Fig. 4: Knockout of Bbc3 gene leads to increased resistance to TKI in SCLL. | Cell Death & Disease

Fig. 4: Knockout of Bbc3 gene leads to increased resistance to TKI in SCLL.

From: Downregulation of PUMA underlies resistance to FGFR1 inhibitors in the stem cell leukemia/lymphoma syndrome

Fig. 4

Schematic diagram of the Puma encoding gene (Bbc3) structure, showing the CRISPR target sites and ___location of primer pairs to demonstrate deletion of the gene (A). Western blot analysis of 24 individual clones identified four (#7, #10, #13, and #14) with complete depletion of Puma protein (B) indicating successful knockout. PCR analysis of the Bbc3 locus confirms the deletion in all four cases (C). Inhibition of growth by BGJ398, as a percentage of DMSO treated controls (D), shows reduced sensitivity in two knockout clones (#10 and #14). IC50 for parental cells = 13.6 nM, for KO#10 = 36.3 nM and for KO#14 = 52.5 nM. Flow cytometry analysis of 7AAD/Annexin V expression levels (E) shows reduced induction of apoptosis in the knockout cells. Western blot analysis of downstream signaling in the knockout clones shows loss of Parp cleavage in the KO cells (see text). Student’s t-test was used for statistical comparisons between groups. **p < 0.01; ***p < 0.001.

Back to article page