Fig. 2
TRAP increases growth and proliferation. Assessment of the impact of TRAP on anchorage-independent growth of TRAP-overexpressing TRAP3high cells (a-c, n = 4). One representative well with colonies is shown in (a). After 3 weeks of culture colony number (b) and colony size (c) were measured. Anchorage-dependent growth of TRAP-overexpressing clone TRAP3high is assessed by spectral quantification of adhered cells cultured for 24 h and 48 h in complete medium (d, n = 3). Measurement of proliferation is quantified based on the incorporation of EdU in the DNA during S-phase, immunocytochemistry and subsequent image analysis (e-h). Assessment of proliferation in TRAP-overexpressing (e, n = 6–14) and in the TRAP knockdown cells (f, n = 3–5) after 24 h culture in complete medium. Proliferation is also quantified in control cells (ctrl) and in TRAP-overexpressing TRAP3high cells after treatment with the small molecule TRAP inhibitor 5-PNA (200 μM) for 24 h (g, n = 4) and after starvation for 24 h and 48 h (h, n = 3). Statistical comparison was performed on biological replicates by t-test (Fig. 2b , f ), Mann-Whitney test (Fig. 2 c) or ANOVA test (Fig. 2d , e , g , h ). Groups are generally compared to their respective controls (ctrl or scr, DMSO treated) or indicated with brackets and significance is annotated with an asterisk (*). ns = non-significant. “n=” indicates the number of replicates