title: Overexpression of miR-183/-96/-182 triggers neuronal cell fate in Human Retinal Pigment Epithelial (hRPE) cells in culture.
abstract: miR-183 cluster, composed of miR-183/-96/-182 genes, is highly expressed in the adult retina, particularly in photoreceptors. It involves in development, maturation and normal function of neuroretina. Ectopic overexpression of miR-183/-96/-182 genes was performed to assess reprogramming of hRPE cells. They were amplified from genomic DNA and cloned independently or in tandem configuration into pAAV.MCS vector. hRPE cells were then transfected with the recombinant constructs. Real-Time PCR was performed to measure the expression levels of miR-183/-96/-182 and that of several retina-specific neuronal genes such as OTX2, NRL, PDC and DCT. The transfected cells also were immunocytochemically examined for retina-specific neuronal markers, including Rhodopsin, red opsin, CRX, Thy1, CD73, recoverin and PKCα, to determine the cellular fate of the transfected hRPE cells. Data showed that upon miR-183/-96/-182 overexpression in hRPE cultures, the expression of neuronal genes including OTX2, NRL, PDC and DCT was also upregulated. Moreover, miR-183 cluster-treated hRPE cells were immunoreactive for neuronal markers such as Rhodopsin, red opsin, CRX and Thy1. Both transcriptional and translational upregulation of neuronal genes in miR-183 cluster-treated hRPE cells suggests that in vitro overexpression of miR-183 cluster could trigger reprogramming of hRPE cells to retinal neuron fate.
authors.affiliation: National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.
authors.forename: Maliheh
authors.initials: M
authors.lastname: Davari
authors.affiliation: National Institute of Genetic Engineering and Biotechnology, Tehran, Iran. Electronic address: soheili@nigeb.ac.ir.
authors.forename: Zahra-Soheila
authors.initials: ZS
authors.lastname: Soheili
authors.affiliation: Blood Transfusion Research Center High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
authors.forename: Shahram
authors.initials: S
authors.lastname: Samiei
authors.affiliation: Blood Transfusion Research Center High Institute for Research and Education in Transfusion Medicine, Tehran, Iran.
authors.forename: Zohreh
authors.initials: Z
authors.lastname: Sharifi
authors.affiliation: Department of Molecular Genetic, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
authors.forename: Ehsan Ranaei
authors.initials: ER
authors.lastname: Pirmardan
mesh_terms: D050976:Basic-Leucine Zipper Transcription Factors; D002478:Cells, Cultured; D065150:Cellular Reprogramming / Q000235:genetics*; D005136:Eye Proteins; D020555:GTP-Binding Protein Regulators; D005786:Gene Expression Regulation; D006801:Humans; D019746:Intramolecular Oxidoreductases; D035683:MicroRNAs / Q000235:genetics*; D005810:Multigene Family / Q000502:physiology*; D009474:Neurons / Q000166:cytology*; D051857:Otx Transcription Factors; D010750:Phosphoproteins; D055213:Retinal Pigment Epithelium / Q000166:cytology* / Q000378:metabolism; D014162:Transfection; D015854:Up-Regulation
chemical_list: D050976:Basic-Leucine Zipper Transcription Factors; D005136:Eye Proteins; D020555:GTP-Binding Protein Regulators; C533449:MIRN183 microRNA, human; C541492:MIRN96 microRNA, human; D035683:MicroRNAs; C538984:Mirn182 microRNA, human; C490037:NRL protein, human; C497659:OTX2 protein, human; D051857:Otx Transcription Factors; D010750:Phosphoproteins; C065631:phosducin; D019746:Intramolecular Oxidoreductases; C042501:dopachrome isomerase
keywords: Retinal neurons; hRPE cells; miR-183 cluster; microRNAs