Figure 2. Azaserine Inhibits Increased De Novo Purine Synthesis in SIRT3 KO MEFs.

(A) Table of the metabolic pathways significantly affected by 6 hr treatment with 30 μM azaserine in SIRT3 KO MEFs. The pathway analysis module in MetaboAnalyst 3.0 was used for the analysis (see also Figures S2A–S2G and Table S2).

(B) Schematic of purine metabolism. Shaded in pink is the de novo purine synthesis pathway, and shaded in blue are intermediates in purine degradation or the salvage pathway. Blue arrows represent purine salvage pathways.

(C) Heatmap comparing metabolite levels in the presence of azaserine (30 μM for 6 hr) in WT and SIRT3 KO MEFs (n = 4) or treated with DMSO (control). Red indicates upregulation and blue indicates downregulation. Asterisk denotes statistically altered metabolites comparing SIRT3 KO with SIRT3 KO + azaserine.

(D and E) Relative levels of carboxyaminoimidazole ribonucleotide (CAIR) (D) and 5-MeTHF (E) in WT and SIRT3 KO MEFs after treatment with 30 μM azaserine for 6 hr.

(F and G) Incorporation of ¹⁴C-glycine (F) or ¹⁴C-aspartate (G) into DNA in WT and SIRT3 KO cells in the presence or absence of 30 mM azaserine for 8 hr (see also Figures S2H–S2J).

(H) Growth of WT and SIRT3 KO MEFs with PPAT knockdown relative to control cells and to WT cells with the corresponding PPAT shRNA (see also Figure S2K).

(I) Relative growth of WT and SIRT3 KO MEFs treated as specified.

Data in this figure are represented as mean ± SEM. *p ≤ 0.05, **p ≤ 0.01, and ***p ≤ 0.001; n.s., not significant.