Hitting the breaks on the migratory capacity of tumoral cells: targeting key regulators of actin dynamics in prostate cancer

GERALDINE GUERON, PÍA VALACCO, BELÉN ELGUERO, FEDERICO COLUCCIO-LESKOW, ALEJANDRA PAEZ, ADRIANA DE SIERVI, ELBA VAZQUEZ; ALEJANDRA PAEZ, ; EMILIANO ORTIZ, ; CARLA PALLAVICINI, ; JIMENA GIUDICE,; PIA VALACCO,; MARIA BINAGHI,; MARCELO MARTI, ; LUCIANA BRUNO, ; VALERIA LEVI,; MARCELO SALIERNO ; JAVIER COTIGNOLA, ; NORA NAVONE,; ELBA VAZQUEZ.

Washington

Encuentro; 22nd Annual PCF Scientific Retreat; 2015

Prostate Cancer Foundation

Background: Metastatic cancer involves the movement of cancer cells from the primary site to distant homing organs. The migratory capacity of cells, requires a dynamic remodeling of the cell cytoskeleton. While actin filaments, microtubules and intermediate filaments, coordinate their functions in normal cells, tumoral cells display abnormal expression of cytoskeletal proteins resulting in an augmented capacity to resist chemotherapy and colonize other organs. The induction of heme oxygenase-1 (HO-1), the rate-limiting enzyme in heme degradation, has a strong anti-tumoral effect in prostate cancer (PCa) and regulates the adhesive properties of PCa cells. Here we have extended our studies to assess the role of HO-1 on cell morphology and dynamics of the actin cytoskeleton at filopodia in PCa cells.Methods: Motility changes were assessed on fiber-like 1D and 2D migration scenarios in cells after hemin treatmemt (HO-1 pharmacological inducer). Actin cytoskeleton visualization was performed on PC3 cells exposed to hemin or siRNAHO-1, fixed and stained with phalloidin-rhodamin. Filopodia from neighboring cells were observed by confocal microscopy and analysed with a custom made algorithm to count contacts. Isolated proteins from PC3 cells transfected with FLAG-tagged HO-1 were immunoprecipitated and subjected to LC/ESI-MSMS analysis. Gene ontology (GO) analyses of HO-1 interacting proteins were performed using DAVID.For the bioinformatics screening of HO-1 interacting proteins, we used Oncomine and the public repository Gene Expression Omnibus (GEO) from the National Center for Biotechnology Information to browse for gene expression microarrays data. Raw expression data was processed using the limma R package from Bioconductor. Results: A reduced frequency in migration events and in migration speed was detected under hemin exposure. Nuclear shape was also altered. A significant higher proportion of filopodia-like protrusions among neighboring cells and cellular contacts were observed under HO-1 induction. HO-1 silencing reversed these effects. The proteomics analysis of HO-1 interacting proteins yielded a 17% of cytoskeletal-associated proteins regulating actin filament dynamics. Our bioinformatics screening of these proteins revealed that most of the genes studied lie within the 10 or less percent of the most consistently low-expressed genes in prostate adenocarcinoma compared to normal tissue.Conclusion: Our experimental findings demonstrate that HO-1 modulation in PCa induces the remodeling of the actin filament architecture at filopodia, alters the migratory patterns and cellular morphology, showcasing the relevance of the cytoskeleton and its interacting partners as potential therapeutic targets against aggressive and metastatic disease.