Supplementary Table S1. Primer pairs used for qPCR. F, forward; R, reverse.

Gene / Primers
Peroxiredoxin1
Peroxiredoxin4
SOD2
Catalase
GSH-Px
Foxo1
Foxo3a
Sestrin1
Sestrin 2
Sestrin 3
p53
p21
actin
hsod2
hFoxo1
hFoxo3a / F,5′-ACACCCAAGAAACAAGGAGGATT-3′
R, 5′-CAACGGGAAGATCGTTTATTGTTA-3′
F 5′-TCCTGTTGCGGACCGAAT-3′
R 5′-GATCTTGGCTTTGCTTAGATGCA-3′
F 5′-ATTAACGCGCAGATCATGCA-3
R 5′-TGTCCCCCACCATTGAACTT-3′
F 5′-GCGTCCAGTGCGCTGTAGA-3′
R 5′-TCAGGGTGGACGTCAGTGAA-3′
F 5′-GGGACTACACCGAGATGAACGA-3′
R 5′-ACCATTCACTTCGCACTTCTCA-3′
F 5′-TACGGAGGATTGAACCAGTA-3′
R 5′-CTGGCATGACCGAATTAGGG-3′
F 5′-TCAGAATGAAGGCACGGGCA-3′
R 5′-TGGAGAGCTGGGAAGGACTG-3′
F 5′-GGACGAGGAACTTGGAATCA-3′
R 5′-ATGCATCTGTGCGTCTTCAC-3′
F 5′-TAGCCTGCAGCCTCACCTAT-3′
R 5′-TATCTGATGCCAAAGACGCA-3′
F 5′-CATGCGTTTCCTCACTCAGA-3′
R 5′-GGCAAAGTCTTCGTACCCAA-3′
F 5′- CTCACTCCAGCTACCTGAAGA -3′
R 5′-AGAGGCAGTCAGTCAGTCTGAGTCA -3′
F 5′- AGTGTGCCGTTGTCTCTTCG-3′
R 5′- ACACCAGAGTGCAAGACAGC-3′
F 5′-GATCATTGCTCCTCCTGAGC-3′
R 5′-AGTCCGCCTAGAAGCACTTG-3′
F 5′-AATTGCTGCTTGTCCAAATCA-3′
R 5′-TCCCCAGCAGTGGAATAAGG-3′
F 5′- GGCGGGCTGGAAGAATTCAA-3’
R 5′-TGTTGTCCATGGATGCAGCTC-3’
F 5′-CCCAGCCTAACCAGGGAAGT-3′
R 5′-AGCGCCCTGGGTTTGG-3′

Supplementary Figure Legends

Figure S1. (a) Survival data for Casp2+/+and Casp2-/-animals. (b) Survival curve for the last 50% survivors of each genotype. Data was analysed using Fishers exact test.

Figure S2. Histological analysis of skin.(a)Haematoxylin and Eosin stainedskin sections from 24 month old male and female,Casp2+/+and Casp2-/-mice. (b)Quantification of the thickness (m) of the dermis, subcutaneous adipose tissue (SAT) and muscle layer from 24 month old female and male mice. In every section, five measurements of the thickness of each layer were measured at different randomly selected positions and the median thickness of n=4 sections for each genotype were calculated. Scale bar=100m. ap<0.05.

Figure S3. (a) Lipid Peroxidation. (b) Protein oxidation levels. (c) SOD enzyme activity. (d) GSH-Px enzyme activity in young Casp2+/+and Casp2-/- mice liver. Values are Mean ± SEM for 6-7 animals per group. ap<0.05, bp<0.01

Figure S4. Western blots from age-matched young (Y) and old (O) Casp2 +/+and Casp2-/- mice showing higher p53 expression in kidney and spleen.

Figure S5. (a) A western blot showing caspase-2 knockdown in IMR90 E1A cells. (b) FoxO3a expression decreases after caspase-2 knockdown while its target gene SOD2 expression remained unaltered.

Figure S6.Assessment of mitochondrial dysfunction in Casp2+/+and Casp2-/- primary MEFS. (a and b) Mitochondrial membrane potential was assessed by JC1 staining in untreated or menadione treated (10µM for 6 h), Casp2+/+ andCasp2-/-MEFs. (b) The percentage of cells with green fluorescence (measure of decreased mitochondrial membrane permeability (Ψm) was determined, showing no difference in Casp2+/+ andCasp2-/-MEFs. All cells expressed some level of red fluorescence indicating the presence of healthy functional mitochondria. (c) No change was observed in the levels of intracellular ATP in untreated and menadione (10 µM for 6 h) treated primary Casp2+/+ andCasp2-/-MEFs.

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