A set of miRNAs participates in the cellular senescence program in human diploid fibroblasts

Abstract

Here we show that replicative senescence in normal human diploid IMR90 fibroblasts is accompanied by altered expression of a set of microRNAs (miRNAs) (senescence-associated miRNAs), with 14 and 10 miRNAs being either up or downregulated (42-fold), respectively, in senescent with respect to young cells. The expression of most of these miRNAs was also deregulated upon senescence induced by DNA damage (etoposide) or mild oxidative stress (diethylmaleate). Four downregulated miRNAs were part of miRNA family-17, recently associated to human cell and tissue aging. Moreover, eight upregulated and six downregulated miRNAs mapped in specific chromosomal clusters, suggesting common transcriptional regulation. Upon adoptive overexpression, seven upregulated miRNAs induced the formation of senescence-associated heterochromatin foci and senescence-associated b-galactosidase staining (Po0.05), which was accompanied, in the case of five of them, by reduced cell proliferation. Finally, miR-210, miR-376a*, miR-486-5p, miR-494, and miR-542-5p induced double-strand DNA breaks and reactive oxygen species accumulation in transfected cells. In conclusion, we have identified a set of human miRNAs induced during replicative and chemically induced senescence that are able to foster the senescent phenotype by prompting DNA damage. Replicative or cellular senescence, a state of irreversible arrest of cell division, was first described in cultures of human fibroblasts. 1 Since then, replicative senescence has been described in various mammalian cells. 2 The mechanisms underlying senescence include telomere shortening, upregulation of the CDKN1A (p21WAF1) and CDKN2A (p16INK4a and p14ARF) loci, and accumulation of DNA damage. 3 Telomeres become progressively shorter at every round of cell division and this leads to critically short telomere length sensed as double-strand DNA breaks. 4 DNA damage and DNA-damage response (DDR) could be common events to cellular senescence programs initiated by telomere dysfunction and aberrant oncogene activation. 5 Senescent cells are marked by lack of DNA replication; expression of senescence-associated b-galactosidase (SA-b-gal); accumulation of discrete nuclear foci that are termed senescence-associated heterochromatin foci (SAHFs); and senescence-associated DNA-damage foci (SDFs). SAHFs are detected by preferential binding of DNA dyes, such as 4 0 ,6-diamidino-2-phenylindole (DAPI), and the presence of certain heterochromatin-associated histone modifications (trimethyl-Lys9 Histone H3). SDFs are nuclear foci containing proteins that are associated to DNA damage (Ser139-phosphorylated histone H2AX -g-H2AX-and p53-binding protein-1-53BP1). 6 Senescent cells show striking changes in gene expression, including upregulation of cell-cycle inhibitors (p21WAF1 and p16INK4a) and secreted proteins involved in microenvironment remodeling (IL-6), 7 and downregulation of genes that facilitate cell-cycle progression (c-FOS, cyclin-A, cyclin-B, PCNA) 8 or that are involved in cell-cycle execution (FOXM1, UBE2C, TYMS). ...

Metformin lowers the threshold for stress-induced senescence: A role for the microRNA-200 family and miR-205

“…Expression profiling of microRNAs (miRNAs, a class or regulatory RNAs responsible for posttranscriptional gene silencing by either degrading the target mRNA or preventing its translation) have revealed that these genes can function as either tumor suppressors or oncogenes in human tumors. Several studies have revealed that a variety of miRNAs are also differentially regulated during replicative senescence of human fibroblasts; [67][68][69][70] importantly, senescent fibroblasts appear to express distinct collections of miRNAs that play a central role in the senescence-elicited tumor suppression. 71 Because miRNAs extensively studied in the EMT of cancer cells (e.g., miR-200 family) [72][73][74][75] have recently been involved in the oxidative stress signature that regulates cellular senescence and tumorigenesis, 76,77 we decided to survey whether EMT-related miRNAs were differentially expressed in dividing (early-passage) HDFs compared with late-passage fibroblasts that had suffered an accelerated replicative senescence due to chronic exposure to metformin.…”

DGCR8-mediated disruption of miRNA biogenesis induces cellular senescence in primary fibroblasts

“…Interestingly, the set of miRNAs with reduced expression during DGCR8-mediated arrest are also downregulated during oncogeneinduced senescence (Fig. 4A), replicative senescence (Bonifacio & Jarstfer, 2010;Marasa et al, 2010;Wang et al, 2011;Faraonio et al, 2012) or DNA-damage induced senescence (Greussing et al, 2013) in different strains of human primary fibroblasts. In addition, some of these miRNAs have been shown to bypass oncogene-induced senescence in human fibroblasts or epithelial cells (Voorhoeve et al, 2006;Borgdorff et al, 2010; and are overexpressed in tumors (Ivanovska et al, 2008).…”

MicroRNAs and their roles in aging

“…Progress has been made in this regard in C. elegans by observing alterations in lifespan of lin-4, mir-71 and other mutants (Boehm and Slack, 2005;de Lencastre et al, 2010). Additionally, it has recently been shown that overexpression of several miRNAs that are upregulated during cellular senescence results in the formation of characteristic features of senescent cells (Faraonio et al, 2011). Nevertheless, studies in mammalian models that constitutively or conditionally lack or overexpress miRNAs will provide compelling evidence for any conserved aging mechanism mediated by miRNAs.…”

“…The miR-17-92 cluster, along with its paralogous clusters miR106a-363 and miR-106b-25, regulate cellular senescence and are all downregulated in several aging models, whereas they are all upregulated in some cancers (Faraonio et al, 2011;. These miRNAs target PTEN, and it is thought that their downregulation during aging causes an increase in PTEN levels, which results in suppression of the IIS pathway (Fig.…”