Figure 2
Characterization of mice carrying the ENU-induced C230X point mutation in the STEP gene. (a) The position of Cys230 within the KIM domain of STEP-61 and -46 isoforms. (b, c) The ENU-induced nonsense mutation at Cys230 within the KIM domain was detected by genomic PCR (b) and by DNA sequencing (c) from the genomic DNA of ENU-mutant mice. KIM, kinase interacting motif; PP, polyproline; PTP domains, protein tyrosine phosphatase; TM, transmembrane domain. (d, e) Western blotting (d) and immunohistochemical (e) analyses using anti-STEP antibody (anti-STEP, 23E5) showing the complete absence of functional STEP isoforms in the brain. (e, lower panel) Nissl-stained brain sections showing the cortex, dorsal and ventral striatum of wild-type and STEPC230X−/− mice. (f) Western blot analysis of STEP-dependent regulation of phospho-ERK levels in the striatum. STEP proteins of 61-kDa and 46-kDa isoforms were detected in the striatum of wild-type mice but not in STEPC230X−/− mice. Phosphorylation of ERK was increased in the striatum of STEPC230X−/− mice. Actin was used as a loading control. +/+, STEP wild-type mice; +/−, STEP heterozygous mice; −/−, STEPC230X−/− mice; cc, corpus callosum; cx, cortex; dSt, dorsal striatum; NAc, nucleus accumbens.
