A phospholipase C from Trypanosoma brucei which selectively cleaves the glycolipid on the variant surface glycoprotein.

Hereld, Dale; Krakow, Jessica L.; Bangs, James D.; Hart, Gerald W.; Englund, Paul T.

doi:10.1016/s0021-9258(18)67092-9

Cited by 191 publications

(24 citation statements)

References 47 publications

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“…7). The molecular weight of the enzyme, as assessed by gel filtration was ~43,000 which corresponds closely with the estimated value of 37,000-39,800 on SDS-PAGE by other laboratories (8,20,27). Sometimes the lipase tended to form higher molecular weight aggregates if the initial solubilization was done in 1% instead of 2 % CHAPS.…”

Section: Studies Using Partially Purified Lipasesupporting

confidence: 84%

“…It appears clear that the remarkable ability of the trypanosome to alter its surface glycoprotein (14) may preclude the development of a conventional vaccine against trypanosomiasis. Since the in-This work was presented in part at the 6th International Congress of Parasitology (Brisbane, Australia) August [25][26][27][28][29]1986. (Grab, D. J., S. Ito, J. D. Lonsdale-Eccles, P. Webster, and Y. Verjee, 1986, ICOPA FI Handbook, 140.…”

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confidence: 99%

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Subcellular localization of a variable surface glycoprotein phosphatidylinositol-specific phospholipase-C in African trypanosomes.

Grab¹,

Webster²,

Ito³

et al. 1987

The Journal of Cell Biology

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Abstract. African trypanosomes contain a membranebound enzyme capable of removing dimyristylglycerol from the membrane-attached form of the variable surface glycoprotein (mfVSG; Ferguson, M. A. J., K. Halder, and G. A. M. Cross, 1985, J. Biol Chem., 260:4963-4968). Although mfVSG phospholipase-C has been implicated in the removal of the VSG from the trypanosome surface (Cardoso de Almeida, M. L., and M. J. Turner, 1983, Nature (Lond.)., 302:349-352; Ferguson, M. A. J., K. Halder, and G. A. M. Cross, 1985, J. Biol Chem., 260:4963-4968), its precise function and subcellular location have not been determined. We have developed a procedure for the separation of the cell fractions and organelles of Trypanosoma brucei brucei (and other trypanosome species) by differential sucrose and isopycnic Percoll R centrifugation. These fractions were tested for mfVSG phospholipase activity using Trypanosoma brucei mfVSG labeled with 3H-myristic acid as substrate.The highest enzyme-specific activity was associated with the flagella and evidence is presented to suggest that it is localized in the flagellar pocket. Some activity was also associated with the Golgi complex. These results suggest that the mfVSG phospholipase is localized primarily in the membrane of the flagella pocket and possibly other membrane organelles derived from and associated with this structure, and may be part of the VSG-membrane recycling system in African trypanosomes.The activity of mfVSG phospholipase amongst various trypanosome species was determined. We show that, in contrast to the bloodstream forms of 1. Abbreviations used in this paper: CRD, cross-reacting determinant; GPDH, glycerol-3-phosphate dehydrogenase; LG fraction, large granule fraction; mfVSG, membrane-attached form of the variable surface glycoprotein; SG fraction, small granule fraction; sVSG, soluble form of the variable surface glycoprotein; VSG, variable surface glycoprotein. tegrity of the surface coat is known to be essential for parasite survival in the host, the purpose of this study was to investigate VSG synthesis, processing, and replacement on the cell surface. A clearer understanding of the cell biology of the VSG may suggest alternative approaches for the possible control of trypanosomiasis.The VSGs from T. brucei are proteins that contain complex carbohydrate moeities of mannose, galactose, glucosamine, and myristilated phosphotidylinositol (17,18,39) conjugated to the protein through the alpha-carboxyl group of the carboxy-terminal amino acid via an ethanolamine residue (13,28,39). The biological importance to VSG of this carbohydrate side chain is evident since it is present on all variable antigens of T. brucei and T. congolense organisms so far studied and is probably responsible for cross-reactivity observed between soluble forms of VSGs (also known as the cross-reacting determinant [CRD]) (14). The carbohydrate side chain may be involved in anchoring the VSG to the plasma membrane (15,17,18,39), and maintaining the integrity of VSG-VSG interactions within the coat ...

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Section: Studies Using Partially Purified Lipasesupporting

confidence: 84%

mentioning

confidence: 99%

Subcellular localization of a variable surface glycoprotein phosphatidylinositol-specific phospholipase-C in African trypanosomes.

Grab¹,

Webster²,

Ito³

et al. 1987

The Journal of Cell Biology

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“…After centrifugation (TLS-55 rotor, 200,000 × g , 2 h, 4°C), gradients were manually fractionated from the top (5 × 280-μl fractions). Samples were fractionated by SDS-PAGE and immunoblotted with anti-VSG221 VSG or anti-CRD antibodies ( 12 ).…”

Section: Methodsmentioning

confidence: 99%

“…BSF cells have a GPI-specific phospholipase C (GPI-PLC) that is particularly enigmatic. Approximately 30,000 GPI-PLC molecules are associated with the cytoplasmic face of intracellular membranes ( 11 – 14 ), though the protein contains no clear N-terminal signal peptide or transmembrane domain. GPI-PLC is tightly regulated, possibly by dynamic acylation ( 15 , 16 ), but when cells are disrupted by mechanical or hypotonic lysis, GPI-PLC gains full access to the cell surface and rapidly cleaves all the VSG GPI anchors ( 17 , 18 ).…”

Section: Introductionmentioning

confidence: 99%

Turnover of Variant Surface Glycoprotein in Trypanosoma brucei Is a Bimodal Process

Garrison

Umaer

Cipriano

et al. 2021

mBio

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“…Trypanosoma brucei contains an endogenous phospholipase C (PLC) known as the GPI-PLC (Bülow and Overath, 1986;Fox et al, 1986;Hereld et al, 1986) that is capable of hydrolyzing the GPI anchor on the VSG, releasing dimyristyl glycerol (Ferguson et al, 1985). The result of this hydrolysis is to convert the hydrophobic membrane form VSG (mfVSG) to a water soluble VSG (sVSG; Cardoso de Almeida and Turner, 1983), and hypotonic lysis (Cardoso de Almeida and Turner, 1983) or stress (Voorheis et al, 1982;Rolin et al, 1996) results in the shedding of the VSG from the membrane.…”

mentioning

confidence: 99%

The GPI-Phospholipase C of Trypanosoma brucei Is Nonessential But Influences Parasitemia in Mice

Webb

Carnall

Vanhamme

et al. 1997

The Journal of Cell Biology

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In the mammalian host, the cell surface of Trypanosoma brucei is protected by a variant surface glycoprotein that is anchored in the plasma membrane through covalent attachment of the COOH terminus to a glycosylphosphatidylinositol. The trypanosome also contains a phospholipase C (GPI-PLC) that cleaves this anchor and could thus potentially enable the trypanosome to shed the surface coat of VSG. Indeed, release of the surface VSG can be observed within a few minutes on lysis of trypanosomes in vitro. To investigate whether the ability to cleave the membrane anchor of the VSG is an essential function of the enzyme in vivo, a GPI-PLC null mutant trypanosome has been generated by targeted gene deletion. The mutant trypanosomes are fully viable; they can go through an entire life cycle and maintain a persistent infection in mice. Thus the GPI-PLC is not an essential activity and is not necessary for antigenic variation. However, mice infected with the mutant trypanosomes have a reduced parasitemia and survive longer than those infected with control trypanosomes. This phenotype is partially alleviated when the null mutant is modified to express low levels of GPI-PLC.

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A phospholipase C from Trypanosoma brucei which selectively cleaves the glycolipid on the variant surface glycoprotein.

Cited by 191 publications

References 47 publications

Subcellular localization of a variable surface glycoprotein phosphatidylinositol-specific phospholipase-C in African trypanosomes.

Subcellular localization of a variable surface glycoprotein phosphatidylinositol-specific phospholipase-C in African trypanosomes.

Turnover of Variant Surface Glycoprotein in Trypanosoma brucei Is a Bimodal Process

The GPI-Phospholipase C of Trypanosoma brucei Is Nonessential But Influences Parasitemia in Mice

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