Figure 4.

Generation and functional characterization of hepatocyte-specific gp130^ΔSTAT/LoxP and gp130^Y757F/LoxP mice. (A) alfpCre gp130^LoxP/LoxP mice were generated by breeding alfpCre mice with animals expressing LoxP-flanked gp130 alleles (25). Hepatocyte-specific gp130^ΔSTAT/LoxP animals (alfpCre gp130^ΔSTAT/LoxP) were generated by crossing alfpCre gp130^LoxP/LoxP with gp130^{ΔSTAT/ΔSTAT} mice, which express a truncated gp130 knockin allele encoding a truncated gp130 protein that lacks the domains mediating STAT1 and STAT3 activation (26). Hepatocyte-specific gp130^Y757F/LoxP animals (alfpCre gp130^Y757F/LoxP) were bred by crossing alfpCre gp130^LoxP/LoxP with gp130^Y757F/Y757F mice, which express a gp130 allele encoding a phenylalanine substitution of the Y757 residue (32), thereby rendering gp130 incapable of recruiting SHP2 and activating the RAS-MAPK pathway. Animals were genotyped by PCR analysis for alfpCre, gp130^LoxP, and gp130^Y757F alleles. (B and E) The functional characterization of hepatocyte-specific gp130 mutant mice. Gp130 downstream-signaling pathways were analyzed by monitoring of phosphorylated STAT3 and phosphorylated ERK2 (p42) expression in whole cell extracts of primary hepatocytes isolated from wild-type (B), alfpCre gp130^LoxP/LoxP (C), alfpCre gp130^Y757F/LoxP (D), or alfp Cre gp130^ΔSTAT/LoxP mice (E). Activation of phosphorylated STAT1 was not detected in any of the 4 mouse strains.