Fig. 5: CCNE1 exerts regulatory functions in TNBC through the ubiquitination modification of ANLN. | Cell Death Discovery

Fig. 5: CCNE1 exerts regulatory functions in TNBC through the ubiquitination modification of ANLN.

From: CCNE1 stabilizes ANLN by counteracting FZR1-mediated the ubiquitination modification to promotes triple negative breast cancer cell stemness and progression

Fig. 5

A The E3-specific ubiquitination sites of the substrate protein ANLN, specifically lysine residues 143, 371, and 405, were mutated to arginine. Flag-tagged plasmids K143R, K371R, and K405R were subsequently constructed, followed by the execution of ubiquitination detection experiments. B After addition of the 40 μM CHX, the stability of ANLN protein was tested in BT-549 and MDA-MB-231 cells with overexpressing either wild-type ANLN (ANLN-WT) or the ubiquitination site-mutated ANLN (ANLN-K143R). The proliferative ability of TNBC cells overexpressing either wild-type ANLN (ANLN-WT) or the ubiquitination site-mutated ANLN (ANLN-K143R) was evaluated by (C) MTT assay and (D) clonal formation assay, respectively. Plasmid overexpressing wild-type ANLN (ANLN-WT) or ANLN with ubiquitination site mutation (ANLN-K143R) was transfected into CCNE1 knockdown and control TNBC cells, and (E) the ball forming ability and (F) expression of stem cell markers were detected. n = 3, *P < 0.05, **P < 0.01, ***P < 0.001.

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