Title: Elevated SGPL1 expression increases metabolic rate and reduces survival in adrenocortical carcinoma patients

Sphingosine-1-phosphate lyase (SGPL1) catalyses the final step in sphingolipid metabolism (Figure 1), irreversibly degrading the lipid signalling molecule sphingosine-1-phosphate (S1P). Loss-of-function mutations in SGPL1 cause a spectrum of disorders, including primary adrenal insufficiency. Given loss of SGPL1 causes reduced steroidogenesis, while increased steroidogenesis is a hallmark of aggressive adrenocortical carcinomas (ACCs), we hypothesise increased SGPL1 expression might regulate ACCs. We identified a potential novel biomarker in ACCs, whereby increased SGPL1 expression correlates with reduced patient survival. We also saw similar correlation with other sphingolipid enzymes which diminish the ceramide pool, while those that replenish it confer a beneficial patient outcome. Increased SGPL1 expression in a cell model increased their resting proliferation and migration, although no effect on apoptosis was seen. There was a global decrease in the sphingolipid pool, possibly indicating increased flux through this pathway. RNAseq analysis identified an increase in expression of oxidative phosphorylation genes, and this correlated with an increase in basal and maximal glycolysis and oxidative phosphorylation. Finally, we showed that the increase in proliferation in the OE cells is dependent on the presence of metabolic substrates which may be used for anabolism and/or the production of ATP. We generated SGPL1 overexpressing H295R cells, as well as SGPL1 KO H295R cells and compared their transcriptomes by RNAseq. We used 3 biological replicates per group.