Fig. 1: p14ARF is modified by SUMO in vitro and in vivo.
From: SUMOylation of the lysine-less tumor suppressor p14ARF counters ubiquitylation-dependent degradation
A In vitro-transcribed/translated 35S-methionine-labeled p14ARF protein is modified by SUMO2 in vitro (upper panel). In vitro SUMOylated p14ARF protein was then subjected to an in vitro deSUMOylation assay in the presence of SENP1 (lower panel). B Modification of transfected p14ARF (upper left panel), p14ARF-HA (upper right panel), p14ARF-myc (lower left panel), and p14ARF-GFP or GFP (lower right panel) proteins by SUMO2 in HEK-293 cells transfected with His6-SUMO2 and UBC9. C Modification of transfected p14ARF-GFP protein by SUMO2 in HEK-293 cells co-transfected with His6-SUMO2, UBC9 and pcDNA or His6-SUMO2, UBC9 and Flag-SENP1. D SUMOylation of endogenous p14ARF in PC3 cells transfected with non-deconjugatable His6-SUMO2-Q90P. E Detection of endogenously SUMOylated p14ARF using SUBES. F Detection of endogenously SUMOylated p14ARF using SUMO2 immunoprecipitation. Arrow indicates unmodified p14ARF protein and both arrowheads and brackets indicate SUMOylated p14ARF. WCE whole cell extracts.
