RHBDD2 AND WWOX PROTEIN INTERACTION AS MODULATOR MECHANISM OF PROLIFERATION AND DIFFERENTIATION OF NORMAL AND NEOPLASTIC BREAST CELLS

FERRETI VIRGINIA; CANZONERI R; LACUNZA E; KORDON E; ALDAZ M; ABBA MC

Congreso; LXI Reunión Anual SAIC; 2016

The Rhomboid gene family constitutes a heterogeneous groupof proteases / pseudoproteases that are conserved throughoutevolution, involved in different biological processes such asEGFR, TNF, ERAD and UPR signaling pathways among others.We previously demonstrated that RHBDD2 is a cancer relatedgene amplified and overexpressed in advanced breast cancer.On the other and, The WW domain containing protein WWOXhas been postulated as a tumor suppressor in breast and othercancers. WWOX modulates the TGFβ signaling pathway viadirect WW-domain mediated binding and potential cytoplasmicsequestration of SMAD3 protein. The aim of this study was tocharacterize the RHBDD2-WWOX protein interaction and theirbiological relevance over the TGFβ signaling pathway in normaland breast cancer cells. Functional proteomics analysis using TAP/MS allowed us to identify a RHBDD2 ? WWOX protein-proteininteraction in HEK293 cancer cells. This result was validated innormal (HC11) and breast cancer cell lines (MCF7, T47D, MDAMB231)by Co-immunoprecipitation (Co-IP) and western-blot. Inaddition, co-localization analysis corroborate the previous finding,demonstrating that such protein-protein interaction occurs at theGolgi complex. HC11 mouse mammary cell line was employedto evaluate Rhbdd2 and Wwox expression at mRNA and proteinlevels by RT-PCR and Co-IP/WB methods respectively. Wedetected a high Rhbdd2 expression at the proliferative stage ofthe HC11 cells with a dramatic decrease at the competent anddifferentiated stages, while Wwox expression was expressed atthe differentiated stage. Oligo-microarray based analysis of MDMBA231RHBDD2 silenced cells (siRNA) suggest that RHBDD2could be involved in the modulation of TGFβ signalling pathwayamong other bioprocess via sequestration of WWOX protein atthe Golgi complex. This study show that the RHBDD2-WWOXprotein interaction is associated with the modulation of the mammary proliferation and differentiation process, suggesting that thisinteraction might play a relevant role in the normal developmentand neoplastic progression.