Conserved Senescence Associated Genes and Pathways in Primary Human Fibroblasts Detected by RNA-Seq

Abstract

Cellular senescence correlates with changes in the transcriptome. To obtain a complete view on senescence-associated transcription networks and pathways, we assessed by deep RNA sequencing the transcriptomes of five of the most commonly used laboratory strains of human fibroblasts during their transition into senescence. In a number of cases, we verified the RNA-seq data by real-time PCR. By determining cellular protein levels we observed that the age-related expression of most but not all genes is regulated at the transcriptional level. We found that 78% of the age-affected differentially expressed genes were commonly regulated in the same direction (either up- or down-regulated) in all five fibroblast strains, indicating a strong conservation of age-associated changes in the transcriptome. KEGG pathway analyses confirmed up-regulation of the senescence-associated secretory phenotype and down-regulation of DNA synthesis/repair and most cell cycle pathways common in all five cell strains. Newly identified senescence-induced pathways include up-regulation of endocytotic/phagocytic pathways and down-regulation of the mRNA metabolism and the mRNA splicing pathways. Our results provide an unprecedented comprehensive and deep view into the individual and common transcriptome and pathway changes during the transition into of senescence of five human fibroblast cell strains.

Unmasking Transcriptional Heterogeneity in Senescent Cells

“…It is noteworthy that the common senescence markers, CDKN2A , CDKN1A , and LMNB1 , were not within the core signature, supporting the idea that known senescence markers lack universality for different cell types and inducers of senescence [10]. Indeed, one of the studies used here [14] showed that p16INK4A mRNA levels are not always significantly changed in senescence in some fibroblast strains. Although RNA levels do not always reflect protein levels, p16INK4A expression is often used as a senescence marker.…”

“…However, the second and third principal components separated the cells according to senescence status, with some influence from the study/dataset of origin (Figure S1A). Nonetheless, one sample within one of the datasets (RS IMR90 [14]) clustered differently from its replicates. The aberrant clustering of this sample was reported in the original study, and this sample was then removed from further analysis (Figure S1A).…”

A lysosomal dimmer switch regulates cellular quiescence depth

“…Similar to deep quiescence, senescence and aging are at least partially driven by ROS and can be counteracted by the lysosome-autophagy pathway 6,8,18,19,36 . With this mechanistic similarity in mind, it was still striking to us that QDS correctly predicted cellular senescence and aging in a wide array of cell lines in vitro 21,22 and tissues in vivo 23,24 (Fig. 6b-d).…”

“…We show that lysosomal function, like a dimmer switch, continuously regulates quiescence depth and thus the proliferative potential of quiescence cells, by reducing the accumulation of intracellular reactive oxygen species (ROS). Furthermore, we found that a gene expression signature developed by comparing deep and shallow quiescent REF cells was able to correctly classify senescent and aging cells in a wide array of cell lines in vitro 21,22 and tissues in vivo 23,24 , suggesting the existence of shared regulatory mechanisms underlying these cell fates and a possible sequential transition from shallow to deep quiescence and eventually to irreversible senescence that may contribute to aging.…”

Transcriptional Signatures of Aging

“…For example, increased expression of genes involved in stress response and oxidative damage is observed in aging human brain, retina, skin and fibroblasts (Table 1) [26, 27, 30, 32, 53]. Similarly, genes involved in stress response and inflammation show increased expression with age in rodent brain, kidney, liver, muscle, and pancreatic cells, and in fruit flies [34, 37, 39, 54].…”