A Functional Polymorphism in the Promoter of MiR-143/145 Is Associated With the Risk of Cervical Squamous Cell Carcinoma in Chinese Women: A Case-Control Study

Abstract

MiR-143/145 is down-regulated in cervical cancer, which may serve as a tumor suppressor by targeting KRAS and Ras-responsive element-binding protein (RREB1). Activated KRAS leads to down-regulation of miR-143/145 transcription in a RREB1-dependent manner, establishing a miR-143/145-KRAS-RREB1 feedback loop. A polymorphism rs4705343C/T in the promoter of miR-143/145 might influence the binding of TATA-binding protein. We hypothesized that the miR-143/145 rs4705343 and KRAS rs712 may be related to the occurrence of cervical squamous cell carcinoma (CSCC). In this study, we genotyped the 2 polymorphisms in 415 patients with CSCC and 504 controls using polymerase chain reaction-restriction fragment length polymorphism. The promoter activities were measured by the Dual-Luciferase Reporter Assay System. We found that the rs4705343TC genotype was associated with an increased risk of CSCC (adjusted odds ratio [OR] = 1.37; 95% confidence interval [CI], 1.05-1.80). The significantly increased association was also observed in a dominant genetic model (adjusted OR = 1.32; 95% CI, 1.01-1.72). Combined analysis showed that individuals carrying the genotypes of rs4705343 TC/CC and rs712GT/TT had a 1.47-fold increased risk of CSCC (adjusted OR = 1.47; 95% CI, 1.01-2.15). By using multifactor dimensionality reduction software method, we identified a significant interaction between the miR-143/145 rs4705343 and KRAS rs712. Dual-Luciferase Reporter Assay showed that the luciferase activity was significantly lower in cells transfected with the rs4705343C allele than that of the rs4705343T allele. These findings indicate that miR-143/145 rs4705343 and KRAS rs712 may contribute to the etiology of CSCC in Chinese women.

FIGURE 1

A: miR-143/145-KRAS-RREB1 feedback loop. B: Bioinformatics analysis predicted a TATA-binding protein (TBP) binding site in the promoter region of miR-143/145 (underlined sequence). If the polymorphic location is T, it can bind to the transcriptional factor TBP; otherwise, if the polymorphic location is C, it cannot bind to the TBP.

FIGURE 2

Deoxyribonucleic acid sequencing of the miR-143/145 rs4705343 and KRAS rs712 polymorphisms.

FIGURE 3

Expression of miR-143/145 in HeLa, A549, and HepG2 cell lines. Real-time polymerized chain reaction was performed to detect miR-143/145 expression. Relative expression was normalized using U6 snRNA. Data were presented as mean ± standard deviation.

FIGURE 4

Effect of the miR-143/145 rs4705343 polymorphism on the transcriptional activity. A: Schematic representation of the promoter of miR-143/145 into pGL3-basic plasmid. B: The pGL3-rs4705343C and pGL3-rs4705343T constructs were transiently transfected into HeLa, A549, and HepG2 cells. The relative luciferase activity was normalized with the internal control of Renilla luciferase activity. Each assay was performed in triplicate. Data were presented as mean ± standard deviation. ^∗∗P < 0.01.