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Infection and Immunity, July 2009, p. 3023-3032, Vol. 77, No. 7
0019-9567/09/$08.00+0 doi:10.1128/IAI.00138-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
Use of L-Proline and ATP Production by Trypanosoma cruzi Metacyclic Forms as Requirements for Host Cell Invasion 
Rafael Miyazawa Martins,1
Charles Covarrubias,1
Robert Galvez Rojas,2
Ariel Mariano Silber,2 and
Nobuko Yoshida1*
Departamento de Microbiologia, Imunologia e Parasitologia, Universidade Federal de São Paulo, R. Botucatu 862, 6° andar, 04023-062 São Paulo, Brazil,1 Departamento de Parasitologia, Universidade de São Paulo, Av. Prof. Lineu Prestes 1374, 05389-970 São Paulo, Brazil2
Received 4 February 2009/ Returned for modification 15 March 2009/ Accepted 25 April 2009
The process of host cell invasion by Trypanosoma cruzi depends on parasite energy. What source of energy is used for that event is not known. To address this and other questions related to T. cruzi energy requirements and cell invasion, we analyzed metacyclic trypomastigote forms of the phylogenetically distant CL and G strains. For both strains, the nutritional stress experienced by cells starved for 24, 36, or 48 h in phosphate-buffered saline reduced the ATP content and the ability of the parasite to invade HeLa cells proportionally to the starvation time. Inhibition of ATP production by treating parasites with rotenone plus antimycin A also diminished the infectivity. Nutrient depletion did not alter the expression of gp82, the surface molecule that mediates CL strain internalization, but increased the expression of gp90, the negative regulator of cell invasion, in the G strain. When L-proline was given to metacyclic forms starved for 36 h, the ATP levels were restored to those of nonstarved controls for both strains. Glucose had no such effect, although this carbohydrate and L-proline were transported in similar fashions. Recovery of infectivity promoted by L-proline treatment of starved parasites was restricted to the CL strain. The profile of restoration of ATP content and gp82-mediated invasion capacity by L-proline treatment of starved Y-strain parasites was similar to that of the CL strain, whereas the Dm28 and Dm30 strains, whose infectivity is downregulated by gp90, behaved like the G strain. L-Proline was also found to increase the ability of the CL strain to traverse a gastric mucin layer, a property important for the establishment of T. cruzi infection by the oral route. Efficient translocation of parasites through gastric mucin toward the target epithelial cells in the stomach mucosa is an essential requirement for subsequent cell invasion. By relying on these closely associated ATP-driven processes, the metacyclic trypomastigotes effectively accomplish their internalization.
* 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: 55-11-5576-4532. Fax: 55-11-5571-1095. E-mail: nyoshida{at}unifesp.br
Published ahead of print on 11 May 2009.
Infection and Immunity, July 2009, p. 3023-3032, Vol. 77, No. 7
0019-9567/09/$08.00+0 doi:10.1128/IAI.00138-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.
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