Long-chain fatty acid oxidation and glutaminolysis are dispensable for most hematopoietic lineages. (A) Schematic representation of relevant metabolic pathways. Pyruvate enters the mitochondria through the MPC, composed of subunits MPC1 and MPC2. Long-chain fatty acids enter the mitochondria via carnitine palmitoyltransferase 1 and 2 (CPT1 and CPT2) and carnitine acylcarnitine translocase (CAT). Within the mitochondria, GLS hydrolyzes glutamine into glutamate. (B) Schematic representation of mixed bone marrow chimera experiments to assess hematopoietic requirements for Cpt2 control (Cpt2^fl/fl; ROSA26 CreER^−/−) or KO (Cpt2^fl/fl; ROSA26 CreER^+/+) cells and Gls control (Gls^fl/fl; ROSA26 CreER^−/−) or KO (Gls^fl/fl; ROSA26 CreER^+/+) cells. (C) CD45.2 chimerism was normalized to either pre-tamoxifen peripheral blood chimerism levels for the spleen or HSC chimerism for the bone marrow of Cpt2 chimeras. Data are shown for B cells, DCs, Gr1^hi and Gr1^lo myeloid cells, NK cells, and T cells for the spleen. Data are shown for HSCs, MPPs, CMPs, GMPs, MEPs, and CLPs in the bone marrow. Each symbol represents an individual mouse. Mean values ± SEM are shown, and data are pooled from three independent experiments. P values >0.05 by two-way ANOVA with post-hoc Tukey’s multiple comparisons test are not depicted. (D) CD45.2⁺ GFP⁻ chimerism was normalized as stated in Fig. 1 C for Gls chimeras. Cell populations assessed are the same as in Fig. 1 C with the addition of platelets in the blood. Each symbol represents an individual mouse. Mean values ± SEM are shown, and data are pooled from two independent experiments. *, P < 0.05 by two-way ANOVA with post-hoc Tukey’s multiple comparisons test.