Computational analysis and experimental validation of gene predictions in Toxoplasma gondii.
BACKGROUND
Toxoplasma gondii is an obligate intracellular protozoan that infects 20 to 90% of the population. It can cause both acute and chronic infections, many of which are asymptomatic, and, in immunocompromised hosts, can cause fatal infection due to reactivation from an asymptomatic chronic infection. An essential step towards understanding molecular mechanisms controlling transitions between the various life stages and identifying candidate drug targets is to accurately characterize the T. gondii proteome.


RESULTS
We have explored the proteome of T. gondii tachyzoites with high throughput proteomics experiments and by comparison to publicly available cDNA sequence data. Mass spectrometry analysis validated 2,477 gene coding regions with 6,438 possible alternative gene predictions; approximately one third of the T. gondii proteome. The proteomics survey identified 609 proteins that are unique to Toxoplasma as compared to any known species including other Apicomplexan. Computational analysis identified 787 cases of possible gene duplication events and located at least 6,089 gene coding regions. Commonly used gene prediction algorithms produce very disparate sets of protein sequences, with pairwise overlaps ranging from 1.4% to 12%. Through this experimental and computational exercise we benchmarked gene prediction methods and observed false negative rates of 31 to 43%.


CONCLUSIONS
This study not only provides the largest proteomics exploration of the T. gondii proteome, but illustrates how high throughput proteomics experiments can elucidate correct gene structures in genomes.
Dybas
Joseph M
JM
Biodefense Proteomics Research Center, Albert Einstein College of Medicine, Bronx, New York, United States of America.
Madrid-Aliste
Carlos J
CJ
Che
Fa-Yun
FY
Nieves
Edward
E
Rykunov
Dmitry
D
Angeletti
Ruth Hogue
RH
Weiss
Louis M
LM
Kim
Kami
K
Fiser
Andras
A
D000465:Algorithms; D000595:Amino Acid Sequence; D000818:Animals; D016000:Cluster Analysis; D019295:Computational Biology; D030541:Databases, Genetic; D020224:Expressed Sequence Tags; D017125:Genes, Protozoan / Q000235:genetics*; D016678:Genome / Q000235:genetics; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis / Q000737:chemistry; D020543:Proteome; D040901:Proteomics; D015800:Protozoan Proteins / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D015203:Reproducibility of Results; D017386:Sequence Homology, Amino Acid; D014122:Toxoplasma / Q000235:genetics*
D010455:Peptides; D020543:Proteome; D015800:Protozoan Proteins