title: Clathrin-dependent pathways and the cytoskeleton network are involved in ceramide endocytosis by a parasitic protozoan, Giardia lamblia.
abstract: Although identified as an early-diverged protozoan, Giardia lamblia shares many similarities with higher eukaryotic cells, including an internal membrane system and cytoskeleton, as well as secretory pathways. However, unlike many other eukaryotes, Giardia does not synthesize lipids de novo, but rather depends on exogenous sources for both energy production and organelle or membrane biogenesis. It is not known how lipid molecules are taken up by this parasite and if endocytic pathways are involved in this process. In this investigation, we tested the hypothesis that highly regulated and selective lipid transport machinery is present in Giardia and necessary for the efficient internalization and intracellular targeting of ceramide molecules, the major sphingolipid precursor. Using metabolic and pathway inhibitors, we demonstrate that ceramide is internalized through endocytic pathways and is primarily targeted into perinuclear/endoplasmic reticulum membranes. Further investigations suggested that Giardia uses both clathrin-dependent pathways and the actin cytoskeleton for ceramide uptake, as well as microtubule filaments for intracellular localization and targeting. We speculate that this parasitic protozoan has evolved cytoskeletal and clathrin-dependent endocytic mechanisms for importing ceramide molecules from the cell exterior for the synthesis of membranes and vesicles during growth and differentiation.
authors.affiliation: Infectious Diseases/Immunology, University of Texas at El Paso, TX 79968-0519, USA.
authors.forename: Yunuen
authors.initials: Y
authors.lastname: Hernandez
authors.forename: Cynthia
authors.initials: C
authors.lastname: Castillo
authors.forename: Sukla
authors.initials: S
authors.lastname: Roychowdhury
authors.forename: Adrian
authors.initials: A
authors.lastname: Hehl
authors.forename: Stephen B
authors.initials: SB
authors.lastname: Aley
authors.forename: Siddhartha
authors.initials: S
authors.lastname: Das
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D001896:Boron Compounds / Q000032:analysis; D002478:Cells, Cultured; D002518:Ceramides / Q000493:pharmacokinetics*; D002966:Clathrin / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism*; D004705:Endocytosis / Q000502:physiology*; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005455:Fluorescent Antibody Technique / Q000379:methods; D005456:Fluorescent Dyes / Q000032:analysis; D016829:Giardia lamblia / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D009685:Nuclear Envelope / Q000378:metabolism; D015800:Protozoan Proteins / Q000378:metabolism*; D011726:Pyridinium Compounds / Q000032:analysis; D000644:Quaternary Ammonium Compounds / Q000032:analysis; D014404:Tubulin / Q000032:analysis
chemical_list: C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D000199:Actins; D001896:Boron Compounds; D002518:Ceramides; D002966:Clathrin; C092350:FM 4-64; D005456:Fluorescent Dyes; D015800:Protozoan Proteins; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; D014404:Tubulin