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Needles in the EST haystack: large-scale identification and analysis of excretory-secretory (ES) proteins in parasitic nematodes using expressed sequence tags (ESTs).

Nagaraj SH, Gasser RB, Ranganathan S - PLoS Negl Trop Dis (2008)

Bottom Line: Of these 4,710 proteins, 2,490 (52.8%) had orthologues in Caenorhabditis elegans, whereas 621 (13.8%) appeared to be novel, currently having no significant match to any molecule available in public databases.We report the large-scale analysis of ES proteins inferred from EST data for a range of parasitic nematodes.This set of ES proteins provides an inventory of known and novel members of ES proteins as a foundation for studies focused on understanding the biology of parasitic nematodes and their interactions with their hosts, as well as for the development of novel drugs or vaccines for parasite intervention and control.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biomolecular Sciences, Macquarie University, Sydney, New South Wales, Australia.

ABSTRACT

Background: Parasitic nematodes of humans, other animals and plants continue to impose a significant public health and economic burden worldwide, due to the diseases they cause. Promising antiparasitic drug and vaccine candidates have been discovered from excreted or secreted (ES) proteins released from the parasite and exposed to the immune system of the host. Mining the entire expressed sequence tag (EST) data available from parasitic nematodes represents an approach to discover such ES targets.

Methods and findings: In this study, we predicted, using EST2Secretome, a novel, high-throughput, computational workflow system, 4,710 ES proteins from 452,134 ESTs derived from 39 different species of nematodes, parasitic in animals (including humans) or plants. In total, 2,632, 786, and 1,292 ES proteins were predicted for animal-, human-, and plant-parasitic nematodes. Subsequently, we systematically analysed ES proteins using computational methods. Of these 4,710 proteins, 2,490 (52.8%) had orthologues in Caenorhabditis elegans, whereas 621 (13.8%) appeared to be novel, currently having no significant match to any molecule available in public databases. Of the C. elegans homologues, 267 had strong "loss-of-function" phenotypes by RNA interference (RNAi) in this nematode. We could functionally classify 1,948 (41.3%) sequences using the Gene Ontology (GO) terms, establish pathway associations for 573 (12.2%) sequences using Kyoto Encyclopaedia of Genes and Genomes (KEGG), and identify protein interaction partners for 1,774 (37.6%) molecules. We also mapped 758 (16.1%) proteins to protein domains including the nematode-specific protein family "transthyretin-like" and "chromadorea ALT," considered as vaccine candidates against filariasis in humans.

Conclusions: We report the large-scale analysis of ES proteins inferred from EST data for a range of parasitic nematodes. This set of ES proteins provides an inventory of known and novel members of ES proteins as a foundation for studies focused on understanding the biology of parasitic nematodes and their interactions with their hosts, as well as for the development of novel drugs or vaccines for parasite intervention and control.

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Identification and analysis of putative excretory-secretory proteins from parasitic nematode EST datasets.The “input” EST dataset and the results obtained from each step of the workflow are shown. All of these steps, excluding two nematode-specific steps (WormHomolog and RNAi-Phenotype), are currently incorporated within EST2Secretome.
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pntd-0000301-g002: Identification and analysis of putative excretory-secretory proteins from parasitic nematode EST datasets.The “input” EST dataset and the results obtained from each step of the workflow are shown. All of these steps, excluding two nematode-specific steps (WormHomolog and RNAi-Phenotype), are currently incorporated within EST2Secretome.

Mentions: EST2Secretome made possible the large-scale analysis and annotation of all publicly available EST data for nematodes that are parasitic in humans, other animals and plants. In total, 452,134 ESTs from 39 parasitic nematodes were downloaded from dbEST [19]. The organisms were broadly categorised on the basis of the host(s) they infect (Table 1) with seven, 18 and 14 nematodes parasitic in humans, other animals and plants, respectively, being selected for secretome analysis. Putative ES proteins were identified in the first two phases of EST2Secretome (see Figure 2). Phase I pre-processing and assembly resulted in a total of 152,702 representative ESTs (rESTs) comprising 53,377 contigs and 99,326 singletons, with 152,702 rESTs being conceptually translated into 101,514 peptide sequences. In Phase II, these conceptually translated peptide sequences were first analysed for the presence of N-terminal signal peptide, followed by the absence of transmembrane helices. We thus identified a total of 4,710 putative soluble ES proteins (2,632 in animal-, 1,292 in plant- and 786 in human-parasitic nematodes) (see Table 2), representing 4.6% of the total number of putative sequences identified. This result is comparable with earlier single organism studies of the bovine lungworm, D. viviparus [23], in which 85 secreted proteins were identified (representing 5.0% of 1685 peptides) and T. vitirinus [24], in which 40 secreted proteins were identified (representing 6.2% of 640 proteins). We manually examined the ES protein sequence data and found that 14 of 4710 entries were low quality sequences containing predominantly long stretches of unknown amino acids (X's), as a result of repeat masking, followed by conceptual translation. These sequences were from organisms like Meloidogyne chitwoodi and Pratylenchus vulnus which lack repeat libraries. These 14 sequences were functionally analysed and annotated in the EST2Secretome pipeline but could not be assigned any function. This step represents one of the challenges involved in the computational analysis of single pass reads from any organism which is not well characterized based on genomic data.


Needles in the EST haystack: large-scale identification and analysis of excretory-secretory (ES) proteins in parasitic nematodes using expressed sequence tags (ESTs).

Nagaraj SH, Gasser RB, Ranganathan S - PLoS Negl Trop Dis (2008)

Identification and analysis of putative excretory-secretory proteins from parasitic nematode EST datasets.The “input” EST dataset and the results obtained from each step of the workflow are shown. All of these steps, excluding two nematode-specific steps (WormHomolog and RNAi-Phenotype), are currently incorporated within EST2Secretome.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2553489&req=5

pntd-0000301-g002: Identification and analysis of putative excretory-secretory proteins from parasitic nematode EST datasets.The “input” EST dataset and the results obtained from each step of the workflow are shown. All of these steps, excluding two nematode-specific steps (WormHomolog and RNAi-Phenotype), are currently incorporated within EST2Secretome.
Mentions: EST2Secretome made possible the large-scale analysis and annotation of all publicly available EST data for nematodes that are parasitic in humans, other animals and plants. In total, 452,134 ESTs from 39 parasitic nematodes were downloaded from dbEST [19]. The organisms were broadly categorised on the basis of the host(s) they infect (Table 1) with seven, 18 and 14 nematodes parasitic in humans, other animals and plants, respectively, being selected for secretome analysis. Putative ES proteins were identified in the first two phases of EST2Secretome (see Figure 2). Phase I pre-processing and assembly resulted in a total of 152,702 representative ESTs (rESTs) comprising 53,377 contigs and 99,326 singletons, with 152,702 rESTs being conceptually translated into 101,514 peptide sequences. In Phase II, these conceptually translated peptide sequences were first analysed for the presence of N-terminal signal peptide, followed by the absence of transmembrane helices. We thus identified a total of 4,710 putative soluble ES proteins (2,632 in animal-, 1,292 in plant- and 786 in human-parasitic nematodes) (see Table 2), representing 4.6% of the total number of putative sequences identified. This result is comparable with earlier single organism studies of the bovine lungworm, D. viviparus [23], in which 85 secreted proteins were identified (representing 5.0% of 1685 peptides) and T. vitirinus [24], in which 40 secreted proteins were identified (representing 6.2% of 640 proteins). We manually examined the ES protein sequence data and found that 14 of 4710 entries were low quality sequences containing predominantly long stretches of unknown amino acids (X's), as a result of repeat masking, followed by conceptual translation. These sequences were from organisms like Meloidogyne chitwoodi and Pratylenchus vulnus which lack repeat libraries. These 14 sequences were functionally analysed and annotated in the EST2Secretome pipeline but could not be assigned any function. This step represents one of the challenges involved in the computational analysis of single pass reads from any organism which is not well characterized based on genomic data.

Bottom Line: Of these 4,710 proteins, 2,490 (52.8%) had orthologues in Caenorhabditis elegans, whereas 621 (13.8%) appeared to be novel, currently having no significant match to any molecule available in public databases.We report the large-scale analysis of ES proteins inferred from EST data for a range of parasitic nematodes.This set of ES proteins provides an inventory of known and novel members of ES proteins as a foundation for studies focused on understanding the biology of parasitic nematodes and their interactions with their hosts, as well as for the development of novel drugs or vaccines for parasite intervention and control.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biomolecular Sciences, Macquarie University, Sydney, New South Wales, Australia.

ABSTRACT

Background: Parasitic nematodes of humans, other animals and plants continue to impose a significant public health and economic burden worldwide, due to the diseases they cause. Promising antiparasitic drug and vaccine candidates have been discovered from excreted or secreted (ES) proteins released from the parasite and exposed to the immune system of the host. Mining the entire expressed sequence tag (EST) data available from parasitic nematodes represents an approach to discover such ES targets.

Methods and findings: In this study, we predicted, using EST2Secretome, a novel, high-throughput, computational workflow system, 4,710 ES proteins from 452,134 ESTs derived from 39 different species of nematodes, parasitic in animals (including humans) or plants. In total, 2,632, 786, and 1,292 ES proteins were predicted for animal-, human-, and plant-parasitic nematodes. Subsequently, we systematically analysed ES proteins using computational methods. Of these 4,710 proteins, 2,490 (52.8%) had orthologues in Caenorhabditis elegans, whereas 621 (13.8%) appeared to be novel, currently having no significant match to any molecule available in public databases. Of the C. elegans homologues, 267 had strong "loss-of-function" phenotypes by RNA interference (RNAi) in this nematode. We could functionally classify 1,948 (41.3%) sequences using the Gene Ontology (GO) terms, establish pathway associations for 573 (12.2%) sequences using Kyoto Encyclopaedia of Genes and Genomes (KEGG), and identify protein interaction partners for 1,774 (37.6%) molecules. We also mapped 758 (16.1%) proteins to protein domains including the nematode-specific protein family "transthyretin-like" and "chromadorea ALT," considered as vaccine candidates against filariasis in humans.

Conclusions: We report the large-scale analysis of ES proteins inferred from EST data for a range of parasitic nematodes. This set of ES proteins provides an inventory of known and novel members of ES proteins as a foundation for studies focused on understanding the biology of parasitic nematodes and their interactions with their hosts, as well as for the development of novel drugs or vaccines for parasite intervention and control.

Show MeSH
Related in: MedlinePlus