Cooperative Repression of Insulin-Like Growth Factor Type 2 Receptor Translation by MicroRNA 195 and RNA-Binding Protein CUGBP1

Abstract

Insulin-like growth factor type 2 (IGF2) receptor (IGF2R) recognizes mannose 6-phosphate-containing molecules and IGF2 and plays an important role in many pathophysiological processes, including gut mucosal adaptation. However, the mechanisms that control cellular IGF2R abundance are poorly known. MicroRNAs (miRNAs) and RNA-binding proteins (RBPs) critically regulate gene expression programs in mammalian cells by modulating the stability and translation of target mRNAs. Here we report that miRNA 195 (miR-195) and RBP CUG-binding protein 1 (CUGBP1) jointly regulate IGF2R expression at the posttranscriptional level in intestinal epithelial cells. Both miR-195 and CUGBP1 interacted with the 3= untranslated region (3=-UTR) of Igf2r mRNA, and the association of CUGBP1 with Igf2r mRNA enhanced miR-195 binding to Igf2r mRNA. Ectopically expressed CUGBP1 and miR-195 repressed IGF2R translation cooperatively without altering the stability of Igf2r mRNA. Importantly, the miR-195-and CUGBP1-repressed levels of cellular IGF2R led to a disruption in the structure of the trans-Golgi network. These findings indicate that IGF2R expression is controlled posttranscriptionally by two factors that associate with Igf2r mRNA and suggest that miR-195 and CUGBP1 dampen IGF signaling by inhibiting IGF2R translation.KEYWORDS insulin superfamily, posttranscriptional regulation, trans-Golgi network, intestinal epithelial cells, RNA-binding proteins, noncoding RNAs M ucosal regeneration/adaptation is an essential process in gut homeostasis and tightly regulated by numerous factors, including insulin-like growth factor type 1 (IGF1) and IGF2 (1, 2). Two structurally distinct types of receptors, the IGF1 receptor (IGF1R) and IGF2R, specifically interact with IGF1 and IGF2 as ligands and regulate diverse biological functions such as cell proliferation, differentiation, and apoptosis (3, 4). Like the insulin receptor, IGF1R is a heterotetrameric transmembrane receptor tyrosine kinase, and its activation results in autophosphorylation of tyrosine residues in the intracellular ␤-subunits, thus initiating the phosphatidylinositol 3-kinase (PI3) kinase/AKT and/or mitogen-activated protein (MAP) kinase signaling cascade (5). In contrast, IGF2R is a single-transmembrane-domain protein and binds IGF2 with greater affinity than IGF1, although it does not accept insulin as a ligand (4, 6). IGF2R also recognizes, via distinct sites, mannose-6-phosphate (M6P)-containing molecules and can therefore associate with other growth factors and cytokines (4). The majority of IGF2R is localized in the trans-Golgi network (TGN) and endosomal compartments and, to a lesser extent, on the cell surface (6). A subpopulation of the IGF2R on the plasma membrane regulates IGF2 internalization and various M6P-containing ligands for their

Long Noncoding RNA uc.173 Promotes Renewal of the Intestinal Mucosa by Inducing Degradation of MicroRNA 195

“…26,39–41 We have recently reported that miRNA195 represses translation of insulin-like growth factor type 2 receptor by cooperating with the RNA-binding protein CUGBP1. 43 Moreover, the use of systemically administered LNA-mediated antagonism in exploring T-UCR function underscores the potential value of developing therapeutic approaches to alter intestinal mucosal growth in the clinical setting. The fact that inhibiting a single T-UCR ( uc.173 ) by LNA-anti-uc.173 oligonucleotides can induce a significant inhibition of mucosal growth in vivo highlights the power of targeting individual T-UCRs in modulating gut mucosal growth.…”

Regulation of Intestinal Epithelial Barrier Function by Long Noncoding RNA uc.173 through Interaction with MicroRNA 29b

“…An expression vector containing a 284-bp fragment flanking the human uc.173 locus under the control of the pCMV promoter was constructed as described previously (29). The chimeric firefly luciferase reporter constructs containing the full-length CLDN1 5= UTR, CR, or different 3= UTR fragments were subcloned into the Dual-Luciferase miRNA target expression vector pmirGLO (Promega, Madison, WI) to generate pmirGLO-Luc-CLDN1-5=UTR, pmirGLO-Luc-CLDN1-CR, and pmirGLO-CLDN1-3=UTR as described previously (49). DNA sequencing and enzyme digestion were used to confirm the sequence and orientation of the fragment in the luciferase reporter.…”

Interaction between HuR and circPABPN1 Modulates Autophagy in the Intestinal Epithelium by Altering ATG16L1 Translation

“…The Atg16l1 mRNA is a potential target of HuR, given that the human ATG16L1 mRNA shows extensive association with HuR (see Table S1 in the supplemental material) (http://starbase.sysu.edu.cn/). To determine whether HuR enhances ATG16L1 translation by directly interacting with the Atg16l1 mRNA, ribonucleoprotein (RNP) immunoprecipitation (RIP) assays were performed in IECs using anti-HuR antibody under conditions that preserved RNP integrity (36). Atg16l1 mRNA was highly enriched in HuR samples compared with control IgG samples, while HuR did not preferentially associate with Atg5 mRNA (Fig.…”