Figure 2.

Tissue survey of mRNA and protein expression of ACSL5 and demonstration of lack of ACSL5 protein content and decreased ACSL activity in ACSL5^−/−mice. Tissues mRNA and protein were analyzed (see Methods) of 6 month old ACSL5^−/− and their littermate male ACSL5^loxP/loxP floxed control mice as generated in Figure 1. In panel A, mean mRNA fold expression of Acsl5 +/− standard error of the mean (SEM) of the indicated tissues of 4–9 mice in each group were determined by semi-quantitative real-time RT-PCR, normalized to Cyclophyllin B, and normalized again to the mean result of the proximal small intestine. Equal protein amounts (in panel B) of protein lysates were resolved by western blot with 5N-1 antibody (see Methods) for ACSL5, producing a single band at approximately 65 kDa. The membrane was reprobed with antibody for GAPDH for loading control. In C, ACSL activity presented as means ± SEM of 3–4 mice was determined by incubation of lysates with 100 μM [1-¹⁴C] palmitic acid (see Methods), *p < 0.05 ACSL comparing ACSL5^−/− (KO) and ACSL5^loxP/loxP (FL).