OBJECTIVE: To investigate whether human RhoA is modified by SUMO. METHODS: Overlap extension PCR and double digestion technique were used to construct the eukaryotic expression vector pcDNA3-3flag-RhoA, which was identified by sequencing. The plasmid was transfected into HEK293T cells and its expression was detected by Western blotting. Immunofluorescence assay was used to detect whether RhoA is co-localized with SUMO. Co-Immunoprecipitation was used to detect whether RhoA is modified by SUMO. RESULTS: The recombinant plasmid pcDNA3-3flag-RhoA was successfully constructed and verified. Western blotting showed that the recombinant plasmid pcDNA3-3flag-RhoA expressed abundant fusion protein in HEK293T cells. Immunofluorescence showed that RhoA was co-localized with SUMO2/3 but not with SUMO1. Co-immunoprecipitation verified that RhoA was modified by SUMO2/3 but not SUMO1. CONCLUSIONS: Human RhoA is modified by SUMO2/3 and probably participates in the regulation of axon regrowth after nervous system injury.
OBJECTIVE: To investigate whether humanRhoA is modified by SUMO. METHODS: Overlap extension PCR and double digestion technique were used to construct the eukaryotic expression vector pcDNA3-3flag-RhoA, which was identified by sequencing. The plasmid was transfected into HEK293T cells and its expression was detected by Western blotting. Immunofluorescence assay was used to detect whether RhoA is co-localized with SUMO. Co-Immunoprecipitation was used to detect whether RhoA is modified by SUMO. RESULTS: The recombinant plasmid pcDNA3-3flag-RhoA was successfully constructed and verified. Western blotting showed that the recombinant plasmid pcDNA3-3flag-RhoA expressed abundant fusion protein in HEK293T cells. Immunofluorescence showed that RhoA was co-localized with SUMO2/3 but not with SUMO1. Co-immunoprecipitation verified that RhoA was modified by SUMO2/3 but not SUMO1. CONCLUSIONS:HumanRhoA is modified by SUMO2/3 and probably participates in the regulation of axon regrowth after nervous system injury.
Authors: Sonia Castillo-Lluva; Michael H Tatham; Richard C Jones; Ellis G Jaffray; Ricky D Edmondson; Ronald T Hay; Angeliki Malliri Journal: Nat Cell Biol Date: 2010-10-10 Impact factor: 28.824
Authors: Helena Cimarosti; Emi Ashikaga; Nadia Jaafari; Laura Dearden; Philip Rubin; Kevin A Wilkinson; Jeremy M Henley Journal: J Cereb Blood Flow Metab Date: 2011-10-12 Impact factor: 6.200