CONICET | Buscador de Institutos y Recursos Humanos

Prostate Cancer (PCa) cells display abnormal expression of proteins resulting in an augmentedcapacity to resist chemotherapy and colonize distant organs. Most of the functional information ofthe cancer-associated genes relies in the proteome, an exceptionally complex biological systeminvolving several proteins. We have previously shown the anti-tumoral role of Heme oxygenase 1(HO-1) -the rate limiting enzyme in heme degradation- in this disease. We propose that HO-1 and itsinteractors reprogram PCa cells and modify the tumoral microenvironment, favoring a lessaggressive phenotype. Towards this end, we undertook a mass spectrometry-based proteomics study to build the HO-1interactome in PCa. In order to identify HO-1 molecular partners which might collaborate with itsbiological function, we constructed a recombinant GSTHO-1 protein. PCa cells were transientlytransfected with GSTHO-1 or the respective control and treated with the stressor agent H 2 O 2 .Immunoprecipitated protein complexes were subjected to LC-ESI MS/MS. The results rendered a setof HO-1 candidate partners that were subjected to protein interaction network and gene ontologyanalysis. Among the candidates, we identified STAT3, MLKN1, HSP27, HSP90, 14-3-3ζ/δ, testin andTrim28, with clinical relevance in PCa. We focused on 14-3-3ζ/δ, a protein that mediates signaltransduction by binding to phosphoserine-containing proteins. To assess and validate the clinicalsignificance of 14-3-3ζ/δ, we performed an integrative bioinformatics analysis using public databaserepositories (29 datasets; 4,401 samples). We identified high expression of 14-3-3ζ/δ to be stronglyassociated with poor prognosis across different PCa datasets. We also performed proteomicsanalysis on PCa and Benign Prostate Hyperplasia human tissue samples, identifying 14-3-3ζ/δ as oneof the proteins highly enriched in PCa.In parallel, we validated in PCa cells our proteomics approach by co-immunoprecipitation analysis,ascertaining HO-1 and 14-3-3ζ/δ interaction. Immunofluorescence assays provided evidence thatHO-1 and 14-3-3ζ/δ co-localize in the cell nuclei under oxidative stress conditions (Manders 1=.57).Further, bioinformatics analysis was performed to investigate the clinical correlation of these twoproteins in human PCa samples. When analyzing the relapse free survival of PCa patients that hadundergone radical prostatectomy ((GSE70769), n=206), those with high expression of YWHAZ,the gene encoding for 14-3-3ζ/δ, showed a higher risk of relapse (HR=3.94; p