MEL-18 controls ESR1 transcription of the suppressing brand new SUMOylation of the ESR1 transcription activities p53 and you will SP1
(A) Cell lysates treated with 20 mM N-ethylmaleimide (NEM) were subjected to immunoblotting. The amount of SUMOylated protein was quantified by measuring the ratio of SUMOylated protein/total protein. (B) Venn diagram showing the relationship between the microarray results for MCF-7 cells expressing MEL-18 shRNA (shMEL) and those for MCF-7 cells treated with RITA (GSE13291) ( 36 ). (C) MCF-7 cells expressing MEL-18 siRNA (siMEL) were cotransfected with WT or SUMOylation-deficient mutant constructs of p53 or SP1 and with ESR1 pro-Luciferase and were subjected to a luciferase reporter assay. The data are presented as the mean ± SD (n = 3). *P < 0.05 vs. siCon/Con; † P < 0.05 siMEL/Con (2-tailed Student's t test). (D) ChIP-qPCR analysis showing the amount of ESR1 transcription factor that was recruited to the ESR1 promoter in the indicated cells. The data are presented as the mean ± SD (n = 3). *P < 0.05 vs. shCon (2-tailed Student's t test). (E) The effect of ginkgolic acid on the expression of ER-? in the MEL-18–silenced cells. Cells were treated with 100 mM ginkgolic acid for 24 hours and subjected to immunoblotting. Parallel samples examined on separate gels are shown. The data were quantified by measuring the immunoblot band densities from three independent experiments (mean ± SD). *P < 0.05 vs. shCon; † P < 0.05 vs. shMEL (2-tailed Student's t test). All data shown are representative of three independent experiments.
For the MEL-18–silenced MCF-7 muscle, the level of the 39-kDa SUMO-1–conjugating particular this new SUMO E2 enzyme UBC9 was graced, whereas the level of the 18-kDa free form out of UBC9 is actually less (Supplemental Figure 13A)
MEL-18 advances deSUMOylation by the suppressing the brand new ubiquitin-proteasome destruction of sentrin-certain protease 1. To help select brand new process by which MEL-18 controls SUMOylation, the effect from MEL-18 to the phrase regarding SUMO-related circumstances is actually checked-out.Leggi tutto »MEL-18 controls ESR1 transcription of the suppressing brand new SUMOylation of the ESR1 transcription activities p53 and you will SP1