Figure 3. PGE2 regulates mitochondrial membrane potential through changes in the malate-aspartate shuttle (MAS).

(A-B) Real time contribution of ETC complexes to Δψ_m in BMMs stimulatd as indicated. Changes in Δψ_m after sequential treatment with rotenone (Rot), succinate (Succ) and antimycin (Ant) (A), or in response to oligomycin (B).

(C) Δψ_m in BMMs stimulated and treated with α-amanitin (α-A) as indicated for 24 h. Mean ± SEM from 6 biological replicates.

(D) Mitochondrial solute carrier gene expression in M(IL-4)+PGE2 stimulated for 6 h compared to M(IL-4).

(E) Schematic of the MAS.

(F) MAS gene expression in BMMs stimulated for 6 h as indicated. Statistically significant (adjusted p value < 0.1) up or down regulated (> 2 fold) genes from 3 biological replicates denoted by *.

(G) LC-MS quantitation of MAS metabolites in BMMs stimulated as indicated for 6 h or 24 h, normalized to M(−) (dotted line). Mean ± SEM from 3 biological replicates.

(H-I) Schematic (H) and results of using GC-MS to trace deuterium (green, H) in BMMs cultured in singly deuterated glucose (²H-Glc) into malate and lactate (I), plus depiction of stimulation strategy.

(J) Δψ_m in BMMs transduced with a control or Got1 targeting shRNA after 24 h stimulation with IL-4 +/− PGE2 as indicated. Means ± SEM from 6–7 biological replicates.

Data in A-C, G, I and J are representative of 3 independent experiments. (C, G, I-J) p*<0.05, p**<0.005, p***<0.0005, p****<0.0001. See also Figure S3.