Phosphatidylserine synthesis is essential for viability of the human fungal pathogen Cryptococcus neoformans

Abstract

Phospholipids are an integral part of the cellular membrane structure and can be produced by a de novo biosynthetic pathway and, alternatively, by the Kennedy pathway. Studies in several yeast species have shown that the phospholipid phosphatidylserine (PS) is synthesized from CDP-diacylglycerol and serine, a route that is different from its synthesis in mammalian cells, involving a base-exchange reaction from preexisting phospholipids. Fungal-specific PS synthesis has been shown to play an important role in fungal virulence and has been proposed as an attractive drug target. However, PS synthase, which catalyzes this reaction, has not been studied in the human fungal pathogen Cryptococcus neoformans. Here, we identified and characterized the PS synthase homolog (Cn Cho1) in this fungus. Heterologous expression of Cn CHO1 in a Saccharomyces cerevisiae cho1⌬ mutant rescued the mutant's growth defect in the absence of ethanolamine supplementation. Moreover, an Sc cho1⌬ mutant expressing Cn CHO1 had PS synthase activity, confirming that the Cn CHO1 encodes PS synthase. We also found that PS synthase in C. neoformans is localized to the endoplasmic reticulum and that it is essential for mitochondrial function and cell viability. Of note, its deficiency could not be complemented by ethanolamine or choline supplementation for the synthesis of phosphatidylethanolamine (PE) or phosphatidylcholine (PC) via the Kennedy pathway. These findings improve our understanding of phospholipid synthesis in a pathogenic fungus and indicate that PS synthase may be a useful target for antifungal drugs. 2 The abbreviations used are:

Crystal structures of phosphatidyl serine synthase PSS reveal the catalytic mechanism of CDP-DAG alcohol O-phosphatidyl transferases

“…PS is an important molecule for all kinds of cells. Not only does it serve as a structural component for cellular lipid membranes or as precursor for other phospholipids such as PE, or PC, but it is also essential for signal transduction, cellular metabolism and is essential for viability of many human pathogens 36 , 43 . Enzymes that synthesize, convert or regulate PS are functionally conserved between bacteria, archaea, and eukaryotes.…”

Molecular Basis of the Anticancer and Antibacterial Properties of CecropinXJ Peptide: An In Silico Study

“…While not interacting with models of mammalian bilayers composed of PC and cholesterol, CXJ is able to specifically recognize PS and PE headgroups, both characterizing the outer leaflet of cancer membranes. These phospholipids play an important role in resistant mechanisms of fungi such as Cryptococcus neoformans or Candida albicans [ 100 , 101 , 161 , 162 ] and in the virulence of parasites such as Plasmodium [ 161 ] or other intracellular pathogens such as Brucella [ 163 ]. CXJ specifically interacts with the carboxylate atoms of PS by means of the side chain NH 2 groups of arginine 1, 13 and 16.…”

Mapping the Substrate-Binding Sites in the Phosphatidylserine Synthase in Candida albicans

“…and was identified to be almost invariably conserved in numerous other lipid biosynthetic enzymes, including those of Gram positive and Gram negative bacteria, archaea, fungi, plants, and mammals (Dryden and Dowhan, 1996;Tanaka et al, 1996;Matsumoto, 1997;Delhaize et al, 1999;Matsuo et al, 2007;Grave et al, 2019;Konarzewska et al, 2019). The CAPT motif in yeast Cpt1 was characterized by alanine substitution mutagenesis, providing information on the importance of specific residues within the conserved motif (Williams and McMaster, 1998).…”

“…The phosphatidylserine (PS) synthase in C. albicans represents a potential drug target for three reasons: 1) It is required for virulence ( Chen et al., 2010 ), indicating that inhibitors of this enzyme would render the organism incapable of causing infection; 2) it is absent in humans ( Braun et al., 2005 ), so inhibitors potentially would have no toxic side effects for the host; and 3) it is conserved among many fungi, so a drug could potentially be effective against multiple fungal pathogens. Recently, deletion of PS synthase in Cryptococcus neoformans was shown to be lethal, suggesting phosphatidylserine synthesis is also essential for the viability of some fungi ( Konarzewska et al., 2019 ). This observation further strengthens the potential of PS synthase as a broad antifungal drug target.…”

“…All of these enzymes bind a CDP-linked molecule and a second small alcohol and catalyze the formation of a phosphodiester bond between the two. Since these initial studies, this CDP-alcohol phosphatidyltransferase (CAPT) motif was narrowed to D-G-(X) 2 -A-R-(X) 8 -G-(X) 3 -D-(X) 3 -D, and was identified to be almost invariably conserved in numerous other lipid biosynthetic enzymes, including those of Gram positive and Gram negative bacteria, archaea, fungi, plants, and mammals ( Dryden and Dowhan, 1996 ; Tanaka et al., 1996 ; Matsumoto, 1997 ; Delhaize et al., 1999 ; Matsuo et al., 2007 ; Grāve et al., 2019 ; Konarzewska et al., 2019 ). The CAPT motif in yeast Cpt1 was characterized by alanine substitution mutagenesis, providing information on the importance of specific residues within the conserved motif ( Williams and McMaster, 1998 ).…”