Figure 3

CARP-1 interacts with TAZ (A, B), TAZ expression interferes with CARP-1-dependent apoptosis (C), and CARP-1 and TAZ co-localize in cytoplasmic/perinuclear region (D). For CARP-1 binding with TAZ, Breast cancer cells (panel A) or HCT-116 colon cancer cells expressing myc-His-tagged wild-type CARP-1 (panel B) were utilized. In panel B, the cells were transfected with plasmids encoding wild-type or mutant TAZ proteins as indicated, prior to generating cell lysates. The cell lysates were then utilized for immunoprecipitation and western blotting using noted antibodies essentially as in figure 2C. In panel C, cells were transfected with plasmids encoding vector or wild-type TAZ followed by their transduction with indicated retroviruses expressing vector or wild-type CARP-1 as detailed in methods. The apoptosis levels were assessed by determining the acridine orange-positive cells as described before [4]. Columns in histogram represent data from 3 independent experiments; bars, SE. In panel D, the cells were transfected with a combination of plasmids encoding myc-His-tagged wild-type CARP-1 and flag-tagged wild-type TAZ proteins, followed by their fluorescent labeling and photography as described in methods.