3.7. EXO1-association replicated in female French centenarians

The functional results prompted us to focus the genetic replication of the association between EXO1 and longevity on the promoter SNP rs1776180. Since the effect of this functionally relevant polymorphism was apparent only in German women (Table 3 and Fig. 2B), we just considered females for validation. We analyzed rs1776180 in a collection of 455 French female centenarians and 109 younger individuals whose age range was comparable to that of the German controls. In this French sample, the APOE-adjusted analysis confirmed the association signal (P = 0.044) (Table 4). The French allele and genotype APOE frequencies were similar to the original findings in the Germans and an enrichment of the C allele in the centenarians compared to the controls was apparent in both populations. Therefore, the combined genetic and functional analyses point to a pivotal role of rs1776180 in the allele-specific regulation of EXO1, which in turn influences the ability to survive to exceptional old age.

An important role of EXO1 in controlling life expectancy has previously been reported in a murine model with a genetic ablation of the nuclease domain of the gene (Exo1(cid:3)/(cid:3)). The respective animals were viable, but their lifespan was significantly shortened (Schaetzlein et al., 2007; Wei et al., 2003). A first report of the Exo1(cid:3)/(cid:3) knock-out mouse model on a mixed genetic background suggested that the beneficial influence of an increased Exo1 activity on lifespan could lie in mutation avoidance and, therefore, in the prevention of tumorigenesis (Wei et al., 2003). However, a recent study has shown that congenized mice carrying the same Exo1 inactivation on a pure C57BL/6J background displayed a considerably shorter survival without a significant

Federal Ministry of Science and Education (BMBF) through an Explorative Project (EP-S23T01) of the National Genome Research Network (NGFN) and through the Pathway Mapping Project (01GS0426) and (3) the French Ministe`re de l’Enseignement Supe´rieur et de la Recherche.

rise in cancer formation, indicating that other factors contribute to the lifespan reduction in Exo1(cid:3)/(cid:3) mice (Schaetzlein et al., 2007). In view of the similarly uncertain role of EXO1 in human carcinogen- esis (reviewed by Liberti and Rasmussen, 2004), the possible links between EXO1 variation, cancer and life expectancy warrant further studies.

Our data implicate E47 as a repressor of EXO1 transcription. The E47protein(encodedbyTCF3/E2A)belongstotheE-Boxsubfamilyof basic helix–loop–helix (bHLH) transcription factors and is involved in the regulation of cell lineage commitment, growth and differentiation (Slattery et al., 2008). A recent study using a TCF3/ E2A-deficient murine lymphoma cell line and hierarchical cluster analysis of DNA microarray data identified E47-dependent gene expression as contributing to cell cycle progression, lipid metabo- lisms, stress response and cell survival (Schwartz et al., 2006). Interestingly, there are several lines of evidence linking E47 to aging processes.E47expressionlevel andactivityarediminishedinspleen and bone marrow of aged mice, compared to young animals (Frasca et al., 2003). In addition, impaired E47 activity has been shown to correlatewithareducedimmunoglobulinclassswitchinsenescingB cells (Frasca et al., 2004). The contribution of E47 to cellular aging is further emphasized by the finding that gene silencing of E47 delays the onset of cellular senescence in human fibroblasts (Zheng et al., 2004). The regulatory crosstalk of E47 and EXO1 observed in this study implies that the concomitant expression of both genes is required for the C allele of rs1776180tobe advantageous. Strikingly, our data reveal an overlap of their expression profiles in a number of tissues,includingliver,thymus, spleen,testesandovary.SinceE-Box proteins(e.g.E47,E12,HEN)arewellknowntoformhomo-aswellas heterodimeric complexes and to interact with other bHLH tran- scriptionfactors(e.g.TAL1,MyoD)(Hsuetal.,1994;Lluisetal.,2005), the complex role of E-Box proteins and interaction partners with regulatory function for human aging awaits further investigation. The pivotal future challenge, however, will be to clarify how increased tissue-specific EXO1 expression contributes to prolonged survival.

Acknowledgments

For excellent technical assistance, we wish to acknowledge the laboratory personnel of the Institute of Clinical Molecular Biology, the members of the Popgen Biobank as well as the staff of the Biological Resource Center of the Fondation Jean Dausset-CEPH. The authors are grateful to Laura Maringele (Newcastle upon Tyne, UK), Richard Kolodner (La Jolla, USA) and Thomas Manke (Berlin, Germany) for helpful discussion.

Appendix A. Supplementary data

Supplementary data associated with this article can be found, in the online version, at doi:

/j.mad.

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