ROR2 inhibits proliferation of melanoma cells by delaying cell cycle progression.

CASTRO MV; BARBERO G; VILLANUEVA MB; LOPEZ BERGAMI P

Mar del Plata

Congreso; Reunión de la Sociedad Argentina de Investigación Clínica; 2018

Sociedad Argentina de Investigación Clínica

The Wnt5a signaling pathway playsan important role in several types of cancer. The aim of this project is todetermine the role of ROR2, a tyrosine kinase receptor of Wnt5a, in melanoma. Wehave shown that ROR2 overexpression decreased proliferation by negativeregulating Cyclin D1 and Cdk4 levels and by increasing p21 levels. Here westudied how ROR2 regulates cell cycle progression. We found that ROR2 overexpressiondecreased Retinoblastoma phosphorylation and its cytoplasmic location, while ROR2silencing increased it. ROR2 overexpression decreased G2/M population (8.9% vs24.5% in control cells, (p <0,01)) at the expense of cells in both G1 and S.Silencing of ROR2 also markedly alter the cell cycle, most noticeably bydecreasing the number of S phase cells from 40.6% to 21.1% (p). Then we performed time course experiments on G1-phase synchronized cellsto evaluate the transition to G2/M phase. Seventeen (17,2% ± 4,8%) percent of A375-empty cellstransitioned to G2/M by 18h vs. 0.6% ± 0.4% of A375-ROR2 (p   ). These cells required an additional 6h to reachsimilar values (xxxx% ± xxx% at 24h). In the case of M2 cells, none(M2-scramble) and 9.6% ± 2.7% cells (M2-shROR2) transitioned to G2/M phaseafter 16h. It took 24h for the M2-scramble cells to reach similar values (xxxx%± xxx%). We then analyzed changes in cell cycle markerson cells treated as in the experiment above. Cyclin A levels rapidly peaks at12h after G1 release and then down curved until almost disappear by 24h. Incontrast, in A375-ROR2 Cyclin A slowly increased until reaching its peak at24h. Meanwhile, in M2 cells Cyclin A peaks at 28h and 16h in scramble and ROR2shRNA cells, respectively. Similar conclusions were obtained from the analysisof Cyclin B levels. These experiments revealed that ROR2 slows-down G1-G2/M transition by regulating theabundance of cell cycle proteins. Thus, ROR2 has an anti-tumorigenic role inmelanoma which has important implicancies for the understanding of Wnt5asignaling.