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. 2001 May 7;193(9):1097-104.
doi: 10.1084/jem.193.9.1097.

The Exocytosis-regulatory protein synaptotagmin VII mediates cell invasion by Trypanosoma cruzi

E V Caler  1 , S Chakrabarti, K T Fowler, S Rao, N W Andrews
Affiliations

The Exocytosis-regulatory protein synaptotagmin VII mediates cell invasion by Trypanosoma cruzi

E V Caler et al. J Exp Med. .

Abstract

The intracellular protozoan parasite Trypanosoma cruzi causes Chagas' disease, which affects millions of people in Latin America. T. cruzi enters a large number of cell types by an unusual mechanism that involves Ca(2+)-triggered fusion of lysosomes with the plasma membrane. Here we show that synaptotagmin VII (Syt VII), a ubiquitously expressed synaptotagmin isoform that regulates exocytosis of lysosomes, is localized on the membranes of intracellular vacuoles containing T. cruzi. Antibodies against the C(2)A domain of Syt VII or recombinant peptides including this domain inhibit cell entry by T. cruzi, but not by Toxoplasma gondii or Salmonella typhimurium. The C(2)A domains of other ubiquitously expressed synaptotagmin isoforms have no effect on T. cruzi invasion, and mutation of critical residues on Syt VII C(2)A abolish its inhibitory activity. These findings indicate that T. cruzi exploits the Syt VII-dependent, Ca(2+)-regulated lysosomal exocytic pathway for invading host cells.

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Figures

Figure 1
Figure 1
Syt VII is localized on the membranes of lysosomes and of T. cruzi–containing intracellular vacuoles. (A) Detection of Syt VII on extracts of different cell types by Western blot using antibodies specific for the unique NH2-terminal Syt VII ectodomain. (B) Double immunofluorescence showing colocalization of Syt VII (red) and the lysosomal glycoprotein Lamp-1 (green) in HEK-293, 3T3, and CHO cells. Arrows point to examples of compartments double positive for Syt VII and Lamp-1. (C) Double immunofluorescence showing colocalization of Syt VII (red) and Lamp-1 (green) on the membranes of intracellular vacuoles containing T. cruzi (arrows). DAPI staining of host cell and parasite DNA is shown in blue.
Figure 2
Figure 2
Antibodies against the Syt VII C2A domain inhibit cell invasion by T. cruzi. (A) DAPI DNA stain (left) and Texas Red–dextran (right) fluorescent images of CHO cells loaded with preimmune rabbit IgG. Arrows point to infected cells previously loaded with dextran and IgG. (B) Same as A, except that the cells were loaded with affinity-purified antibodies against the Syt VII C2A domain. Arrowheads point to uninfected cells previously loaded with dextran and IgG. (C) Quantitation of the number of intracellular trypomastigotes found in cells loaded (black bars) or unloaded (white bars) with dextran, in the presence of PBS only or of preimmune (PI) or anti–Syt VII C2A antibodies (α-C2A). The data is expressed as the mean ± SD of triplicate infections.
Figure 3
Figure 3
Expression of the Syt VII C2A domain inhibits cell invasion by T. cruzi but not by T. gondii or S. typhimurium. (A) T. cruzi invasion of CHO cells untransfected (gray bars; unt) or transfected (white bars) with GFPv, Syt I C2A–GFP, and Syt VII C2A–GFP. (B) T. gondii invasion of CHO cells transfected with GFPv, Syt I C2A–GFP, and Syt VII C2A–GFP. (C) S. typhymurium invasion of CHO cells transfected with GFPv, Syt I C2A–GFP, and Syt VII C2A–GFP. The data is expressed as the mean ± SD of triplicate infections. The color images below panels A, B, and C illustrate the immunofluorescence detection assays used to distinguish intracellular and extracellular pathogens: GFP is shown in green, antibodies against T. cruzi, Toxoplasma, or Salmonella in red, and DAPI DNA stain in blue. Arrows point to extracellular organisms and arrowheads to intracellular ones.
Figure 4
Figure 4
The inhibitory effect on T. cruzi invasion is specific for Syt VII and is abolished by mutation of critical C2A domain residues. The number of intracellular trypomastigotes was quantitated after infection of: (A) NRK cells transfected with GFPv (pEGFP-N2 vector alone), Syt I C2A–GFP, or Syt VII C2A–GFP; (B) NRK cells microinjected with his-tagged Syt I C2A or Syt VII C2A; (C) CHO cells transfected with GFPv or GFP-fused C2A domains of the synaptotagmin isoforms Syt I, VII, IX, or VIII; (D) CHO cells transfected with GFPv, Syt VII C2A–GFP, or mSyt VII C2A. The data is expressed as the mean ± SD of triplicate infections.
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References

    1. Schenkman S., Robbins E.S., Nussenzweig V. Attachment of Trypanosoma cruzi to mammalian cells requires parasite energy, and invasion can be independent of the target cell cytoskeleton. Infect. Immun. 1991;59:645–654. - PMC - PubMed
    1. Tardieux I., Webster P., Ravesloot J., Boron W., Lunn J.A., Heuser J.E., Andrews N.W. Lysosome recruitment and fusion are early events required for trypanosome invasion of mammalian cells. Cell. 1992;71:1117–1130. - PubMed
    1. Rodríguez A., Samoff E., Rioult M.G., Chung A., Andrews N.W. Host cell invasion by trypanosomes requires lysosomes and microtubule/kinesin-mediated transport. J. Cell Biol. 1996;134:349–362. - PMC - PubMed
    1. Rodríguez A., Rioult M.G., Ora A., Andrews N.W. A trypanosome soluble factor induces IP3 formation, intracellular Ca2+ mobilization, and microfilament rearrangement in host cells. J. Cell Biol. 1995;129:1263–1273. - PMC - PubMed
    1. Dorta M.L., Ferreira A.T., Oshiro M.E.M., Yoshida N. Ca2+ signal induced by Trypanosoma cruzi metacyclic trypomastigote surface molecules implicated in mammalian cell invasion. Mol. Biochem. Parasitol. 1995;73:285–289. - PubMed

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