Abstract 1478: Role of Stat3 Ser727 phosphorylation in mouse hepatocarcinogenesis

Abstract

Abstract Inappropriate activation of signal transducer and activator of transcription 3 (Stat3) occurs with extreme frequency in a wide variety of cancers including HCC, and constitutive Stat3 activation in hepatocytes was demonstrated to accelerate hepatocarcinogenesis, while repression of Stat3 activation suppresses hepatocarcinogenesis. Diverse factors such as cytokines, growth factors and oncoproteins can activate Stat3, where phosphorylation of the conserved tyrosine residue at the C-terminus (Y705) results in the Stat3 homodimer or the heterodimer with Stat1, and phosphorylation of the serine residue on the transactivation domain (S727) contributes to its maximal transcriptional activity. The activated Stat dimers bind to specific DNA-response elements in the promoter of target genes and thereby induce unique gene expression programs cooperating with other transcription factors and cofactors. We investigated Stat3 activation in mouse hepatocarcinogenesis induced by neonatal treatment with diethylnitrosamine (DEN) and found that Stat3 phosphorylation occurred by two steps, where S727 phosphorylation was seen in the preneoplastic stage, whereas Y705 phosphorylation was further associated with malignant progression. In mouse HCC cells, serum treatment induced Ser727 phosphorylation via MAPK activation, associating with cell proliferation and Stat3 nuclear translocation, whereas IL-6 induced Tyr705 phosphorylation via JAK activation without stimulating cell proliferation and the Stat3 nuclear translation. On the other hand, the Ser727 phosphorylation dead Stat3 (S727A), in which the serine residue was replaced by alanine, reduced the cell proliferation and Stat3 nuclear translocation, while the Ser727-phosphorylation mimic Stat3 (S727D), in which the serine residue was replaced by aspartic acid, enhanced the cell proliferation and Stat3 nuclear translocation. To further investigate the role of Stat3 Ser727 phosphorylation, the Stat3 knockdown HCC cell lines were established by introducing the Stat3 siRNA into the mouse HCC cells. These cell lines showed retarded growth and less attachment to the substrate forming spheroidal aggregates, whereas the parent cells showed rapid proliferation and attached to the substrate with extended morphology. cDNA array analysis revealed that the Stat3 knockdown cells showed variety of gene expression changes relating to cell survival and growth, and RT-PCR demonstrated that the expression of EGFR, FGFR and E-cadherin was markedly reduced in the Stat3 knockdown cells. Our findings indicate that the Ser727-phosphorylated Stat3 plays an important role from the early stage in mouse hepatocarcinogenesis, presumably by controlling cellular growth and survival. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1478.