Mass spectrometry-based profiling of phospholipids and sphingolipids in extracts from Saccharomyces cerevisiae

Lipids are rapidly moving to centre stage in many fields of biological sciences. Lipidomics, the systems-level scale analysis of lipids and their interacting factors, is thus an emerging field which holds great promise for drug and biomarker discovery. Here we present a mass spectrometry-based approach for profiling of polar lipids, in particular phospholipids and sphingolipids, in Saccharomyces cerevisiae. The first step includes semi-quantitative surveys of lipids in an untargeted fashion, which is particularly powerful for detection of changes that cannot easily be anticipated. This leads to the identification of ions with increased or decreased signal intensities. Comprehensive theoretical calculation of the masses of yeast phospholipid and sphingolipid molecular species, based on fatty acyl and headgroup heterogeneity, is next used to tentatively assign ions of interest. Subsequent targeted analysis using tandem mass spectrometry allows for characterization and quantification of phospholipids and sphingolipids. Given the high degree of conservation in pathways of lipid metabolism between different organisms, it can be expected that this method will lead to the discovery of novel enzymatic activities and modulators of known ones, particularly when used in combination with genetic and chemogenetic libraries and screens. We validated the method using the EUROSCARF library of non-essential deletion mutants. Mutants of SCS7, a lipid hydroxylase, and SLC1, a putative acyl transferase with unknown substrate specificity, were profiled for their phospholipid and sphingolipid content. The observed changes in lipid profiles are consistent with previous observations and extend our knowledge on in vivo substrate use under permissive growth conditions.