Discovery of differentially expressed novel miRNAs in breast normal cells and their putative targets

MicroRNAs (miRNAs) play critical role in normal breast development and their altered expression may lead to breast cancer. Identification of new miRNAs allows us to understand the normal physiological process and associated disease pathophysiology. In the present study we identify the novel miRNAs in withaferin A treated breast normal cells (MCF-10A) using small RNA sequencing. The pathophysiological potential of the identified miRNAs was checked by studying their expression pattern in MDA-MB-231 and MCF-7 breast cancer cells using qRT-PCR technique. The secondary/tertiary structure of the identified miRNAs, target gene enrichment in Gene Ontology terms and KEGG pathway, miRNA-mRNA interaction of the sorted target genes, miRNA-mRNA/miRNA-argonaute protein/miRNA-mRNA-argonaute protein interaction and stability, were studied using bioinformatics tools/software, and molecular dynamics simulations. Hsa-miR-N88585 and hsa-miR-N461089 were identified and validated as novel miRNAs in normal breast cells. Up-expression of identified miRNAs in MDA-MB-231 and MCF-7 cells indicates their oncogenic nature. Identified target genes were enriched in classical signaling pathways (AMPK and Ras) and important GO terms. PLXDC2, BHLHE40, ARMC8, and PECAM1, CDC27, KCNK3 genes were sorted as putative targets for hsa-miR-N88585 and hsa-miR-N461089, respectively. MD simulation revealed stable hsa-miR-N88585/hsa-miR-N461089-AGO protein complex formation which indicates their further processing. In conclusion, the study identifies hsa-miR-N88585 and hsa-miR-N461089 as novel miRNAs in breast normal cells which are significantly inversely expressed in breast cancer cells. Further experiments are required to study the role of identified novel miRNAs in normal breast development and pathophysiology of breast cancer.

Automated summary

In this study, novel miRNAs were identified in breast normal cells treated with Withaferin A and their expression patterns were studied in breast cancer cells. Bioinformatics tools and molecular dynamics simulations were used to analyze the secondary/tertiary structure of the identified miRNAs, target gene enrichment, miRNA-mRNA interaction, and stability. Hsa-miR-N88585 and hsa-miR-N461089 were identified as novel miRNAs in breast normal cells, showing overexpression in breast cancer cells and interaction with genes involved in important signaling pathways and GO terms. Molecular dynamics simulations indicated stable complex formation between hsa-miR-N88585/hsa-miR-N461089 and AGO protein. These findings suggest the potential role of these novel miRNAs in breast cancer pathophysiology.