complicated with JAK2 and STAT3, suggesting that AGK might be involved while in the regulation of
JAK2/STAT3 signaling. Indeed, we identified that between these JH2 interacting partners, overexpres sion of AGK radically enhanced, whereas silencing of AGK decreased, STAT3 luciferase reporter activity and also the expres sion levels of phosphorylated JAK2 and phosphorylated STAT3.
Moreover, via analysis of AGK expression and STAT3 regulated gene signatures via gene set enrichment analysis kinase inhibitor SAR245409 in published ESCC patient expression profiles, we observed that AGK levels between typical and tumor tissues and within tumors were positively correlated with the STAT3 activated gene signatures and inversely correlated with all the STAT3 suppressed gene signatures. Taken together, these effects propose that AGK contributes towards the activation of JAK2/STAT3 signaling in ESCC. AGK interacts with JAK2 via binding immediately to its JH2 domain. To additional investigate the physical association among AGK along with the JAK2/STAT3 complex, the effect of AGK knockdown on the interaction between endogenous JAK2 and STAT3 was examined. As proven in figure 2A, AGK silencing didn’t greatly reduce the binding affinity of JAK2 for STAT3, indicating that AGK will not contribute to JAK2/STAT3 interaction.
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