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De Novo sequencing and transcriptome analysis for Tetramorium bicarinatum: a comprehensive venom gland transcriptome analysis from an ant species.

Bouzid W, Verdenaud M, Klopp C, Ducancel F, Noirot C, Vétillard A - BMC Genomics (2014)

Bottom Line: Moreover, our results revealed for the first time the presence of toxin-like peptides that have been previously identified from unrelated venomous animals such as waprin-like (snakes) and agatoxins (spiders and conus).The non-toxin transcripts were mainly represented by contigs involved in protein folding and translation, consistent with the protein-secretory function of the venom gland tissue.To the best of our knowledge, this work reports the first large-scale analysis of genes transcribed by the venomous gland of the ant species T. bicarinatum and helps with the identification of Hymenoptera toxin arsenal.In addition, results from this study demonstrate that de novo transcriptome assembly allows useful venom gene expression analysis in a species lacking a genome sequence database.

View Article: PubMed Central - PubMed

Affiliation: Venoms and Biological Activities Laboratory, EA 4357, PRES-University of Toulouse, Jean-François Champollion University Center, Albi, France. angelique.vetillard@univ-jfc.fr.

ABSTRACT

Background: Arthropod venoms are invaluable sources of bioactive substances with biotechnological application. The limited availability of some venoms, such as those from ants, has restricted the knowledge about the composition and the potential that these biomolecules could represent. In order to provide a global insight on the transcripts expressed in the venom gland of the Brazilian ant species Tetramorium bicarinatum and to unveil the potential of its products, high-throughput approach using Illumina technology has been applied to analyze the genes expressed in active venom glands of this ant species.

Results: A total of 212,371,758 pairs of quality-filtered, 100-base-pair Illumina reads were obtained. The de novo assemblies yielded 36,042 contigs for which 27,873 have at least one predicted ORF among which 59.77% produce significant hits in the available databases. The investigation of the reads mapping toxin class revealed a high diversification with the major part consistent with the classical hymenopteran venom protein signature represented by venom allergen (33.3%), followed by a diverse toxin-expression profile including several distinct isoforms of phospholipase A1 and A2, venom serine protease, hyaluronidase, protease inhibitor and secapin. Moreover, our results revealed for the first time the presence of toxin-like peptides that have been previously identified from unrelated venomous animals such as waprin-like (snakes) and agatoxins (spiders and conus).The non-toxin transcripts were mainly represented by contigs involved in protein folding and translation, consistent with the protein-secretory function of the venom gland tissue. Finally, about 40% of the generated contigs have no hits in the databases with 25% of the predicted peptides bearing signal peptide emphasizing the potential of the investigation of these sequences as source of new molecules. Among these contigs, six putative novel peptides that show homologies with previously identified antimicrobial peptides were identified.

Conclusions: To the best of our knowledge, this work reports the first large-scale analysis of genes transcribed by the venomous gland of the ant species T. bicarinatum and helps with the identification of Hymenoptera toxin arsenal. In addition, results from this study demonstrate that de novo transcriptome assembly allows useful venom gene expression analysis in a species lacking a genome sequence database.

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Related in: MedlinePlus

Composition of the venom-gland ‘over-expressed’ protein/peptides predicted from the venom gland transcripts ofT. bicarinatum. Details on the composition of the toxin and non-toxin group are given. Unknown group (9%) designates the contigs that matched hypothetical or uncharacterized proteins.
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Fig3: Composition of the venom-gland ‘over-expressed’ protein/peptides predicted from the venom gland transcripts ofT. bicarinatum. Details on the composition of the toxin and non-toxin group are given. Unknown group (9%) designates the contigs that matched hypothetical or uncharacterized proteins.

Mentions: In order to assess differentially expressed genes in the venom glands, we used the previously described DESeq approach implemented in R package which allows investigation of differential expressed transcripts from high-throughput data without replicates [15]. Regarding a likely important expression level variation between the two tested samples, the analysis failed to give workable results. This bias could be linked to the necessary pre-amplification of the venom gland sample prior to library construction compared to ant whole body sample from which extracted RNA quantity was sufficient to be directly processed to cDNA library construction (see Methods section). To bypass this bias probably originating from the amplification step, we set an arbitrary 103 fold expression-threshold to the 37,818 contigs obtained from both sequenced samples. Due to a lower quality of the sequencing of the read2, we decided to perform our analysis using only the read1 for both samples. In total, we generated a list of 502 contigs that are 1000 fold more expressed in the venom gland, that we called the ‘venom-gland over-expressed’ transcripts. Analysis of the functional diversification among this group revealed a significant abundance (77%) of contigs that have no hits in the databases (Figure 3). On the other hand, contigs that matched predicted toxins and non-toxin proteins, accounted for 11% and 3%, respectively. The remaining group of 9% consisted of uncharacterized compounds (Figure 3). Moreover, about 235 contigs have significant hit with available bacterial sequences. These sequences have not been eliminated given that their low number does not affect the overall assemblage quality. The distribution of the possible bacterial community cohabiting with T. bicarinatum according to their contig number is provided in (Additional file 1: Figure S1).Figure 3


De Novo sequencing and transcriptome analysis for Tetramorium bicarinatum: a comprehensive venom gland transcriptome analysis from an ant species.

Bouzid W, Verdenaud M, Klopp C, Ducancel F, Noirot C, Vétillard A - BMC Genomics (2014)

Composition of the venom-gland ‘over-expressed’ protein/peptides predicted from the venom gland transcripts ofT. bicarinatum. Details on the composition of the toxin and non-toxin group are given. Unknown group (9%) designates the contigs that matched hypothetical or uncharacterized proteins.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4256838&req=5

Fig3: Composition of the venom-gland ‘over-expressed’ protein/peptides predicted from the venom gland transcripts ofT. bicarinatum. Details on the composition of the toxin and non-toxin group are given. Unknown group (9%) designates the contigs that matched hypothetical or uncharacterized proteins.
Mentions: In order to assess differentially expressed genes in the venom glands, we used the previously described DESeq approach implemented in R package which allows investigation of differential expressed transcripts from high-throughput data without replicates [15]. Regarding a likely important expression level variation between the two tested samples, the analysis failed to give workable results. This bias could be linked to the necessary pre-amplification of the venom gland sample prior to library construction compared to ant whole body sample from which extracted RNA quantity was sufficient to be directly processed to cDNA library construction (see Methods section). To bypass this bias probably originating from the amplification step, we set an arbitrary 103 fold expression-threshold to the 37,818 contigs obtained from both sequenced samples. Due to a lower quality of the sequencing of the read2, we decided to perform our analysis using only the read1 for both samples. In total, we generated a list of 502 contigs that are 1000 fold more expressed in the venom gland, that we called the ‘venom-gland over-expressed’ transcripts. Analysis of the functional diversification among this group revealed a significant abundance (77%) of contigs that have no hits in the databases (Figure 3). On the other hand, contigs that matched predicted toxins and non-toxin proteins, accounted for 11% and 3%, respectively. The remaining group of 9% consisted of uncharacterized compounds (Figure 3). Moreover, about 235 contigs have significant hit with available bacterial sequences. These sequences have not been eliminated given that their low number does not affect the overall assemblage quality. The distribution of the possible bacterial community cohabiting with T. bicarinatum according to their contig number is provided in (Additional file 1: Figure S1).Figure 3

Bottom Line: Moreover, our results revealed for the first time the presence of toxin-like peptides that have been previously identified from unrelated venomous animals such as waprin-like (snakes) and agatoxins (spiders and conus).The non-toxin transcripts were mainly represented by contigs involved in protein folding and translation, consistent with the protein-secretory function of the venom gland tissue.To the best of our knowledge, this work reports the first large-scale analysis of genes transcribed by the venomous gland of the ant species T. bicarinatum and helps with the identification of Hymenoptera toxin arsenal.In addition, results from this study demonstrate that de novo transcriptome assembly allows useful venom gene expression analysis in a species lacking a genome sequence database.

View Article: PubMed Central - PubMed

Affiliation: Venoms and Biological Activities Laboratory, EA 4357, PRES-University of Toulouse, Jean-François Champollion University Center, Albi, France. angelique.vetillard@univ-jfc.fr.

ABSTRACT

Background: Arthropod venoms are invaluable sources of bioactive substances with biotechnological application. The limited availability of some venoms, such as those from ants, has restricted the knowledge about the composition and the potential that these biomolecules could represent. In order to provide a global insight on the transcripts expressed in the venom gland of the Brazilian ant species Tetramorium bicarinatum and to unveil the potential of its products, high-throughput approach using Illumina technology has been applied to analyze the genes expressed in active venom glands of this ant species.

Results: A total of 212,371,758 pairs of quality-filtered, 100-base-pair Illumina reads were obtained. The de novo assemblies yielded 36,042 contigs for which 27,873 have at least one predicted ORF among which 59.77% produce significant hits in the available databases. The investigation of the reads mapping toxin class revealed a high diversification with the major part consistent with the classical hymenopteran venom protein signature represented by venom allergen (33.3%), followed by a diverse toxin-expression profile including several distinct isoforms of phospholipase A1 and A2, venom serine protease, hyaluronidase, protease inhibitor and secapin. Moreover, our results revealed for the first time the presence of toxin-like peptides that have been previously identified from unrelated venomous animals such as waprin-like (snakes) and agatoxins (spiders and conus).The non-toxin transcripts were mainly represented by contigs involved in protein folding and translation, consistent with the protein-secretory function of the venom gland tissue. Finally, about 40% of the generated contigs have no hits in the databases with 25% of the predicted peptides bearing signal peptide emphasizing the potential of the investigation of these sequences as source of new molecules. Among these contigs, six putative novel peptides that show homologies with previously identified antimicrobial peptides were identified.

Conclusions: To the best of our knowledge, this work reports the first large-scale analysis of genes transcribed by the venomous gland of the ant species T. bicarinatum and helps with the identification of Hymenoptera toxin arsenal. In addition, results from this study demonstrate that de novo transcriptome assembly allows useful venom gene expression analysis in a species lacking a genome sequence database.

Show MeSH
Related in: MedlinePlus