SUMOylation Blocks the Ubiquitin-Mediated Degradation of the Nephronophthisis Gene Product Glis2/NPHP7
Fig 6
Lysine 195 is required for Glis2 stability.
(A) HEK 293T cells, transfected with Glis2 wild type (WT) or Glis2.K195R along with V5-tagged GFP, were treated with MG132 (12 μM) for 2 hours to inhibit proteasomal degradation. DMSO was used as a vehicle control. The cell lysates were resolved on SDS-PAGE, and the protein levels of Glis2 were detected using anti-V5 antibody. (B) Bars represent the quantification of Glis2.WT and Glis2.K195R protein levels normalised to V5-tagged GFP levels from three independent experiments (n = 3). (C) In vivo ubiquitylation assays demonstrate the increased ubiquitylation of F.Glis2.K195R in comparison to Glis2 wild type (WT). (D) In vivo ubiquitylation assays were performed with the plasmids as indicated to compare the SUMO-3 mediated inhibition of ubiquitylation of F.Glis2 wild type (WT) and F.Glis2.K195R.