Fig. 5: Interaction of UBR5 with CDC73/PAF1C.
From: UBR5 targets tumor suppressor CDC73 proteolytically to promote aggressive breast cancer
A Physical interaction of endogenous UBR5 with endogenous CDC73 was detected by co-IP with anti-UBR5 antibody in 4T1 cells which were treated with MG132 (10 μM) for 8 h before being collected for co-IP. B Physical interaction of exogenous UBR5 with exogenous CDC73 was monitored by co-IP with anti-Myc antibody in HEK293T cells transfected with empty vector, Myc-CDC73 or/and EGFP-UBR5 expressing plasmids for 40 h and then treated with MG132 (10 μM) for 8 h before being collected for co-IP. C The co-localization of endogenous UBR5 and CDC73 (i), UBR5 and PAF1 (ii), and CDC73 and PAF1 (iii) in 4T1 cells was monitored by immunofluorescence with a fluorescent microscope. The protein levels of UBR5 and the subunits of PAF1C including CDC73, PAF1, CTR9, LEO1, RTF1, and WDR61 were detected by western blotting in 4T1/GFP and 4T1/Ubr5−/− cells reconstituted with EGFP or UBR5 (D), 4T1/GFP shNC, 4T1/GFP shCdc73, 4T1/Ubr5−/− shNC, and 4T1/Ubr5−/− shCdc73 cells (E), shNC and UBR5 knockdown MDA-MB-231 cells (F), and shNC and CDC73 knockdown MDA-MB-231 cells (G).
