Parasites, Hosts and Diseases

The Korea Society for Parasitology and Tropical Medicine (대한기생충학·열대의학회)
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Molecular Characterization of Trypanosoma cruzi Tc8.2 Gene Indicates Two Differential Locations for the Encoded Protein in Epimastigote and Trypomastigote Forms

  • Received : 2015.05.14
  • Accepted : 2015.07.21
  • Published : 2015.08.31
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Abstract

This report describes the molecular characterization of the Tc8.2 gene of Trypanosoma cruzi. Both the Tc8.2 gene and its encoded protein were analyzed by bioinformatics, while Northern blot and RT-PCR were used for the transcripts. Besides, immunolocalization of recombinant protein was done by immunofluorescence and electron microscopy. Analysis indicated the presence of a single copy of Tc8.2 in the T. cruzi genome and 2-different sized transcripts in epimastigotes/amastigotes and trypomastigotes. Immunoblotting showed 70 and 80 kDa polypeptides in epimastigotes and trypomastigotes, respectively, and a differential pattern of immunolocalization. Overall, the results suggest that Tc8.2 is differentially expressed during the T. cruzi life cycle.

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References

  1. World Health Organization. Chagas disease (American trypanosomiasis). WHO Technical Report Series. Geneva, Switzerland. WHO. 2012.
  2. El Sayed NM, Myler PJ, Bartholomeu DC, Nilsson D, Aggarwal G, Tran AN, Ghedin E, Worthey EA, Delcher AL, Blandin G, Westenberger SJ, Caler E, Cerqueira GC, Branche C, Haas B, Anupama A, Arner E, Aslund L, Attipoe P, Bontempi E, Bringaud F, Burton P, Cadag E, Campbell DA, Carrington M, Crabtree J, Darban H, da Silveira JF, de Jong P, Edwards K, Englund PT, Fazelina G, Feldblyum T, Ferella M, Frasch AC, Gull K, Horn D, Hou L, Huang Y, Kindlund E, Klingbeil M, Kluge S, Koo H, Lacerda D, Levin MJ, Lorenzi H, Louie T, Machado CR, McCulloch R, McKenna A, Mizuno Y, Mottram JC, Nelson S, Ochaya S, Osoegawa K, Pai G, Parsons M, Pentony M, Pettersson U, Pop M, Ramirez JL, Rinta J, Robertson L, Salzberg SL, Sanchez DO, Seyler A, Sharma R, Shetty J, Simpson AJ, Sisk E, Tammi MT, Tarleton R, Teixeira S, Van Aken S, Vogt C, Ward PN, Wickstead B, Wortman J, White O, Fraser CM, Stuart KD, Andersson B. The genome sequence of Trypanosoma cruzi, etiologic agent of Chagas disease. Science 2005; 309: 409-415. https://doi.org/10.1126/science.1112631
  3. Medina-Acosta E, Cross GAM. Rapid isolation of DNA from trypanosomatid protozoa using a simple ‘mini-prep’ procedure. Mol Biochem Parasitol 1993; 59: 327-329. https://doi.org/10.1016/0166-6851(93)90231-L
  4. Towbin H, Staehelin T, Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedures and some applications. Proc Natl Acad Sci USA 1979; 76: 4350-4354. https://doi.org/10.1073/pnas.76.9.4350
  5. Porcile PE, Santos MR, Souza RT, Verbisck NV, Brandao A, Urmenyi T, Silva R, Rondinelli E, Lorenzi H, Levin MJ, Degrave W, Franco da Silveira J. A refined molecular karyotype for the reference strain of the Trypanosoma cruzi genome project (clone CL Brener) by assignment of chromosome markers. Gene 2003; 308: 53-65. https://doi.org/10.1016/S0378-1119(03)00489-X
  6. Alsford S, Turner DJ, Obado SO, Sanchez-Flores A, Glover L, Berriman M, Hertz-Fowler C, Horn D. High-throughput phenotyping using parallel sequencing of RNA interference targets in the African trypanosome. Genome Res 2011; 21: 915-924. https://doi.org/10.1101/gr.115089.110
  7. Kabani S, Fenn K, Ross A, Ivens A, Smith TK, Ghazal P, Matthews K. Genome-wide expression profiling of in vivo-derived bloodstream parasite stages and dynamic analysis of mRNA alterations during synchronous differentiation in Trypanosoma brucei. BMC Genomics 2009; 10: 427. https://doi.org/10.1186/1471-2164-10-427
  8. Jensen BC, Sivam D, Kifer CT, Myler PJ, Parsons M. Widespread variation in transcript abundance within and across developmental stages of Trypanosoma brucei. BMC Genomics 2009; 10: 482. https://doi.org/10.1186/1471-2164-10-482
  9. Siegel TN, Hekstra DR, Wang X, Dewell S, Cross GA. Genome-wide analysis of mRNA abundance in two life-cycle stages of Trypanosoma brucei and identification of splicing and polyadenylation sites. Nucleic Acids Res 2010; 38: 4946-4957. https://doi.org/10.1093/nar/gkq237
  10. Kramer S, Queiroz R, Ellis L, Webb H, Hoheisel JD, Clayton C, Carrington M. Heat shock causes a decrease in polysomes and the appearance of stress granules in trypanosomes independently of eIF2(alpha) phosphorylation at Thr169. J Cell Sci 2008; 121: 3002-3014. https://doi.org/10.1242/jcs.031823
  11. Karniely S, Pines O. Single translation-dual destination: mechanism of dual protein targeting in eukaryotes. EMBO J 2005; 6: 420-425. https://doi.org/10.1038/sj.embor.7400394
  12. Benabdellah K, Gonzalez-Rey E, Gonzalez A. Alternative trans-splicing of the Trypanosoma cruzi LYT1 gene transcript results in compartmental and functional switch for the encoded protein. Mol Microbiol 2007; 65: 1559-1567. https://doi.org/10.1111/j.1365-2958.2007.05892.x
  13. Manning-Cela R, Cortes A, Gonzales-Rey E, Van Voorhis WC, Swindle J, Gonzalez A. LYT1 protein is required for efficient in vitro infection by Trypanosoma cruzi. Infect Immun 2001; 69: 3916-3923. https://doi.org/10.1128/IAI.69.6.3916-3923.2001
  14. Manning-Cela R, Gonzalez A, Swindle J. Alternative splicing of LYT1 transcripts in Trypanosoma cruzi. Infect Immun 2002; 70: 4726-4728. https://doi.org/10.1128/IAI.70.8.4726-4728.2002