Fig. 3: SPOP shortens ASCT2 protein half-life and promotes ASCT2 ubiquitylation. | Nature Communications

Fig. 3: SPOP shortens ASCT2 protein half-life and promotes ASCT2 ubiquitylation.

From: Neddylation inhibition induces glutamine uptake and metabolism by targeting CRL3SPOP E3 ligase in cancer cells

Fig. 3: SPOP shortens ASCT2 protein half-life and promotes ASCT2 ubiquitylation.

a MDA-MB-231 cells were transfected with FLAG-SPOP and various mutants for 48 h, followed by immunoblotting. b MDA-MB-231 cells were transfected siRNAs targeting SPOP for 48 h, followed by immunoblotting. c, d MDA-MB-231 cells were transfected with siRNA targeting SPOP (si-SPOP#1) or scramble control siRNA for 48 h, then incubated with cycloheximide (CHX). Cells were harvested at indicated time points for immunoblotting (c). The band density of ASCT2 was quantified using ImageJ software and normalized to α-tubulin (mean ± SD, n = 3) (d). e–h HEK293 cells were transfected with indicated SPOP plasmids (e), indicated ASCT2 plasmids (f), or with indicated ubiquitin mutants (h) lysed under denaturing conditions, followed by Ni-beads pulldown, and immunoblotting for ASCT2 (top panels). The bottom panels are direct immunoblotting using whole cell extracts. Source data are provided as a Source Data file.

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