title: Hybrid-polymerase chain reaction to identify novel target genes of miR-134 in paclitaxel resistant human ovarian carcinoma cells.
abstract: Increasing evidence has shown that miR-134 is involved in the promotion of tumorigenesis and chemoresistance. However, whether miR-134 participates in ovarian cancer chemoresistance and its functional targets still remains unclear. The objective of this study was to apply hybrid-polymerase chain reaction (PCR) to screen target genes of miR-134 in ovarian carcinoma paclitaxel resistant SKOV3-TR30 cells, and to provide a number of novel targets of miR-134 for further study of ovarian cancer paclitaxel resistance. The current study found that miR-134 was decreased in SKOV3-TR30 cells compared with the parental SKOV3 cell line. By applying hybrid-PCR, 8 putative target genes of miR-134 in SKOV3-TR30 cells were identified, includingC16orf72,PNAS-105,SRM,VIM,F-box protein 2,GAPDH,PRPF6andRPL41. Notably, the target sites ofVIMandPRPF6were not located in 3'untranslated region, but rather in the coding sequence region. By conducting a luciferase reporter assay, miR-134 was demonstrated to recognize the putative binding sites of these target genes includingVIMandPRPF6. Transfecting SKOV3-TR30 cells with miR-134 mimic and performing reverse transcription-PCR in addition to western blot analysis confirmed that miR-134 regulates vimentin expression at a post transcriptional level. This finding provides a novel perspective for studying the mechanism of miR-134/mRNA interaction. In conclusion, this study was the first to apply an effective method of hybrid-PCR to screen putative target mRNAs of miR-134 in paclitaxel resistant SKOV3-TR30 cells and indicate that miR-134 may contribute to the induction of SKOV3-TR30 paclitaxel resistance by targeting these genes.
authors.affiliation: Department of Obstetrics and Gynecology, Shengjing Hospital Affiliated to China Medical University, Shenyang, Liaoning 110004, P.R. China.
authors.forename: Ting
authors.initials: T
authors.lastname: Shuang
authors.affiliation: Department of Obstetrics and Gynecology, Shengjing Hospital Affiliated to China Medical University, Shenyang, Liaoning 110004, P.R. China.
authors.forename: Min
authors.initials: M
authors.lastname: Wang
authors.affiliation: Department of Obstetrics and Gynecology, Shengjing Hospital Affiliated to China Medical University, Shenyang, Liaoning 110004, P.R. China.
authors.forename: Shuang
authors.initials: S
authors.lastname: Chang
keywords: SKOV3-TR30; hybrid-polymerase chain reaction; miR-134; ovarian cancer; paclitaxel resistance; target mRNAs