Figure 4.

TTC5 prevents apoptosis of THP1 cells through regulation of BCL2 expression. THP1 cells were infected with lentiviruses expressing shRNAs targeting TTC5 for KD, or a non-targeting control (NTC) shRNA, with puromycin drug resistance as the selectable marker. Bar chart shows (a) mean±S.E.M. transcript expression levels of the indicated genes in KD cells relative to control cells at the indicated time points following lentiviral infection (n=2). (b) Representative western blots show expression of the indicated proteins, 7 days after lentiviral infection. Bar charts show (c) mean±S.E.M. TTC5 or (d) BCL2 transcript levels in control (EV, empty vector) or BCL2 overexpressing TTC5 KD THP1 cells relative to control cells, 72 h after lentiviral infection (n=3). Bar charts show (e) mean±S.E.M. percentage of apoptotic cells, as determined by annexin V binding, 7 days after lentiviral infection (n=3) and (f) mean±S.E.M. colony-forming cell (CFC) frequencies of drug-resistant cells enumerated following 10 days in semisolid culture (n=3). * Indicates P⩽0.05 (Mann–Whitney U-test) for comparison of KD versus control conditions, or for the indicated comparisons. NS=not significant. (g) Bar chart shows mean±S.E.M. percentage of TTC5 KD cells in G1 or S phase of cell cycle relative to that observed in control cells (for BCL2 over expressing and control THP1 lines) 96 h after initiation of KD (n=3). * Indicates P⩽0.05 for comparison of KD versus control conditions using one-way ANOVA and Fisher's least significant difference post hoc test