Fig. 5

YB-1 phosphorylation at S102 confers CDK4/6i resistance. A. The plasmids YB-1-S102A and YB-1-S102E were used to simulate dephosphorylation and hyperphosphorylation, respectively, and two plasmids were transfected separately into MCF7 cells. The PARP1 level was almost invisible in the YB-1-S102A group and was significantly increased in the YB-1-S102E group compared to the wild-type YB-1 group, as the Western blot data show. B. The fluctuation of CDK4/6i sensitivity was confirmed by MTT assay when cells were transfected with YB-1-S102A and YB-1-S102E plasmids. C. The two mutant YB-1 plasmids were transfected into MCF7-shYB-1 cells; the Western blot data showed that the PARP1 level in the shYB-1 group was lower than that in the control group. D–E. The change in CDK4/6i sensitivity for mutant YB-1 plasmid transfection was confirmed by MTT assay. F. The inhibition efficiency of cell viability for the YB-1-S102A-MCF7-shYB-1 group reached approximately 13% when transfected with the mutant YB-1 plasmid when selecting the cell viability of abemaciclib at 10 μM, G–H. The YB-1 nuclear translocation was increased under abemaciclib treatment, as determined by an immunofluorescence assay. I. We constructed an upstream 2000 bp fragment in the promoter of PARP1 into a luciferin vector plasmid for a double fluorescein reporter gene assay. The effect of mutant plasmid transfection on PARP1 transcription was determined by the change in fluorescence intensity under transfection with the YB-1-WT/S102A/S102E plasmids. The results presented have been repeated in 3 biological replicates. Data, means ± SEMs, *, P < 0.05, **, P < 0.01, ***, P < 0.001