Fig. 3 | Signal Transduction and Targeted Therapy

Fig. 3

From: Conversion of Ku80 K568 crotonylation to SUMOylation facilitates DNA non-homologous end joining and cancer radioresistance

Fig. 3

HDAC8 and PCAF coregulate the crotonylation of Ku80. a Functional analysis of the effects of TSA (1 μM, 10 h) and NAM (10 mM, 10 h) treatment on the decrotonylation of Ku80. b IP and western blotting were performed to identify the key regulating decrotonylase for Ku80 with the indicated antibodies. c HEK-293T cells expressing exogenous Flag-Ku80 were transfected with the indicated siRNAs and plasmids. Later, Ku80 crotonylation degrees were evaluated on IP products of the Flag antibody by immunoblotting. d Treatment with the HDAC8-specific inhibitor PCI-34051 (10 μM, 6 h) increased the crotonylation level of Ku80 in HEK-293T cells. e HDAC8 disrupted Ku80 crotonylation in a dose-dependent manner in HeLa cells. f PCAF increased Ku80 crotonylation in a dose-dependent manner in HeLa cells. g Impaired Ku80 crotonylation by PCAF siRNA is reversed by transfection of PCAF siRNA-resistant plasmids in HEK-293T cells. h PCAF crotonylates and HDAC8 decrotonylates Ku80 in vitro cell-free reaction assay. i Co-IP with DNase I treatment shows increased interaction between Ku80 and the decrotonylase HDAC8 after radiation exposure. Three independent replications of WB experiments. β-actin was assayed to ensure equivalent loading and transfer

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