miR-183 Inhibits UV-Induced DNA Damage Repair in Human Trabecular Meshwork Cells by Targeting of KIAA0101.
OBJECTIVE
The purpose of this study was to investigate the mechanisms by which miR-183 may contribute to the phenotypic alterations associated with stress-induced senescence of human trabecular meshwork (HTM) cells.


METHODS
Changes in gene expression induced by miR-183 in HTM cells were evaluated by gene array analysis, confirmed by quantitative-PCR (Q-PCR), and analyzed by MetaCore pathway analysis. Effects of miR-183 on cell proliferation were assessed by incorporation of bromodeoxyuridine incorporation, and DNA damage by CometAssay after ultraviolet (UV) irradiation in primary HTM cells, and confirmed in human diploid fibroblasts (HDF) and HeLa cells. A plasmid expressing KIAA0101 without its 3'-untranslated region (3'-UTR) was cotransfected with miR-183 to evaluate the role of KIAA0101 on the effects induced by miR-183.


RESULTS
miR-183 affected the expression of multiple genes involved in cell cycle regulation and DNA damage response in HTM cells. Forced expression of miR-183 in HTM and HDF resulted in a significant decrease in proliferation in primary HTM and HDF cells but not in HeLa cells. In all cell types tested, overexpression of miR-183 resulted in increased DNA damage under UV irradiation. Expression of KIAA0101 lacking the 3'-UTR region partially prevented the effects of miR-183 on cell proliferation and completely reversed the effects on UV-induced DNA damage.


CONCLUSIONS
Our results suggest that the observed up-regulation of miR-183 after stress-induced senescence in HTM cells may contribute to reinforce cellular senescence by inhibiting cell cycle progression through multiple gene targets and limiting the DNA repair mechanisms through inhibition of KIAA0101.
Guorong
G
Li
Coralia
C
Luna
Pedro
P
Gonzalez
D015153:Blotting, Western; D002352:Carrier Proteins / Q000502:physiology*; D002478:Cells, Cultured; D020552:Comet Assay; D004249:DNA Damage / Q000528:radiation effects*; D004260:DNA Repair / Q000502:physiology*; D004268:DNA-Binding Proteins; D005347:Fibroblasts / Q000502:physiology; D015870:Gene Expression / Q000502:physiology; D006367:HeLa Cells / Q000502:physiology; D006801:Humans; D035683:MicroRNAs / Q000502:physiology*; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D014129:Trabecular Meshwork / Q000166:cytology / Q000502:physiology / Q000528:radiation effects*; D014466:Ultraviolet Rays / Q000009:adverse effects
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; C533449:MIRN183 microRNA, human; D035683:MicroRNAs; C425208:PCLAF protein, human