Conclusion
We propose that DNA demethylation followed by increased expression of key genes may be one of the factors responsible for the progression and maintenance of the resistant cell phenotype that includes exosome-induced resistance.
Methods
The cell viability was assessed by the MTT assay, exosomes were purified by successive centrifugations, immunoblotting was used to evaluate protein expression, AP-1 activity was analyzed using reporter assay.
Results
In experiments on the MCF-7 cell line (breast cancer) and the MCF-7/Rap subline that is resistant to rapamycin, the capability of resistant cell exosomes to trigger a similar rapamycin resistance in the parent MCF-7 cells was demonstrated. Exosome-induced resistance reproduces the changes revealed in MCF-7/Rap resistant cells, including the activation of ERK/AP-1 signaling, and it remains for a long time, for at least several months, after exosome withdrawal. We have shown that both the MCF-7 subline resistant to rapamycin and cells having exosome-triggered resistance demonstrate a stable decrease in the expression of DNMT3A, the key enzyme responsible for DNA methylation. Knockdown of DNMT3A in MCF-7 cells by siRNA leads to partial cell resistance to rapamycin; thus, the DNMT3A suppression is regarded as one of the necessary elements for the development of acquired rapamycin resistance.