MicroRNA-99 family members suppress Homeobox A1 expression in epithelial cells

• Published in: PloS one 2013-12, Vol.8 (12), p.e80625-e80625
• Main Authors: Chen, Dan, Chen, Zujian, Jin, Yi, Dragas, Dragan, Zhang, Leitao, Adjei, Barima S, Wang, Anxun, Dai, Yang, Zhou, Xiaofeng
• Format: Article
• Language: English
• Subjects: Actin; Algorithms; Apoptosis; Bcl-2 protein; Bioinformatics; Cancer; Cell adhesion & migration; Cell growth; Cell Line; Cell migration; Cell proliferation; Chromatin; Dentistry; Downstream effects; Embryogenesis; Embryonic growth stage; Epithelial cells; Gene expression; Gene Expression Profiling; Gene Expression Regulation - physiology; Gene sequencing; Genes; Genome-Wide Association Study; Genomes; Healing; Homeobox; Homeobox genes; Homeodomain Proteins - biosynthesis; Homeodomain Proteins - genetics; Hospitals; HOXA1 protein; Humans; Immunoprecipitation; Kinases; Luciferase; Maxillofacial surgery; Membrane proteins; MicroRNA; MicroRNAs; MicroRNAs - biosynthesis; MicroRNAs - genetics; miRNA; Molecular biology; N-Myristoyltransferase; Oral diseases; Pancreatic cancer; Penicillin; Physiological aspects; Physiology; Proto-Oncogene Proteins - biosynthesis; Proto-Oncogene Proteins - genetics; Rapamycin; Reporter gene; Ribonucleic acid; RNA; Rodents; Skin; Studies; Surgery; Tissues; TOR protein; Transcription Factors - biosynthesis; Transcription Factors - genetics; Transfection; Tumorigenesis; Wound healing
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0080625

The miR-99 family is one of the evolutionarily most ancient microRNA families, and it plays a critical role in developmental timing and the maintenance of tissue identity. Recent studies, including reports from our group, suggested that the miR-99 family regulates various physiological processes in adult tissues, such as dermal wound healing, and a number of disease processes, including cancer. By combining 5 independent genome-wide expression profiling experiments, we identified a panel of 266 unique transcripts that were down-regulated in epithelial cells transfected with miR-99 family members. A comprehensive bioinformatics analysis using 12 different sequence-based microRNA target prediction algorithms revealed that 81 out of these 266 down-regulated transcripts are potential direct targets for the miR-99 family. Confirmation experiments and functional analyses were performed to further assess 6 selected miR-99 target genes, including mammalian Target of rapamycin (mTOR), Homeobox A1 (HOXA1), CTD small phosphatase-like (CTDSPL), N-myristoyltransferase 1 (NMT1), Transmembrane protein 30A (TMEM30A), and SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A member 5 (SMARCA5). HOXA1 is a known proto-oncogene, and it also plays an important role in embryonic development. The direct targeting of the miR-99 family to two candidate binding sequences located in the HOXA1 mRNA was confirmed using a luciferase reporter gene assay and a ribonucleoprotein-immunoprecipitation (RIP-IP) assay. Ectopic transfection of miR-99 family reduced the expression of HOXA1, which, in consequence, down-regulated the expression of its downstream gene (i.e., Bcl-2) and led to reduced proliferation and cell migration, as well as enhanced apoptosis. In summary, we identified a number of high-confidence miR-99 family target genes, including proto-oncogene HOXA1, which may play an important role in regulating epithelial cell proliferation and migration during physiological disease processes, such as dermal wound healing and tumorigenesis.