Upregulation of ryanodine receptor-1 dependent calcium signaling contributes to the malignance of high-grade serous ovarian cancer (HGSOC).

Abstract

Ca2+ signaling plays crucial roles in cancer metastasis and progression. We analyzed RNAseq data from TCGA database and our patient samples; and identified over-expression of ryanodine receptor 1 (RYR1), the target of the FDA-approved drug dantrolene, is associated with poor survival of HGSOC patients. RYR1 expression is also markedly higher in several HGSOC cell lines compared to normal human ovarian surface epithelial cells (HOSE) and fallopian tube epithelial cells. Suppression of RYR1 expression/activity by shRNA or dantrolene significantly inhibited proliferation and migration of HGSOC cells (OVCA432, OVCA433, OV90). Ca2+ fluorescence measurement showed that the resting [Ca2+] and Ca2+ release triggered by the RYR-agonist 4-chloro-m-cresol (4-cmc) were significantly higher in HGSOC cells than the HOSE cells. Store-operated Ca2+ entry (SOCE) induced by 4-cmc was also enhanced in HGSOC cells. The elevated resting [Ca2+], the 4-cmc-induced Ca2+ release and SOCE in the HGSOC cells were reduced to the control level by shRNA against RYR1; and were significantly potentiated by over-expression of RYR1. RYR-dependent endoplasmic reticulum (ER)-mitochondrial Ca2+ transfer was examined by expressing the ER specific Ca2+ biosensor G-CEPIA1er and mitochondria Ca2+ biosensor CEPIA2mt in HGSOC cells. Activation of RYR by 4-cmc triggered a large reduction in ER [Ca2+], which was associated with a robust increase in mitochondrial [Ca2+]. Activation of RYRs by photorelease of caged cADP-ribose in a subcellular region caused an immediate transient local reduction in ER [Ca2+] and a fast transient local increase in mitochondria [Ca2+]. In contrast, RyR-dependent ER-mitochondrial Ca2+ transfer was not observed in the HOSE cells. These observations are the first direct evidence of enhanced RYR-mediated ER-mitochondrial Ca2+ transfer in ovarian cancer cells, and upregulation of RYR1-dependent Ca2+-signaling pathway contributes to the malignant phenotypes of HGSOC cells.