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Updated: May 25, 2026
Investigating the Phagocytosis of Leishmania using Confocal Microscopy
Published on: July 29, 2021
Trafficking and release of Leishmania metacyclic HASPB on macrophage invasion
Lorna M Maclean1, Peter J O'Toole, Meg Stark
1Centre for Immunology and Infection, Department of Biology/Hull York Medical School, University of York, York YO10 5DD, UK.
Abstract:
Proteins of the Leishmania hydrophilic acylated surface protein B (HASPB) family are only expressed in infective parasites (both extra- and intracellular stages) and, together with the peripheral membrane protein SHERP (small hydrophilic endoplasmic reticulum-associated protein), are essential for parasite differentiation (metacyclogenesis) in the sand fly vector. HASPB is a 'non-classically' secreted protein, requiring N-terminal acylation for trafficking to and exposure on the plasma membrane. Here, we use live cell imaging methods to further explore this pathway to the membrane and flagellum. Unlike HASPB trafficking in transfected mammalian cells, we find no evidence for a phosphorylation-regulated recycling pathway in metacyclic parasites. Once at the plasma membrane, HASPB18-GFP (green fluorescent protein) can undergo bidirectional movement within the inner leaflet of the membrane and on the flagellum. Transfer of fluorescent protein between the flagellum and the plasma membrane is compromised, however, suggesting the presence of a diffusion barrier at the base of the Leishmania flagellum. Full-length HASPB is released from the metacyclic parasite surface on to macrophages during phagocytosis but while expression is maintained in intracellular amastigotes, HASPB cannot be detected on the external surface in these cells. Thus HASPB may be a dual function protein that is shed by the infective metacyclic but retained internally once Leishmania are taken up by macrophages.
Insights
Leishmania hydrophilic acylated surface protein B (HASPB) is essential for parasite differentiation. This study reveals HASPB
Area of Science:
- Parasitology
- Cell Biology
- Molecular Biology
Background:
- Proteins of the Leishmania hydrophilic acylated surface protein B (HASPB) family are crucial for infective parasite stages.
- HASPB and SHERP are essential for parasite differentiation (metacyclogenesis) in the sand fly vector.
- HASPB is a non-classically secreted protein requiring N-terminal acylation for plasma membrane localization.
Purpose of the Study:
- To investigate the trafficking pathway of HASPB to the plasma membrane and flagellum in metacyclic parasites.
- To explore the dynamics and localization of HASPB at the cell surface and flagellum.
- To understand the fate of HASPB during host cell invasion.
Main Methods:
- Live cell imaging of HASPB-green fluorescent protein (GFP) fusion proteins.
- Analysis of protein trafficking and membrane dynamics.
- Investigation of protein release during host cell interaction.
Main Results:
- No evidence for a phosphorylation-regulated recycling pathway of HASPB in metacyclic parasites.
- HASPB exhibits bidirectional movement within the plasma membrane and on the flagellum.
- A diffusion barrier at the flagellar base restricts HASPB transfer.
- HASPB is shed from the parasite surface onto macrophages during phagocytosis.
- HASPB is not detected on the external surface of intracellular amastigotes.
Conclusions:
- HASPB trafficking to the membrane and flagellum is independent of phosphorylation-regulated recycling in metacyclic parasites.
- A diffusion barrier exists at the base of the Leishmania flagellum.
- HASPB may function as a dual-role protein, shed by infective stages but retained internally after macrophage uptake.
