Fig. 5: Mechanism dissection of how Enz influences miR-361-3p expression.

a Quantitative real-time PCR analysis shows that miR-361-3p expression in C4-2 Enz-R cells is lower than in their parental Enz-sensitive C4-2 cells. b Western blot analysis shows that androgen receptor splicing variant 7 (ARv7) protein expression increased in C4-2 Enz-R cells compared to parental Enz-sensitive C4-2 cells. c, d Quantitative real-time PCR analysis shows that Enz decreases miR-361-3p expression in (c) C4-2 and (d) LNCaP parental cells after 72 h, but not at shorter times (24 or 48 h). e C4-2 cells treated with Enz for 24 h have decreased miR-361-3p expression under hypoxia (left); meanwhile, the expression of hypoxia-responsive genes is as expected (right). f Rescue assays show that decreased miR-361-3p expression in C4-2 cells could be reversed when hypoxia is antagonized by HIF-2α inhibitor TC-S 7009 (left). Expression of hypoxia-responsive genes is indeed reduced after treating with the inhibitor (right). g LNCaP cells treated with Enz for 24 h have decreased miR-361-3p expression under hypoxia (left); meanwhile, the expression of hypoxia-responsive genes is as expected (right). h Rescue assays show that decreased miR-361-3p expression in LNCaP cells could be reversed when hypoxia is antagonized by HIF-2α inhibitor TC-S 7009 (left). Expression of hypoxia-responsive genes is indeed reduced after treating with the inhibitor (right). i Hypoxia-responsive genes are increased in C4-2 Enz-R cells. Representative results are shown as mean ± SD. *P < 0.05.