1. Genomic instability in cancer and aging Jan Vijg, University of Texas Health Science Center, San Antonio, Texas

3. DNA mutations AGC TCG Point mutation Transposition Deletion/Insertion Inversion

4. Role of genome instability in aging Evolutionary logic of genome instability Evidence for genome deterioration in aging Association of DNA repair defects with accelerated aging

5. Effect of deleterious mutations on fitness in E. coli Number of mutations ln mean fitness Elena & Lenski, 1997, Nature 390: 395

7. “The problem in this theory is that of developing a quantitative measure of mutations in somatic cells” Howard J. Curtis, 1963

8. LacZ plasmid recovery

9. Spontaneous mutant frequencies with age in heart and small intestine

10. Spontaneous mutant frequencies with age in mouse organs

11. Mutant spectra in vivo and in vitro

12. Point mutational spectra in organs from Young (3m) and Old (32m) mice Mutant frequency (x10 -5 ) Percentage BrainHeartLiverSpleenSmall intestineMock recovery Lymphoma G:C  A:TG:C  T:A G:C  C:G A:T  n:n Del(-1) G:C  A:TG:C  T:A G:C  C:G A:T  n:n Del(-1) G:C  A:TG:C  T:A G:C  C:G A:T  n:n Del(-1) G:C  A:TG:C  T:A G:C  C:G A:T  n:n Del(-1) G:C  A:TG:C  T:A G:C  C:G A:T  n:n Del(-1) G:C  A:T G:C  T:A G:C  C:G A:T  n:n Del(-1) Mutant frequency (x10 -5 ) Percentage G:C  AT at CpG sites Del (-1) at reiterated sites, i.e. a sequence of 3 or more of the same nucleotide O Y

13. Mutant frequencies in vivo and in vitro

14. MEF 6 Growth curve Mutant frequency spectra 3% 20% Total MF Point Mut Size change

15. Point mutations in MEFs at high and low oxygen

16. Genome maintenance and aging Growth & Reproduction Increased Longevity Somatic Maintenance Cancer Aging DNA Damage Apoptosis Cell Senescence Transcriptional Interference Mutation Altered Chromatin

17. Aging in DNA repair-deficient mice Mutant geneDefectLife span (weeks)Aging Phenotype Xpa KO Ercc1 KO Ercc1 HP Complete NER NER and crosslink repair Same 100 3-4 25 None Weight reduction, renal and liver failure, cachexia, sarcopenia, kyphosis, neuronal degeneration Same

18. Ratio of mRNA expression levels in old v/s young livers (normalized) 72 Arrays 6 Old, 6 Young Animals M = log2(red/green) A = 1/2 log2(red*green) Total Intensity Ratio of Intensity

19. Ratio of mRNA expression levels in Ercc1 -/- knockout v/s wildtype liver M = log2(red/green) A = 1/2 log2(red*green) Over-expressed genes* Under-expressed genes* Significant genes that Overlap with aging* *significant by SAM

20. Annotation of genes co-expressed in aging and Ercc1 in liver Function AgingErcc1 Immune Response Liver Regeneration Genotoxic Response Peroxisome Proliferators DNA Cross Linkers Oxidative Stress

21. Mutation accumulation in liver of Ercc1 and Xpa mutant mice 24 weeks52 weeks

22. Point mutations in liver of 1-year old Xpa null mice Mutant Frequency (x10 -5 )

23. Point mutations in liver of 5mo Ercc1 null mice Mutant Frequency (x10 -5 )

24. Conclusions Different types of mutations accumulate with age in an organ-specific manner Liver-specific gene expression profile of mice with defects in the repair of double-strand lesions, i.e., Ercc1, is similar to normal aging DNA repair defects cause shifts in the spectrum of age-accumulating mutations Genome rearrangements appear to be associated more with aging than with cancer

26. Acknowledgements Mutation Analysis Rita Busuttil Martijn Dollé Wendy Snyder Microarray Felix Calderon Debbie Muñoz Prakash Nair Bioinformatics Brent Calder John David Garza Paul Lohman Accelerated Genomics Mangkey Bounpheng Nathalie van Orsouw Erasmus University Jan Hoeijmakers Laura Niedernhofer Dana-Farber Cancer Institute Frederick Li Seoul National University Yousin Suh RIVM Harry van Steeg LBNL Judy Campisi