WS4.6 MicroRNA-30s negatively regulate planar cell polarity genes in human cystic fibrosis bronchial epithelial cells

Abstract

Objectives: Mucociliary clearance (MCC), abnormal in CF, is physiologically regulated by epithelial ion transport together with mechanical phenomenon such as cilia movement and cough. Planar Cell Polarity (PCP) is a tightly controlled protein network that drives ciliogenesis and cilia function. Cilia structure and function have been studies in CF. Although the majority of these studies showed no structural abnormality and a normal cilia beat frequency (CBF), it has also been showed that ciliary disorientation, rather than ultrastructural abnormalities or slow CBF, may occur secondary to lung inflammation and result in delayed MCC. We hypothesized that CF HBEs may display abnormalities in PCP network which could further impair coordinated cilia function in the plan of the epithelium. Methods: We profiled PCP genes expression by PCR in HBEs. By quantitative RT-PCR, we determined that expression of CELSR3 and Vangl-1 (Van-Gogh like 1) is down-regulated in CF (F508del/F508del) HBEs as compared to NL cells. In contrast, Fz3, Fz6, Pk1 and Vangl-2 genes expression is upregulated in CF cells. In silico analysis revealed that Vangl-1 and Vangl-2, Pk1, Dvl1, Dvl2, and CELSR3 are putative targets of miR-30 family. Expression of miR-30a, miR-30b, miR-30c, miR-30d and miR-30e is elevated is CF-HBEs. Knock-down of miR-30s expression in CF-HBEs significantly increased expression of CELSR3, Fz6, Vangl-1, Pk1, Dvl 1 and Dvl2 but did not modify expression of Fz3, Vangl-2 and Dvl3. Conclusion: Our results validate several PCP genes as miR-30s target genes and suggest that decreasing miR-30s expression in CF HBEs could modulate cilia function through restoration of PCP genes expression.