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The GPI-phospholipase C of Trypanosoma brucei is nonessential but influences parasitemia in mice.

Webb H, Carnall N, Vanhamme L, Rolin S, Van Den Abbeele J, Welburn S, Pays E, Carrington M - J. Cell Biol. (1997)

Bottom Line: 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 mutant trypanosome has been generated by targeted gene deletion.Thus the GPI-PLC is not an essential activity and is not necessary for antigenic variation.This phenotype is partially alleviated when the mutant is modified to express low levels of GPI-PLC.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Cambridge University, United Kingdom.

ABSTRACT
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 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 mutant is modified to express low levels of GPI-PLC.

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Time course of trypanosome growth on differentiation  of control and PLC− trypanosomes from bloodstream to procyclic forms in vitro.
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Figure 4: Time course of trypanosome growth on differentiation of control and PLC− trypanosomes from bloodstream to procyclic forms in vitro.

Mentions: On ingestion by tsetse flies, bloodstream trypanosomes differentiate to procyclic forms, a process that can be mimicked in vitro (Ziegelbauer et al., 1990). Since the PLC− mutant was first generated as a procyclic cell line, this differentiation would complete the life cycle. Bloodstream-form cells were harvested from irradiated mice 7 d after infection when the population was predominantly stumpy in morphology and hence predisposed to synchronous differentiation (Ziegelbauer et al., 1990). A plot of cell density against time after cells were induced to differentiate is given in Fig. 4. The control trypanosomes displayed a 15-h lag before exponential growth of procyclic forms commenced. This is similar to results previously reported with stumpy-form AnTat 1.1 cells (Ziegelbauer et al., 1990; Rolin et al., 1993). The lag and subsequent growth rate of the PLC− trypanosomes were indistinguishable from the control and are within the normal range expected for this cell line. The distinct morphological changes that occur in wild-type cells during this time (Rolin et al., 1993) were also apparent during differentiation of the PLC− cells (data not shown).


The GPI-phospholipase C of Trypanosoma brucei is nonessential but influences parasitemia in mice.

Webb H, Carnall N, Vanhamme L, Rolin S, Van Den Abbeele J, Welburn S, Pays E, Carrington M - J. Cell Biol. (1997)

Time course of trypanosome growth on differentiation  of control and PLC− trypanosomes from bloodstream to procyclic forms in vitro.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139819&req=5

Figure 4: Time course of trypanosome growth on differentiation of control and PLC− trypanosomes from bloodstream to procyclic forms in vitro.
Mentions: On ingestion by tsetse flies, bloodstream trypanosomes differentiate to procyclic forms, a process that can be mimicked in vitro (Ziegelbauer et al., 1990). Since the PLC− mutant was first generated as a procyclic cell line, this differentiation would complete the life cycle. Bloodstream-form cells were harvested from irradiated mice 7 d after infection when the population was predominantly stumpy in morphology and hence predisposed to synchronous differentiation (Ziegelbauer et al., 1990). A plot of cell density against time after cells were induced to differentiate is given in Fig. 4. The control trypanosomes displayed a 15-h lag before exponential growth of procyclic forms commenced. This is similar to results previously reported with stumpy-form AnTat 1.1 cells (Ziegelbauer et al., 1990; Rolin et al., 1993). The lag and subsequent growth rate of the PLC− trypanosomes were indistinguishable from the control and are within the normal range expected for this cell line. The distinct morphological changes that occur in wild-type cells during this time (Rolin et al., 1993) were also apparent during differentiation of the PLC− cells (data not shown).

Bottom Line: 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 mutant trypanosome has been generated by targeted gene deletion.Thus the GPI-PLC is not an essential activity and is not necessary for antigenic variation.This phenotype is partially alleviated when the mutant is modified to express low levels of GPI-PLC.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry, Cambridge University, United Kingdom.

ABSTRACT
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 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 mutant is modified to express low levels of GPI-PLC.

Show MeSH
Related in: MedlinePlus