miR-24 Regulates Apoptosis by Targeting the Open Reading Frame (ORF) Region of FAF1 in Cancer Cells

Abstract

BackgroundmicroRNAs (miRNAs) are small noncoding RNAs that regulate cognate mRNAs at the post-transcriptional stage. Several studies have shown that miRNAs modulate gene expression in mammalian cells by base pairing to complementary sites in the 3′-untranslated region (3′-UTR) of the target mRNAs.Methodology/Principal FindingsIn the present study, miR-24 was found to target fas associated factor 1(FAF1) by binding to its amino acid coding sequence (CDS) region, thereby regulating apoptosis in DU-145 cells. This result supports an augmented model whereby animal miRNAs can exercise their effects through binding to the CDS region of the target mRNA. Transfection of miR-24 antisense oligonucleotide (miR-24-ASO) also induced apoptosis in HGC-27, MGC-803 and HeLa cells.Conclusions/SignificanceWe found that miR-24 regulates apoptosis by targeting FAF1 in cancer cells. These findings suggest that miR-24 could be an effective drug target for treatment of hormone-insensitive prostate cancer or other types of cancers. Future work may further develop miR-24 for therapeutic applications in cancer biology.

Exogenous plant MIR168a specifically targets mammalian LDLRAP1: evidence of cross-kingdom regulation by microRNA

“…A luciferase reporter assay was employed to demonstrate the direct binding of MIR168a to exon 4 of LDLRAP1. The same strategy has been used by others in studying miRNA targeting in coding regions [30][31][32]. In this experiment, the wild-type (WT) or mutant (MUT) MIR168a complementary site (CS), WT or MUT LDLRAP1 binding site (BS), the human LDLRAP1 exon 4, and human LDLRAP1 coding sequence (CDS) were cloned into a luciferase reporter plasmid ( Figure 3G) and transfected into HepG2 cells combined with pre-MIR168a.…”

“…We found that the rate of the inhibition of luciferase activity by MIR168a was dependent on the ratio of pre-MIR168a to BS-containing luciferase reporter (Supplementary information, Figure S3A) and that the efficiency of MIR168a inhibition of LDLRAP1 expression was similar to the inhibition of target genes by endogenous miRNAs (miR-16, miR-21, and miR-150; Supplementary information, Figure S3B). To determine whether MIR168a could directly target LDLRAP1 within its CDS, we followed the strategy proposed by others [30][31][32] and created a construct that expressed the full-length ORF of LDLRAP1 ( Figure 3I). Furthermore, we used site-directed mutagenesis to create a variant of the LDLRAP1 construct in which the MIR168a target site was mutated while leaving the LDLRAP1 aminoacid sequence intact ( Figure 3I).…”

Major apoptotic mechanisms and genes involved in apoptosis

“…Another oncogenic miRNA exerting its anti-apoptotic effects by inhibiting FasL directly is miR-21, which is found to be upregulated in advanced pancreatic cancer patients [165]. miR-24 regulates apoptosis by binding to coding sequence of Fas-associated factor 1 (FAF1) mRNA and induced apoptosis of several different types of cancer [166]. miR-21 also negatively regulates PTEN, a tumor suppressor gene, involved in the apoptotic pathway through the formation of DISC complex in many tumors such as breast and gastric cancers [167,168].…”

Aryl hydrocarbon receptor nuclear translocator in human liver is regulated by miR-24

“…In animals, miRNA target sites are located mainly in the 3'-untranslated region (3'-UTR) of target mRNAs (Chekulaeva and Filipowicz, 2009), although there are examples of target sites in the coding region (Qin et al, 2010). Until now, over 1400 miRNAs have been identified in humans.…”