Surface coat remodeling during differentiation of Trypanosoma brucei.
African trypanosomes (Trypanosoma brucei) are digenetic parasites whose lifecycle alternates between the mammalian bloodstream and the midgut of the tsetse fly vector. In mammals, proliferating long slender parasites transform into non-diving short stumpy forms, which differentiate into procyclic forms when ingested by the tsetse fly. A hallmark of differentiation is the replacement of the bloodstream stage surface coat composed of variant surface glycoprotein (VSG) with a new coat composed of procylin. An undefined endoprotease and endogenous glycosylphosphatidylinositol-specific phospholipase C (GPI-PLC) have been implicated in releasing the old VSG coat. However, GPI hydrolysis has been considered unimportant because (i) GPI-PLC null mutants are fully viable and (ii) cytosolic GPI-PLC is localized away from cell surface VSG. Utilizing an in vitro differentiation assay with pleomorphic strains we have investigated these modes of VSG release. Shedding is initially by GPI hydrolysis, which ultimately accounts for a substantial portion of total release. Surface biotinylation assays indicate that GPI-PLC does gain access to extracellular VSG, suggesting that this mode is primed in the starting short stumpy population. Proteolytic release is up-regulated during differentiation and is stereoselectively inhibited by peptidomimetic collagenase inhibitors, implicating a zinc metalloprotease. This protease may be related to TbMSP-B, a trypanosomal homologue of Leishmania major surface protease (MSP) described in the accompanying paper (LaCount, D. J., Gruszynski, A. E., Grandgenett, P. M., Bangs, J. D., and Donelson, J. E. (2003) J. Biol. Chem. 278, 24658-24664). Overall, our results demonstrate that surface coat remodeling during differentiation has multiple mechanisms and that GPI-PLC plays a more significant role in VSG release than previously thought.
Gruszynski
Amy E
AE
Department of Biomolecular Chemistry, University of Wisconsin Medical School, Madison 53706, USA.
DeMaster
Andrew
A
Hooper
Nigel M
NM
Bangs
James D
JD
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D054800:Glycosylphosphatidylinositol Diacylglycerol-Lyase; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D014346:Trypanosoma brucei brucei / Q000166:cytology / Q000502:physiology*; D010738:Type C Phospholipases / Q000502:physiology*; D014643:Variant Surface Glycoproteins, Trypanosoma / Q000502:physiology*
D014643:Variant Surface Glycoproteins, Trypanosoma; D010738:Type C Phospholipases; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054800:Glycosylphosphatidylinositol Diacylglycerol-Lyase