Host membrane glycosphingolipids and lipid microdomains facilitate Histoplasma capsulatum internalisation by macrophages

Recognition and internalisation of intracellular pathogens by host cells is a multifactorial process, involving both stable and transient interactions. The plasticity of the host cell plasma membrane is fundamental in this infectious process. Here, the participation of macrophage lipid microdomains during adhesion and internalisation of the fungal pathogen Histoplasma capsulatum (Hc) was investigated. An increase in membrane lateral organisation, which is a characteristic of lipid microdomains, was observed during the first steps of Hc–macrophage interaction. Cholesterol enrichment in macrophage membranes around Hc contact regions and reduced levels of Hc–macrophage association after cholesterol removal also suggested the participation of lipid microdomains during Hc–macrophage interaction. Using optical tweezers to study cell-to-cell interactions, we showed that cholesterol depletion increased the time required for Hc adhesion. Additionally, fungal internalisation was significantly reduced under these conditions. Moreover, macrophages treated with the ceramide-glucosyltransferase inhibitor (P4r) and macrophages with altered ganglioside synthesis (from B4galnt1 −/− mice) showed a deficient ability to interact with Hc. Coincubation of oligo-GM1 and treatment with Cholera toxin Subunit B, which recognises the ganglioside GM1, also reduced Hc association. Although purified GM1 did not alter Hc binding, treatment with P4 significantly increased the time required for Hc binding to macrophages. The content of CD18 was displaced from lipid microdomains in B4galnt1 −/− macrophages. In addition, macrophages with reduced CD18 expression (CD18 low ) were associated with Hc at levels similar to wild-type cells. Finally, CD11b and CD18 colocalised with GM1 during Hc–macrophage interaction. Our results indicate that lipid rafts and particularly complex gangliosides that reside in lipid rafts stabilise Hc–macrophage adhesion and mediate efficient internalisation during histoplasmosis.Fil: Guimarães, Allan J.. Universidade Federal Fluminense; Brasil. Yeshiva University; Estados UnidosFil: de Cerqueira, Mariana Duarte. Universidade Federal do Rio de Janeiro; BrasilFil: Zamith-Miranda, Daniel. Universidade Federal do Rio de Janeiro; BrasilFil: Lopez, Pablo. The Johns Hopkins School Of Medicine; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Universidad Nacional de Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra; ArgentinaFil: Rodrigues, Marcio L.. Fundación Oswaldo Cruz; Brasil. Universidade Federal do Rio de Janeiro; BrasilFil: Pontes, Bruno. Universidade Federal do Rio de Janeiro; BrasilFil: Viana, Nathan B.. Universidade Federal do Rio de Janeiro; BrasilFil: DeLeon-Rodriguez, Carlos M.. Johns Hopkins Bloomberg School Of Public Health; Estados UnidosFil: Rossi, Diego Conrado Pereira. Johns Hopkins Bloomberg School Of Public Health; Estados UnidosFil: Casadevall, Arturo. Johns Hopkins Bloomberg School Of Public Health; Estados UnidosFil: Gomes, Andre M.O.. Universidade Federal do Rio de Janeiro; BrasilFil: Martinez, Luis R.. University of Texas at El Paso; Estados UnidosFil: Schnaar, Ronald L.. The Johns Hopkins School Of Medicine; Estados UnidosFil: Nosanchuk, Joshua D.. Yeshiva University; Estados UnidosFil: Nimrichter, Leonardo. Universidade Federal do Rio de Janeiro; Brasi