Role of Sphingomyelin Synthase in Controlling the Antimicrobial Activity of Neutrophils against Cryptococcus neoformans

Abstract

The key host cellular pathway(s) necessary to control the infection caused by inhalation of the environmental fungal pathogen Cryptococcus neoformans are still largely unknown. Here we have identified that the sphingolipid pathway in neutrophils is required for them to exert their killing activity on the fungus. In particular, using both pharmacological and genetic approaches, we show that inhibition of sphingomyelin synthase (SMS) activity profoundly impairs the killing ability of neutrophils by preventing the extracellular release of an antifungal factor(s). We next found that inhibition of protein kinase D (PKD), which controls vesicular sorting and secretion and is regulated by diacylglycerol (DAG) produced by SMS, totally blocks the extracellular killing activity of neutrophils against C. neoformans. The expression of SMS genes, SMS activity and the levels of the lipids regulated by SMS (namely sphingomyelin (SM) and DAG) are up-regulated during neutrophil differentiation. Finally, tissue imaging of lungs infected with C. neoformans using matrix-assisted laser desorption-ionization mass spectrometry (MALDI-MS), revealed that specific SM species are associated with neutrophil infiltration at the site of the infection. This study establishes a key role for SMS in the regulation of the killing activity of neutrophils against C. neoformans through a DAG-PKD dependent mechanism, and provides, for the first time, new insights into the protective role of host sphingolipids against a fungal infection.

Real-time in vivo imaging reveals the ability of neutrophils to remove Cryptococcus neoformans directly from the brain vasculature

“…In vitro, neutrophils are able to kill Cn via an intracellular [6,7] or extracellular killing mechanism [8]. Both oxidative [6,9] and nonoxidative [10] killing of Cn by neutrophils have been described in vitro.…”

Simultaneous Host-Pathogen Transcriptome Analysis during Granulibacter bethesdensis Infection of Neutrophils from Healthy Subjects and Patients with Chronic Granulomatous Disease

“…Class A genes (Table 3), upregulated primarily in normal PMN but not in CGD PMN, included dual-specificity phosphatase 16 (DUSP16) (increased 21-to 35-fold), which is acetylated by the Mycobacterium tuberculosis Eis protein, resulting in the suppression of certain host responses (37). Class C genes, altered primarily in CGD but not normal PMN, included several factors that might represent an attempt to control microbial growth, such as hepcidin (HAMP) (increased 3-fold), an antimicrobial peptide and hormone that decreases bioavailable iron in the circulation, and sphingomyelin synthase 2 (SGMS2) (increased 23-to 72-fold) which is important for PMN killing of Cryptococcus (38). It is unclear whether these differences reflect the continued survival of Granulibacter in CGD PMN and its killing by normal PMN or whether they reflect different host responses to Granulibacter.…”

Cryptococcus Neoformans Modulates Extracellular Killing by Neutrophils

“…We had previously observed that neutrophils obtained from immunocompetent mice having a fully functional immune system were able to kill C. neoformans (Qureshi et al, 2010). To this aim, we employed fresh murine neutrophils from Tgε26 mice, and carried out the killing assay as described in the Section “Materials and Methods”.…”