MicroRNA-143 Regulates Adipocyte Differentiation

Abstract

MicroRNAs (miRNAs) are endogenously expressed 20 -24 nucleotide RNAs thought to repress protein translation through binding to a target mRNA (1-3). Only a few of the more than 250 predicted human miRNAs have been assigned any biological function. In an effort to uncover miRNAs important during adipocyte differentiation, antisense oligonucleotides (ASOs) targeting 86 human miRNAs were transfected into cultured human pre-adipocytes, and their ability to modulate adipocyte differentiation was evaluated. Expression of 254 miRNAs in differentiating adipocytes was also examined on a miRNA microarray. Here we report that the combination of expression data and functional assay results identified a role for miR-143 in adipocyte differentiation. miR-143 levels increased in differentiating adipocytes, and inhibition of miR-143 effectively inhibited adipocyte differentiation. In addition, protein levels of the proposed miR-143 target ERK5 (4) were higher in ASO-treated adipocytes. These results demonstrate that miR-143 is involved in adipocyte differentiation and may act through target gene ERK5.The first miRNA 1 was identified in Caenorhabditis elegans as a gene important for timing of larval development (5). miRNAs have since been implicated in many processes in invertebrates, including cell proliferation and apoptosis (6, 7), fat metabolism (6), and neuronal patterning (8). As many miRNAs are conserved across species (9 -11), they are likely to be involved in developmental processes in all animals. Only a few mammalian miRNAs have been assigned any function, and at least two of these are involved in developmental processes: miR-181 promotes B cell development in mice (12) and miR196a regulates several Hox genes (13), which code for a family of transcription factors involved in various developmental programs in animals (14).We hypothesized that miRNAs may play a role in maturation of human adipocytes. Understanding the molecular events involved in adipocyte differentiation is of interest for development of therapeutics for metabolic diseases such as obesity and diabetes. In vitro cell culture systems, such as human primary subcutaneous pre-adipocytes, have been crucial in uncovering signaling pathways important for adipocyte differentiation (15). These cells can be cultured with differentiation-promoting hormonal stimuli, causing them to develop into cells that morphologically and functionally resemble mature adipocytes. In this study we have inhibited a panel of miRNAs in pre-adipocytes using antisense oligonucleotides and evaluated the effect on adipocyte differentiation. Combined with expression analysis of miRNAs in differentiating adipocytes by microarray, one miRNA, miR-143, was identified which normally promotes adipocyte differentiation. These results indicate that miRNAs do play a role in adipocyte differentiation and are potential therapeutic targets for obesity and metabolic diseases. EXPERIMENTAL PROCEDURESOligonucleotide Synthesis-Oligonucleotides were prepared using conventional phosphoramidite chemistry and...

Role of anti-oncomirs miR-143 and -145 in human colorectal tumors

“…However, the upregulation of miR-7 and -21 and downregulation of miR-34a are significant markers in cancers. ERK5 was already determined to be a target gene of miR-143 by us [7][8][9][10][11] and Esau et al 23 Oncogene K-ras 24 MicroRNA-143 and -145 in human colorectal tumors Y Akao et al checkpoint kinase CHEK2 have also been suggested to be candidate target genes of miR-143. As putative targets of miR-145 with potential oncogenic functions are transcription factors such as MYCN, FOS, YES and FLI, cell-cycle promoters such as cyclin D2 and CDK3 and mitogenactivated protein kinase pathway (MAPK) transduction proteins such as MAP3K3 and MAPK4K4 (http:// microrna.sanger.ac.uk/targets/v5).…”

Chemical modification and design of anti-miRNA oligonucleotides

“…Several of these newer 2¢-modified chemistries have been used in AMOs and show similar benefits. Esau et al 24 described use of the 2¢-O-methyoxyethyl (2¢MOE) (Figure 1) modification in AMOs, where the oligonucleotides were fully 2¢MOE-PS modified ( Table 1). The 2¢MOE modification improves nuclease resistance and increases T m and has been shown to be more potent than 2¢OMe RNA when used in traditional anti-mRNA antisense LNA/2′OMe-PS Same as target Lennox 18 miR-122…”

Implications of micro-RNA profiling for cancer diagnosis

“…However, upregulation of specific miRNAs, such as the mir-17-92 cluster described above, appears to be associated with oncogenesis (He et al, 2005b;Volinia et al, 2006). In addition, other miRNAs, including miR125b and miR-143, have been directly associated with the differentiation and proliferation of certain cell types (Esau et al, 2004;Lee et al, 2005). Finally, miRNA expression patterns may change when cells are treated with differentiation-promoting agents (Kasashima et al, 2004;Stegmaier et al, 2004), and recent studies have shown that undifferentiated human embryonic stem cells express certain miRNAs (Suh et al, 2004).…”