Gene Survey of the Pathogenic Protozoan Trypanosoma cruzi

Abstract

We have performed a survey of the active genes in the important human pathogen Trypanosoma cruzi by analyzing 5013 expressed sequence tags (ESTs) generated from a normalized epimastigote cDNA library. Clustering of all sequences resulted in 771 clusters, comprising 54% of the ESTs. In total, the ESTs corresponded to 3054 transcripts that might represent one-fourth of the total gene repertoire in T. cruzi. About 33% of the T. cruzi transcripts showed similarity to sequences in the public databases, and a large number of hitherto undiscovered genes predicted to be involved in transcription, cell cycle control, cell division, signal transduction, secretion, and metabolism were identified. More than 140 full-length gene sequences were derived from the ESTs.Comparisons with all open reading frames in yeast and in Caenorhabditis elegans showed that only 12% of the T. cruzi transcripts were shared among diverse eukaryotic organisms. Comparison with other kinetoplastid sequences identified 237 orthologous genes that are shared between these evolutionarily divergent organisms. The generated data are a useful resource for further studies of the biology of the parasite and for development of new means to combat Chagas' disease.[The sequence data described in this paper have been submitted to the dbEST database under nos.

Analysis of expressed sequence tags from Trypanosoma cruzi amastigotes

“…1 shows the distribution among eight functional categories of all ESTs to which a putative identity could be assigned using Blast analyses. Similar to the EST distribution obtained in previous EST studies (Verdun et al 1998, Porcel et al 2000, with epimastigote cDNA libraries and Agüero et al 2004, with an amastigote cDNA library), a large fraction of mRNAs encoding components of the translation machinery is found in amastigotes. Because the two amastigote libraries were not normalized, we expected a significant number of cDNAs to correspond to abundant transcripts present in this stage of the parasite.…”

“…The full list of the identified ESTs and a detailed functional classification can be found at http:/ /www.icb.ufmg.br/~nage/aest/supplemental_table.html The high frequency of ribosomal protein cDNAs (9% of the total number of cDNAs) is in agreement with the results of the amastigote EST survey reported by Agüero et al (2004), who found an even larger percentage (17.4%) of total ESTs generated from the CL Brener amastigote library encoding ribosomal proteins. This proportion is also similar in the data set from the two epimastigote cDNA libraries (Verdun et al 1998, Porcel et al 2000, thus indicating that both forms of T. cruzi depend on the maintenance of the protein synthesis apparatus characteristic of rapidly dividing cells. In line with that, microarray studies of gene expression during trypomastigote (a non-dividing stage) to amastigote differentiation also identified several genes encoding ribosomal proteins as a group of amastigote up-regulated genes (Minning et al 2003).…”

“…The largest cluster contains 18 ESTs, and 12 clusters were found to contain more than 5 ESTs (see below). This clustering analysis indicates that the redundancy of the library estimated from the percentage of clones assembled into clusters is relatively low, considering the levels of redundancy reported by Porcel et al (2000) (54%) and Verdun et al (1998) (28%) for cDNAs libraries constructed with RNA derived from T. cruzi epimastigotes.…”

“…EST data has also been useful in the final steps of genome annotation and gene prediction of whole genome sequencing projects. Beginning in 1994, the T. cruzi EST sequencing project has generated more than 10,000 ESTs, mainly derived from a normalized cDNA library constructed with RNA from epimastigote cultures (Brandão et al 1997, Verdun et al 1998, Porcel et al 2000 and, more recently, from trypomastigotes and amastigotes of the CL Brener strain (Agüero et al 2004) (http://cruzi.cgb.ki.se/ TcruziEST/index.html). After an intense effort initiated by an International Consortium of three sequencing centers (The Institute for Genomic Research, TIGR, Seattle Biomedical Research Institute, and Karolinska Institute), the complete genome sequence of the 60 Mb haploid genome of the CL Brener clone is expected to be published shortly.…”

A Solanesyl-diphosphate Synthase Localizes in Glycosomes of Trypanosoma cruzi

“…We determined the complete sequence of the T. cruzi cDNA clone TENU4155 (accession number AW324852) (36), which showed similarities to polyprenyl synthases. The sequence surrounding the first ATG complied with the published rules for start codons in protozoa (37).…”