Global Alternative Splicing Defects in Human Breast Cancer Cells

SIMPLE SUMMARY: Aberrant alternative splicing (AS) regulation plays a pivotal role in breast cancer development, progression, and resistance to therapeutical interventions. Indeed, cancer cells can adapt their own transcriptome by changing different AS programs, thus generating cancer-specific AS isoforms involved in every hallmark of cancer. Here, we investigated global AS errors occurring in human breast cancer cells by using RNA-mediated oligonucleotide annealing, selection, and ligation coupled with next-generation sequencing. Our results identified several dysregulated AS events potentially relevant for breast cancer-related biological processes and that provide a better comprehension of the molecular mechanisms that orchestrate the malignant transformation. ABSTRACT: Breast cancer is the most frequently occurred cancer type and the second cause of death in women worldwide. Alternative splicing (AS) is the process that generates more than one mRNA isoform from a single gene, and it plays a major role in expanding the human protein diversity. Aberrant AS contributes to breast cancer metastasis and resistance to chemotherapeutic interventions. Therefore, identifying cancer-specific isoforms is the prerequisite for therapeutic interventions intended to correct aberrantly expressed AS events. Here, we performed RNA-mediated oligonucleotide annealing, selection, and ligation coupled with next-generation sequencing (RASL-seq) in breast cancer cells, to identify global breast cancer-specific AS defects. By RT-PCR validation, we demonstrate the high accuracy of RASL-seq results. In addition, we analyzed identified AS events using the Cancer Genome Atlas (TCGA) database in a large number of non-pathological and breast tumor specimens and validated them in normal and breast cancer samples. Interestingly, aberrantly regulated AS cassette exons in cancer tissues do not encode for known functional domains but instead encode for amino acids constituting regions of intrinsically disordered protein portions characterized by high flexibility and prone to be subjected to post-translational modifications. Collectively, our results reveal novel AS errors occurring in human breast cancer, potentially affecting breast cancer-related biological processes.