Several lines of evidence suggest a link between genome integrity and ageing, but perhaps the most compelling is the premature ageing seen in mammals that have defects in DNA-repair processes. Typically, the mechanism linking DNA damage to ageing is postulated to be mutation accumulation or increased cell death. However, we pose the question: if DNA repair was 100% efficient and there is no opportunity for mutations to accumulate, would the very presence of ongoing repair processes have an effect on the physiology?
To investigate this idea our laboratory has become increasingly interested in the premature ageing Werner syndrome (WS). Clinically, WS individuals prematurely show many features of normal ageing including, greying hair, aged skin, and cataracts, and show pre-disposition to age-related diseases such as type II diabetes, atherosclerosis, and osteoporosis1. Additionally, there is an elevated susceptibility to rare mesenchymal tumours. WS is caused by loss of the RecQ helicase WRNp, resulting in defects in DNA repair and recombination2 that lead to chromosomal rearrangements and increased replication fork stalling3.
The molecular mechanism of in vivo ageing in WS appears to be associated with accelerated cell ageing. WS cells have a shortened in vitro lifespan and resemble normal senescent cells in that they are enlarged, flattened and granular, even at a young age, and have prominent F-actin stress fibres reminiscent of cells undergoing stress-induced premature senescence (SIPS) 4. Recent studies suggest that the shortened lifespan reflects a synergy between telomere-driven senescence and a novel telomere independent senescence mechanism that involves the p38MAPK stress-signalling pathway4. We hypothesise that increased replication fork stalling in WS cells is sensed as replication stress that triggers p38 activation leading to SIPS. Treatment with the p38 inhibitor, SB203580, rescues the senescent-like morphology of WS cells and increases the lifespan and growth rate to within the normal range, i.e., effectively preventing premature cell ageing in WS4. This suggests that p38 activation contributes to the abnormal cellular phenotype in WS and, furthermore, that some of the clinical phenotypes, such as the elevated rates of inflammatory disease, including type II diabetes, atherosclerosis, and osteoporosis seen in WS, may in part be due to chronic activation of p38. WS thus provides a powerful system to dissect the interplay between senescence, genome instability and cancer.
However, the question remains as to whether this phenomenon is simply a unique "private" mechanism at work in WS, or is a general ability of cells to respond to physiological levels of cell intrinsic genome instability. To address this, further study is underway to investigate whether a similar response is seen in genome instability syndromes such as Rothmund-Thomson, Bloom, Hutchinson-Gilford, Nijmegen-breakage (NBS1) and Cockayne (CSA), amongst others. Initial studies focus on determining whether primary fibroblasts show any of the changes in cell behaviour that are seen in WS, e.g., changes in growth rate, cell morphology, production of stress fibres and p38 activation.
Morphologically, fibroblasts from other genome instability syndromes show similarities to WS in that they are enlarged and granular displaying distinct stress fibres. Of particular note is the fenestrated appearance seen in many of the cells, most prominently in NBS1 and CSA. Treatment with p38 inhibitors, such as SB203580, BIRB796 and VX745, show varying effects in the different syndromes, e.g. they dramatically rescue the senescent-like and stress fibre phenotypes of NBS1 fibroblasts, but have little effect on Bloom syndrome fibroblasts.
To date, evidence suggests that fibroblasts from all of the syndromes show aberrant physiology, suggesting that that they are undergoing some form of stress, perhaps in response to ongoing genome instability. However, preliminary studies with p38 inhibitors suggest that, either p38 is not active in all cell strains, or indeed if it is, it does not always contribute to their phenotype. Therefore, it seems that p38MAPK upregulation is specific to WS, and that while genome instability per se might trigger a stress-signalling cascade, this may not exclusively involve p38.
Figure legend
Effect of BIRB796 on RTS fibroblasts, shown by phase contrast. Untreated cells (left) and BIRB796 treated cells (right).
