Human longevity and variation in DNA damage response and repair: a study of the contribution of sub-processes using competitive gene-set analysis.

Birgit Debrabant*,1,8, Mette Sørensen*,1,2,8, Friederike Flachsbart3, Serena Dato4, Jonas Mengel-From1,2, Tinna Stevnsner5, Vilhelm A Bohr5,6, Torben A Kruse2, Stefan Schreiber3, Almut Nebel3, Kaare Christensen1,2,7, Qihua Tan1,2 and Lene Christiansen1,2.

DNA damage response and repair are crucial to maintaining genetic stability and are consequently considered central to aging and longevity. Here we investigate whether this pathway generally associates with longevity and whether specific sub-processes are more strongly associated with longevity than others. Data were applied to 592 SNPs from 77 genes involved in nine sub-processes: DNA damage response, base excision repair (BER), nucleotide excision repair, mismatch repair, non-homologous end-joining, homologous recombinational repair (HRR), RecQ helicase activities (RECQ), telomere functioning and mitochondrial DNA processes. The study population consisted of 1089 long-lived and 736 middle-aged Danes. A self-contained set-based test of all SNPs displayed association with longevity (P-value 9.9 x 10^-5), supporting the idea that the overall pathway could affect longevity. Investigation of the nine sub-processes using the competitive gene-set analysis by Wang et al indicated that BER, HRR and RECQ were more strongly associated with longevity than the respective remaining genes of the pathway (P-values 0.004-0.048). For HRR and RECQ only one gene contributed to the significance whereas for BER several genes contributed. These associations did not, however, generally pass correction for multiple testing. Still, these findings indicate that of the entire pathway, variation in BER might influence longevity the most. These modest-sized P-values were not replicated in a German sample. This might, though, be due to differences in genotyping procedures and investigated SNPs, potentially inducing differences in the coverage of gene regions. Specifically, five genes were not covered at all in the German data. Therefore, investigations in additional study populations are needed before a final conclusion can be drawn.

European Journal of Human Genetics advance online publication, 12 February 2014; doi:10.1038/ejhg.2013.299

Keywords: competitive gene-set analysis; human longevity; SNPs; case-control data; DNA damage response and repair

INTRODUCTION

It has been estimated that about 25% of the variation in human lifespan is caused by genetic factors1, a contribution which is likely to be minimal before age 60 years and most pronounced from age 85 years onwards.2 Candidate longevity genes encode proteins involved in several biological processes including DNA damage response and DNA repair,3 which are essential for cellular function due to the constant exposure of living cells to various kinds of endogenous and exogenous agents. Such agents can damage diverse cellular components, including the DNA, and potentially induce genomic instability or mutagenesis. To maintain the integrity of the genome, DNA damage response processes detect the DNA lesions and activate cell cycle arrest and DNA repair processes.4 The DNA repair processes each repair a certain subset of DNA lesions: base excision repair removes single-base lesions (eg oxidative lesions induced by reactive oxygen species (ROS)); nucleotide excision repair operates on bulky lesions (eg induced by UV light); mismatch repair among other repair errors arising during DNA replication while double-strand break repair (DSB) removes DSB breaks and complex lesions (eg induced by ionizing radiation or chemotherapeutics). DSB is divided into homologous recombinational repair and non-homologous end-joining. The RecQ helicases are, among other things, important for ensuring the proper structure of the DNA helix during DNA repair.8 Severe mutations in two of the RecQ helicase genes, WRN and RECQL4, can cause Werner and Rothmund-Thomson premature aging syndromes, respectively.9,10 Additional mechanisms affect the stability of the chromosome ends, the telomeres. Telomeres undergo shortening with every replication cycle in cells lacking telomerase until the telomeres reach a critical length at which the cell enters cellular senescence.11,12 Telomere shortening is associated with organismal aging; in humans, leukocyte telomere length is inversely related to age13,14 and is associated with an increased risk of age-related disease and with higher mortality.15-18 Finally, mitochondrial DNA (mtDNA) is exposed to specifically high levels of ROS due to its vicinity to oxidative phosphorylation and is thus potentially exposed toresponse and repair genes is associated with human longevity. We used a combined analysis of sets of SNPs and genes to reduce the multiple testing burden and increase power. DNA was isolated from blood spot cards or full blood using QIAamp DNA Kits, and genotyping of all samples was performed on the Illumina GoldenGate platform. Data cleaning was done to exclude samples and SNPs with low call rates, close clusters, clusters of low intensity, heterozygote cluster shifting towards a homozygote cluster, excess heterozygosity, and SNPs located on the X chromosome. Overall, 8.8% of the individuals and 10.4% of the SNPs were excluded, leaving data on 592 SNPs in 77 genes from 1089 oldest-old and 736 controls.

The study focused on common variation in 77 DNA-damage response and repair genes composing nine sub-processes. The association of the overall pathway with longevity was investigated first, followed by a gene-set method put forward by Wang et al to test whether any of the sub-processes is more associated with longevity than the other sub-processes. The analyses were carried out in a study population of 1089 oldest-old and 736 middle-aged Danes, whereas replication was done in a German study population of 763 long-lived individuals and 1085 controls.