Figure 1
From: FBXW7 regulates DISC1 stability via the ubiquitin-proteosome system
DISC1 turnover is regulated by the ubiquitin-proteosome system. (a) HEK293 cells were treated with cycloheximide (50 μg ml–1) for indicated times before cellular lysates were isolated and probed for DISC1 and another known FBXW7-substrate Myc. Levels of DISC1 protein were evaluated by densitometry (bar-chart, n=3) (b) HEK293 cells were treated with indicated concentrations of MG132 (10–60 μM) for 1 h before cellular lysates were isolated and probed for DISC1. (c) HEK293 cells were transfected with HA-DISC1 and treated with MG132 (10 μM) for indicated times before cellular lysates were prepared (upper panels) and blotted for HA and Tubulin. HA-DISC1 levels were evaluated using densitometry (bar-chart, n=3). ‘Nt’ indicates no treatment control. The HA-tag was immunoprecipitated and blotted for HA and ubiquitin (lower panels). (d) HEK293 cells were transfected with HA-DISC1 and treated with MG132 (10 μM) for 1 h before cellular lysates were prepared and HA-tag immunoprecipitated and blotted for HA (lower panel) and ubiquitin chains linked specifically via lysine 48 (upper panel). (e) A siRNA library against human F-Box proteins was transfected individually into HEK293 cells, before cellular lysates were recovered and blotted for endogenous DISC1 and GAPDH. DISC1 levels were evaluated by densitometry (bar-chart, n=1).
