Fig. 6: NEDD4L is the E3 ligase responsible for MLCK ubiquitination and degradation. | Communications Biology

Fig. 6: NEDD4L is the E3 ligase responsible for MLCK ubiquitination and degradation.

From: Elevated uric acid induces erectile dysfunction in rats by interacting with MLCK and inhibiting its ubiquitin-mediated degradation

Fig. 6: NEDD4L is the E3 ligase responsible for MLCK ubiquitination and degradation.

A MLCK ubiquitination in CCSMC regulated by HUA using Co-IP under MG132 treatment for 12 h. B Representative WB images of MLCK protein levels in CCSMC transfected with MLCK-WT or N803A plasmid for 48 h under HUA conditions or not. C The dot plot of MLCK protein levels in CCSMC analyzed by the two-tailed unpaired t-test from (M) Bars corresponds to Mean ± SD. *P < 0.05, **P < 0.01. n = 3 independent biological replicates per group. D MLCK ubiquitination in ARPE19 transfected with MLCK-WT or N803A plasmid for 48 h under HUA condition or not using Co-IP under MG132 treatment for 12 h. E Network view for retrieved E3 ligases for the query protein: MYLK (species: H.sapiens) using an in-silico ubiquitination prediction tool, UbiBrowser. F Representative WB images of NEDD4L protein levels from CCSMC treated with HUA or not. n = 3 independent biological replicates per group. G Representative WB images of NEDD4L and MLCK protein levels in CCSMC treated with HUA (or control) transfected with NEDD4L plasmid (or negative control plasmid) for 48 h. H The interaction between endogenous MLCK and NEDD4L in CCSMC for 48 h under MG132 treatment for 12 h by Co-IP. I The interaction between MLCK, NEDD4L and ubiquitin in ARPE19 with Flag-NEDD4L and either wild-type (WT) MLCK or the N803A mutant for 48 h under MG132 treatment for 12 h byCo-IP. HUA means 10% DMEM with an extra 450 μM uric acid for 48 h after 4 h starvation. MG132, 50 μM.

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