Through Genome-Wide Association Studies (GWAS), IRX4 has been identified to be associated with prostate cancer (PCa) risk at chromosome 5p15. Interestingly, our group discovered a long non-coding RNA (IRX4lncRNA) in the anti-sense strand of IRX4 and a novel Insertion-Deletion Polymorphism (INDEL) regulating androgen mediated IRX4 and IRX4lncRNA expression. The potential of lncRNAs to encode regulatory small peptides with indispensable regulatory functions, called micropeptides (miPEPs) has recently been discovered. Our preliminary in silico analysis indicated that IRX4lncRNA can potentially encode for the miPEPs. We aim to determine the role of IRX4lncRNA/miPEP and validate its efficacy as PCa biomarker.
Sequential Window acquisition of all theoretical fragment ion spectra mass spectrometry (SWATH-MS) and Multiple Reaction Monitoring (MRM) were performed to identify and validate micropeptides coded by IRX4lncRNA in prostate cancer cells. To determine the functional role of IRX4lncRNA/micropeptides in prostate cancer, in-vitro and in-vivo assays assays are being performed using overexpression and knockdown models. Further, identification of these peptides in clinical patient serum/tissue samples will determine its expression and utility as a biomarker.
SWATH-MS and MRM analysis validated the expression of two peptides encoded by IRX4lncRNA in PCa cells (PC3, VCaP and LnCaP). Overexpression models of the ORF encoding these peptides resulted in increased proliferation of prostate cancer cells. Additionally, mass spectrometry analysis of the overexpression and knockdown models of the ORF identified changes in the pathways asscoaued with ubiquitin mediated proteolysis, enrichment in Rap1signalling pathway and changes in fatty acid metabolism. These pathways will be analysed in detail to determine specific function of the IRX4lncRNA/miPEP. Moreover, these peptides are being tested in clinical patient serum/tissue samples to determine its expression and utility as a biomarker
The findings from this study will elucidate the role of IRX4lncRNA/miPEP and their biological impact in PCa.