Regulatory interaction of HNF1-alpha to microRNA-194 gene during intestinal epithelial cell differentiation.
ABSTRACT Maintenance of intestinal epithelium is based on well-balanced molecular mechanisms that confer the stable and continuous supply of specialized epithelial cell lineages from multipotent progenitors. Lineage commitment decisions in intestinal epithelium system involve multiple regulatory systems that interplay each other to establish the cellular identities. Here, we demonstrate that the microRNA system could be involved in intestinal epithelial cell differentiation and that microRNA-194 (miR194) is highly induced during this process and controlled by a transcription factor, HNF-alpha, that is well known to regulate gene expression in intestinal epithelial cells. Thus, the 5' conserved genomic region of miR-1942 gene, the inducible class of miR-194 parental gene, contains a binding motif for HNF1-alpha. This consensus region is required for the transcription of miR-1942 and active in intestinal epithelial cell line, Caco-2, in-vivo. Our observations indicate that microRNA genes could be targets of lineage specific transcription factors and that microRNAs are regulated in intestinal epithelial cells in a tissue specific manner. Given that role of microRNA in fine tuning of gene expression patterns, our results suggest that HNF1-alpha regulates the gene expression program by not only direct activation of genes but also modulation through induction of microRNAs such as miR-194, in intestinal epithelial cells. This represents a novel molecular machinery that might specify the fates of intestinal epithelial cell lineages during their differentiation.
- [Show abstract] [Hide abstract]
ABSTRACT: Micro (mi)RNAs are abundant small noncoding RNAs found in plants and animals, the regulatory functions of which are not fully understood in fish. To identify potential miRNAs, we screened an miRNA microarray with total RNA from zebrafish infected with Vibrio harveyi and another from uninfected zebrafish. Six miRNAs were obtained from the microarray screening. We studied miRNA expression patterns of 2 miRNAs (miR-122 and miR-194) after bacterial infection of transgenic zebrafish (containing tilapia hepcidin (TH)2-3) and non-transgenic zebrafish from which the 2 miRNAs were obtained from the microarray experiment. The results indicated that miR-122 and miR-194 were higher in PBS-injected zebrafish compared with TH2-3 zebrafish or wild-type (WT) zebrafish after V. harveyi infection. Overexpression of miRNAs (miR-122, miR-192, and miR-194a) was seen in zebrafish liver (ZFL) cells after lipopolysaccharide (LPS) treatment and in untreated fish. Our results showed that after 24 h of doxycycline treatment without LPS stimulation, interleukin (IL)-22, lysozyme, toll-like receptor (TLR)1, TLR3, TLR4a, and tumor necrosis factor (TNF)-α gene expressions were, respectively, upregulated by ~14-, 22-, 2.2-, 13-, 200-, and 38-fold in miR-122-transfected compared with non-transfected (WT) ZFL cells. In cells transfected with miR-192 and treated with LPS after 8-12 h, IL-22, lysozyme, TLR1, TLR3, TLR4a, and TNF-α expressions significantly differed between WT and miR-192-overexpressing ZFL cells. However, we observed significantly higher IL-22 expression levels after 12 h of LPS treatment in miR-192-transfected ZFL cells compared with non-transfected cells. In contrast, IL-22, lysozyme, and TNF-α were markedly upregulated (>100-fold) after miR-194a transfection and overexpression in ZFL cells and treatment with LPS. Our cloning and expression analyses indicated that miR-122, miR-192, and miR-194a play important roles in zebrafish immunology.Fish Physiology and Biochemistry 03/2012; 38(5):1299-310. · 1.55 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Trastuzumab, a humanized monoclonal antibody directed against the extracellular domain of the HER2 oncoprotein, can effectively target HER2-positive breast cancer through several mechanisms. Although the effects of trastuzumab on cancer cell proliferation, angiogenesis and apoptosis have been investigated in depth, the effect of trastuzumab on microRNA (miRNA) has not been extensively studied. We have performed miRNA microarray profiling before and after trastuzumab treatment in SKBr3 and BT474 human breast cancer cells that overexpress HER2. We found that trastuzumab treatment of SKBr3 cells significantly decreased five miRNAs and increased three others, whereas treatment of BT474 cells significantly decreased two miRNAs and increased nine. The only change in miRNA expression observed in both cell lines following trastuzumab treatment was upregulation of miRNA-194 (miR-194) that was further validated in vitro and in vivo. Forced expression of miR-194 in breast cancer cells that overexpress HER2 produced no effect on apoptosis, modest inhibition of proliferation, significant inhibition of cell migration/invasion in vitro and significant inhibition of xenograft growth in vivo. Conversely, knockdown of miR-194 promoted cell migration. Increased miR-194 expression markedly reduced levels of the cytoskeletal protein talin2 and specifically inhibited luciferase reporter activity of a talin2 wild-type 3'-untranslated region, but not that of a mutant reporter, indicating that talin2 is a direct downstream target of miR-194. Trastuzumab treatment inhibited breast cancer cell migration and reduced talin2 expression in vitro and in vivo. Knockdown of talin2 inhibited cell migration/invasion. Knockdown of trastuzumab-induced miR-194 expression with a miR-194 inhibitor compromised trastuzumab-inhibited cell migration in HER2-overexpressing breast cancer cells. Consequently, trastuzumab treatment upregulates miR-194 expression and may exert its cell migration-inhibitory effect through miR-194-mediated downregulation of cytoskeleton protein talin2 in HER2-overexpressing human breast cancer cells.PLoS ONE 07/2012; 7(7-10.1371/journal.pone.0041170). · 3.73 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: MicroRNAs (miRNAs) are members of non-coding RNAs. They are involved in diverse biological functions. MiRNAs are precisely regulated in a tissue- and developmental-specific manner, but dysregulated in many human diseases, in particular cancers. Transcriptional regulation, post-transcriptional regulation, as well as genetic alterations, are the three major mechanisms controlling the spatial and temporal expression of miRNAs. Emerging evidence now indicates that transcriptional and epigenetic regulations play major roles in miRNA expression. This review summarizes the current knowledge and discusses the future challenges.Cancer letters 12/2012; · 4.86 Impact Factor