PLA2G15

The epidermis contains many structurally diverse ceramides, which form the skin permeability barrier (skin barrier). Mutations in genes involved in the synthesis of ω-O-acylceramides (acylceramides) and protein-bound ceramides cause ichthyosis.

We aimed to elucidate the relationship between the degree of skin barrier impairment and changes in epidermal ceramide profiles caused by mutations in acylceramide synthesis genes.

Knockout (KO) mice of three genes-fatty acid (FA) ω-hydroxylase Cyp4f39 (human CYP4F22 ortholog), FA elongase Elovl1, and acyl-CoA synthetase Fatp4-were subjected to transepidermal water loss measurement, toluidine blue staining, and epidermal ceramide profiling via liquid chromatography coupled with tandem mass spectrometry.

Transepidermal water loss was highest in Cyp4f39 KO mice, followed by Elovl1 KO and Fatp4 KO mice, and Cyp4f39 KO mice also showed the strongest degree of toluidine blue staining. In Cyp4f39 KO, Elovl1 KO, and Fatp4 KO mice, acylceramide levels were 0.6%, 1.6%, and 12%, respectively, of those in wild-type mice. Protein-bound ceramide levels were 0.2%, 30%, and 33%, respectively, of those in wild-type mice. We also observed a near-complete absence of ω-hydroxy ceramides in Cyp4f39 KO mice, reduced total ceramide levels and shortened FA moieties in Elovl1 KO mice, and increased hydroxylated ceramide levels and slightly shortened FA moieties in Fatp4 KO mice.

The degree of reduction in protein-bound ceramide levels is probably related to the severity of skin barrier defects in these three strains. However, reduced acylceramide levels and other changes in ceramide composition unique to each KO strain are also involved.

Cold exposure is an environmental stress that elicits a rapid metabolic shift in endotherms and is required for survival. The liver provides metabolic flexibility through its ability to rewire lipid metabolism to respond to an increased demand in energy for thermogenesis. We leveraged cold exposure to identify novel lipids contributing to energy homeostasis and found that lysosomal bis(monoacylglycero)phosphate (BMP) lipids were significantly increased in the liver during acute cold exposure. BMP lipid changes occurred independently of lysosomal abundance but were dependent on the lysosomal transcriptional regulator transcription factor EB (TFEB). Knockdown of TFEB in hepatocytes decreased BMP lipid levels. Through molecular biology and biochemical assays, we found that TFEB regulates lipid catabolism during cold exposure and that TFEB knockdown mice were cold intolerant. To identify how TFEB regulates BMP lipid levels, we used a combinatorial approach to identify TFEB target Pla2g15 , a lysosomal phospholipase, as capable of degrading BMP lipids in in vitro liposome assays. Knockdown of Pla2g15 in hepatocytes led to a decrease in BMP lipid species. Together, our studies uncover a required role of TFEB in mediating lipid liver remodeling during cold exposure and identified Pla2g15 as an enzyme that regulates BMP lipid catabolism.

The mesenteric lymph nodes (MLN) function as a barrier to systemic spread for both commensal and pathogenic bacteria in the gut. Listeria monocytogenes, a facultative intracellular foodborne pathogen, readily overcomes this barrier and spreads into the bloodstream, causing life-threatening systemic infections. We show here that intracellular replication protected L. monocytogenes from clearance by monocytes and neutrophils and promoted colonization of the small intestine-draining MLN (sMLN) but was not required for dissemination to the colon-draining MLN (cMLN). Intestinal tissue had enough free lipoate to support LplA2-dependent extracellular growth of L. monocytogenes, but exogenous lipoate in the MLN was severely limited, and so the bacteria could replicate only inside cells, where they used LplA1 to scavenge lipoate from host peptides. When foodborne infection was manipulated to allow ΔlplA1 L. monocytogenes to colonize the MLN to the same extent as wild-type bacteria, the mutant was still never recovered in the spleen or liver of any animal. We found that intracellular replication in the MLN promoted actin-based motility and cell-to-cell spread of L. monocytogenes and that rapid efficient exit from the MLN was actA dependent. We conclude that intracellular replication of L. monocytogenes in intestinal tissues is not essential and serves primarily to amplify bacterial burdens above a critical threshold needed to efficiently colonize the cMLN. In contrast, intracellular replication in the MLN is absolutely required for further systemic spread and serves primarily to promote ActA-mediated cell-to-cell spread.