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Infection and Immunity, January 2001, p. 353-359, Vol. 69, No. 1
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.1.353-359.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Targeted Reduction in Expression of Trypanosoma cruzi Surface Glycoprotein gp90 Increases Parasite Infectivity

Sergio Málaga and Nobuko Yoshida*

Departamento de Microbiologia, Imunologia e Parasitologia, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, Brazil

Received 15 June 2000/Returned for modification 4 August 2000/Accepted 11 October 2000

A previous study had shown that the expression of gp90, a stage-specific surface glycoprotein of Trypanosoma cruzi metacyclic trypomastigotes, is inversely correlated with the parasite's ability to invade mammalian cells. By using antisense oligonucleotides complementary to a region of the gp90 gene implicated in host cell adhesion, we investigated whether the selective inhibition of gp90 synthesis affected the capacity of metacyclic forms to enter target cells. Parasites were incubated for 24 h with 20 µM PS1, a phosphorothioate oligonucleotide based on a sequence of the gp90 coding strand; PS2, the antisense counterpart of PS1; or PO2, the unmodified version of PS2 containing phosphodiester linkages, and the expression of surface molecules was analyzed by flow cytometry and immunoblotting using specific monoclonal antibodies. PS2 but not PS1 or PO2 inhibited the expression of gp90. Inhibition by PS2 was dose dependent. Northern blot analysis revealed that steady-state gp90 mRNA levels were diminished in PS2-treated parasites compared to untreated controls. Treatment with PS2 did not affect the expression of other metacyclic stage surface glycoproteins involved in parasite-host cell interaction, such as gp82 and the mucin-like gp35/50. Expression of gp90 was also inhibited by other phosphorothioate oligonucleotides targeted to the gp90 gene (PS4, PS5, PS6, and PS7) but not by PS3, with the same base composition as PS2 but a mismatched sequence. Parasites treated with PS2, PS4, or PS5 entered HeLa cells in significantly higher numbers than untreated controls, whereas the invasive capacity of PS1- and PS3-treated parasites was unchanged, confirming the inverse association between infectivity and gp90 expression.

* Corresponding author. Mailing address: Escola Paulista de Medicina, Universidade Federal de São Paulo, R. Botucatu, 862- 6° andar, 04023-062, São Paulo, S.P., Brazil. Phone: 55-11-576-4532. Fax: 55-11-571-1095. E-mail: nyoshida{at}ecb.epm.br.

Infection and Immunity, January 2001, p. 353-359, Vol. 69, No. 1
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.1.353-359.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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