Glycosylphosphatidylinositol biosynthetic enzymes are localized to a stable tubular subcompartment of the endoplasmic reticulum in Leishmania mexicana.
Glycosylphosphatidylinositols (GPI) are essential components in the plasma membrane of the protozoan parasite Leishmania mexicana, both as membrane anchors for the major surface macromolecules and as the sole class of free glycolipids. We provide evidence that L.mexicana dolichol-phosphate-mannose synthase (DPMS), a key enzyme in GPI biosynthesis, is localized to a distinct tubular subdomain of the endoplasmic reticulum (ER), based on the localization of a green fluorescent protein (GFP)-DPMS chimera and subcellular fractionation experiments. This tubular membrane (termed the DPMS tubule) is also enriched in other enzymes involved in GPI biosynthesis, can be specifically stained with the fluorescent lipid, BODIPY-C5-ceramide, and appears to be connected to specific subpellicular microtubules that underlie the plasma membrane. Perturbation of microtubules and DPMS tubule structure in vivo results in the selective accumulation of GPI anchor precursors, but not free GPIs. The DPMS tubule is closely associated morphologically with the single Golgi apparatus in non-dividing and dividing cells, appears to exclude luminal ER resident proteins and is labeled, together with the Golgi apparatus, with another GFP chimera containing the heterologous human Golgi marker beta1,2-N-acetylglucosaminyltransferase-I. The possibility that the DPMS-tubule is a stable transitional ER is discussed.
Ilgoutz
S C
SC
Departments of Biochemistry and Molecular Biology, University of Melbourne, Royal Parade, Parkville, Victoria 3052, Australia.
Mullin
K A
KA
Southwell
B R
BR
McConville
M J
MJ
D000818:Animals; D015415:Biomarkers / Q000032:analysis; D002455:Cell Division; D002458:Cell Fractionation; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism; D005453:Fluorescence; D017261:Glycosylphosphatidylinositols / Q000096:biosynthesis* / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism; D006801:Humans; D007425:Intracellular Membranes / Q000737:chemistry / Q000201:enzymology* / Q000378:metabolism; D007894:Leishmania mexicana / Q000166:cytology / Q000201:enzymology* / Q000254:growth & development / Q000378:metabolism; D008364:Mannosyltransferases / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D008928:Mitochondria / Q000378:metabolism; D008938:Mitosis; D011498:Protein Precursors / Q000737:chemistry / Q000378:metabolism; D015800:Protozoan Proteins / Q000737:chemistry / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D015415:Biomarkers; D017261:Glycosylphosphatidylinositols; D011498:Protein Precursors; D015800:Protozoan Proteins; D011993:Recombinant Fusion Proteins; D008364:Mannosyltransferases; C022755:dolichyl-phosphate beta-D-mannosyltransferase