title: Micro-RNA30c negatively regulates REDD1 expression in human hematopoietic and osteoblast cells after gamma-irradiation.
abstract: We recently demonstrated that a novel cell stress response gene REDD1 protects human fetal osteoblast cell line (hFOB) cells from γ-radiation-induced premature senescence. Here we show that levels of endogenous REDD1 are very low in human hematopoietic progenitor CD34+ cells regardless of radiation, but highly expressed in differentiated hematopoietic cells (14 day cultured CD34+ cells) in response to radiation, which might be associated with radiation tolerance of the latter cells. To further understand the mechanisms of radiation-induced damage in different cells, microRNA (miRNA)-arrays were performed using purified miRNAs from CD34+ and hFOB cells before and post-irradiation and real-time reverse transcription (RT)-PCR was used to validate the expression profiles of miRNAs in the radiation-damaged cells. The results indicate that γ-radiation downregulated 16 miRNAs in CD34+ cells and 14 in hFOB cells. Radiation-induced upregulation was observed for 15 miRNAs in CD34+ cells and 18 miRNAs in hFOB cells. The profiles of radiation-induced miRNA expression were completely different in CD34+ vs. hFOB cells. Radiation up-regulated miRNA (miR)-30b, miR-30c and miR-30d in CD34+ cells, whereas it inhibited miR-30c expression in hFOB cells. Since miR-30 has potential target sites located in the 3'untranslated region (UTR) of the REDD1 gene and radiation regulated miR-30c expression in both CD34+ and hFOB cells, we further explored the effects of miR-30c on REDD1 expression using miR-30c inhibitor and precursor (pre-miR-30c). The results show that pre-miR-30c transfection suppressed REDD1 expression in 14 day cultured CD34+ cells and hFOB cells and resulted in hFOB cell death. In contrast, inhibition of miR-30c expression significantly enhanced clonogenicity in CD34+ cells. Our data suggest that CD34+ and hFOB cells have different miRNA expression patterns after irradiation and miR-30c plays a key role in radiation-induced cell damage which might be through regulation of REDD1 expression.
authors.affiliation: Radiation Countermeasures Program, Armed Forces Radiobiology Research Institute, Uniformed Services University of the Health Sciences, Bethesda, Maryland, United States of America.
authors.forename: Xiang Hong
authors.initials: XH
authors.lastname: Li
authors.forename: Cam T
authors.initials: CT
authors.lastname: Ha
authors.forename: Dadin
authors.initials: D
authors.lastname: Fu
authors.forename: Mang
authors.initials: M
authors.lastname: Xiao
mesh_terms: D018952:Antigens, CD34 / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D016000:Cluster Analysis; D005720:Gamma Rays; D005786:Gene Expression Regulation / Q000528:radiation effects*; D006412:Hematopoietic Stem Cells / Q000378:metabolism / Q000528:radiation effects*; D006801:Humans; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D020411:Oligonucleotide Array Sequence Analysis; D010006:Osteoblasts / Q000378:metabolism / Q000528:radiation effects*; D060888:Real-Time Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
chemical_list: D018952:Antigens, CD34; C469991:DDIT4 protein, human; C546086:MIRN30b microRNA, human; D035683:MicroRNAs; D014157:Transcription Factors