Additional Supporting Information may be found in the online version of this article.

HEP_24329_sm_suppinfo.doc33KSupporting Information
HEP_24329_sm_suppinfofig1.tif1997KSupporting Fig. 1. Expression of constitutively active (ca) Stat3 versions and U-Stat3 in Ras-transformed hepatocytes. (A) Schematic representation of wtStat3 and Stat3 mutants. Stat3 is composed of the N-terminal domain, the coiled-coil, DNA-binding, linker, Src homology 2 (SH2) and transcriptional activation domain (TAD). (B) Overexpression of Stat3 as shown by immunoblotting using anti-Stat3 and anti-Actin antibodies. Actin was used as loading control.
HEP_24329_sm_suppinfofig2.tif1767KSupporting Fig. 2. Characteristics of Stat3Δhc/p19ARF-/- hepatocytes. (A) The loss of Stat3 and p19ARF in single and double knocked-out hepatocytes was analyzed by linear semi-quantitative RT-PCR (wt, wild type; ko, knock-out). (B) pY-Stat3 levels were analyzed by immunoblotting after stimulation with IL-6 (20 ng/ml). Actin was used as loading control. (C) Hepatocellular marker expression analyzed by linear semi-quantitative RT-PCR in primary hepatocytes (prim. hep.), parental MIM-1-4 hepatocytes (MIM-1-4) and the two single cell clones MIM-Stat3Δhc-1 and MIM-Stat3Δhc-2. The constitutive expression of RhoA is shown as loading control. One representative out of three experiments is shown. HNF, hepatocyte nuclear factor.
HEP_24329_sm_suppinfofig3.tif9550KSupporting Fig. 3. Knock-down of p14ARF in human Hep3B cells. (A) Real-time PCR analysis of p14ARF in p14ARF-deficient MCM-1, p14ARF-positive SW480 and Hep3B cells. (B) Levels of p14ARF and pY-Stat3 in Hep3B cells after expression of either scrambled control shRNA (sh-control) or shRNAs against p14ARF (sh-p14-1, sh-p14-2, sh-p14-3) as determined by immunoblot analysis. Actin was used as loading control. (C) Phase contrast and confocal immunofluorescence images after staining of Hep3B cells and those expressing sh-control or sh-p14 with anti-p14ARF antibody. Red, phalloidin; green, p14ARF; blue, DNA. Arrows indicate p14ARF localization in cell nucleoli. Error bars depict SD from at least three individual experiments.

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