Identification of microRNA-mRNA functional interactions in UVB-induced senescence of human diploid fibroblasts

Ruth Greussing,Christoph Mueck,Marcel Scheideler,Pidder Jansen-Dürr,Matthias Hackl,Michael Neuhaus,Zlatko Trajanoski,Rossella Monteforte,Johannes Grillari,Alexander Pauck,Maria Cavinato,Pornpimol Charoentong,Lucia Micutkova,Edith Hofer

doi:10.1186/1471-2164-14-224

Abstract

BackgroundCellular senescence can be induced by a variety of extrinsic stimuli, and sustained exposure to sunlight is a key factor in photoaging of the skin. Accordingly, irradiation of skin fibroblasts by UVB light triggers cellular senescence, which is thought to contribute to extrinsic skin aging, although molecular mechanisms are incompletely understood. Here, we addressed molecular mechanisms underlying UVB induced senescence of human diploid fibroblasts.ResultsWe observed a parallel activation of the p53/p21WAF1 and p16INK4a/pRb pathways. Using genome-wide transcriptome analysis, we identified a transcriptional signature of UVB-induced senescence that was conserved in three independent strains of human diploid fibroblasts (HDF) from skin. In parallel, a comprehensive screen for microRNAs regulated during UVB-induced senescence was performed which identified five microRNAs that are significantly regulated during the process. Bioinformatic analysis of miRNA-mRNA networks was performed to identify new functional mRNA targets with high confidence for miR-15a, miR-20a, miR-20b, miR-93, and miR-101. Already known targets of these miRNAs were identified in each case, validating the approach. Several new targets were identified for all of these miRNAs, with the potential to provide new insight in the process of UVB-induced senescence at a genome-wide level. Subsequent analysis was focused on miR-101 and its putative target gene Ezh2. We confirmed that Ezh2 is regulated by miR-101 in human fibroblasts, and found that both overexpression of miR-101 and downregulation of Ezh2 independently induce senescence in the absence of UVB irradiation. However, the downregulation of miR-101 was not sufficient to block the phenotype of UVB-induced senescence, suggesting that other UVB-induced processes induce the senescence response in a pathway redundant with upregulation of miR-101.ConclusionWe performed a comprehensive screen for UVB-regulated microRNAs in human diploid fibroblasts, and identified a network of miRNA-mRNA interactions mediating UVB-induced senescence. In addition, miR-101 and Ezh2 were identified as key players in UVB-induced senescence of HDF.

Highlights

Cellular senescence can be induced by a variety of extrinsic stimuli, and sustained exposure to sunlight is a key factor in photoaging of the skin
We have addressed molecular mechanisms underlying the establishment of premature senescence in human fibroblasts exposed to repeated series of mild Ultraviolet light band B (UVB) irradiations, as a model system to monitor molecular processes involved in extrinsic skin aging
Together the data suggest that mild UVB stress induces premature cellular senescence and that both the p53/p21WAF1 axis and the p16INK4A/Retinoblastoma protein (pRb) pathway are involved in the senescence response, consistent with previous reports [23,24,25]

Summary

Introduction

Cellular senescence can be induced by a variety of extrinsic stimuli, and sustained exposure to sunlight is a key factor in photoaging of the skin. Irradiation of skin fibroblasts by UVB light triggers cellular senescence, which is thought to contribute to extrinsic skin aging, molecular mechanisms are incompletely understood. We addressed molecular mechanisms underlying UVB induced senescence of human diploid fibroblasts. Using genome-wide transcriptome analysis, we identified a transcriptional signature of UVB-induced senescence that was conserved in three independent strains of human diploid fibroblasts (HDF) from skin. Several new targets were identified for all of these miRNAs, with the potential to provide new insight in the process of UVB-induced senescence at a genome-wide level. Upregulation of the CDK inhibitor p16INK4a leads to the inhibition of phosphorylation of the retinoblastoma protein (pRb), thereby enforcing prolonged cell cycle arrest [3], which is a useful marker for senescence in human tissues [4]. It should be noted that, besides these central pathways, a variety of signals have been shown to initiate a senescence response [8]

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