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Transcriptomic analyses reveal novel genes with sexually dimorphic expression in the zebrafish gonad and brain.

Sreenivasan R, Cai M, Bartfai R, Wang X, Christoffels A, Orban L - PLoS ONE (2008)

Bottom Line: We found that five members of the GTP-binding septin gene family, from which only one member (septin 4) has previously been implicated in reproduction in mice, were all strongly expressed in the gonads.Our microarray would be useful for studying gonad development, differentiation and function not only in zebrafish but also in related teleosts via cross-species hybridizations.Since several genes have been shown to play similar roles in gonadogenesis in zebrafish and other vertebrates, our array may even provide information on genetic disorders affecting gonadal phenotypes and fertility in mammals.

View Article: PubMed Central - PubMed

Affiliation: Reproductive Genomics Group, Temasek Life Sciences Laboratory, Singapore, Singapore.

ABSTRACT

Background: Our knowledge on zebrafish reproduction is very limited. We generated a gonad-derived cDNA microarray from zebrafish and used it to analyze large-scale gene expression profiles in adult gonads and other organs.

Methodology/principal findings: We have identified 116638 gonad-derived zebrafish expressed sequence tags (ESTs), 21% of which were isolated in our lab. Following in silico normalization, we constructed a gonad-derived microarray comprising 6370 unique, full-length cDNAs from differentiating and adult gonads. Labeled targets from adult gonad, brain, kidney and 'rest-of-body' from both sexes were hybridized onto the microarray. Our analyses revealed 1366, 881 and 656 differentially expressed transcripts (34.7% novel) that showed highest expression in ovary, testis and both gonads respectively. Hierarchical clustering showed correlation of the two gonadal transcriptomes and their similarities to those of the brains. In addition, we have identified 276 genes showing sexually dimorphic expression both between the brains and between the gonads. By in situ hybridization, we showed that the gonadal transcripts with the strongest array signal intensities were germline-expressed. We found that five members of the GTP-binding septin gene family, from which only one member (septin 4) has previously been implicated in reproduction in mice, were all strongly expressed in the gonads.

Conclusions/significance: We have generated a gonad-derived zebrafish cDNA microarray and demonstrated its usefulness in identifying genes with sexually dimorphic co-expression in both the gonads and the brains. We have also provided the first evidence of large-scale differential gene expression between female and male brains of a teleost. Our microarray would be useful for studying gonad development, differentiation and function not only in zebrafish but also in related teleosts via cross-species hybridizations. Since several genes have been shown to play similar roles in gonadogenesis in zebrafish and other vertebrates, our array may even provide information on genetic disorders affecting gonadal phenotypes and fertility in mammals.

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Venn diagram depicting the clones used for in silico normalization for the generation of the Gonad Uniclone Microarray.The number of adult and differentiating gonad-derived zebrafish sequences present in our Gonad Uniclone EST collection and their extent of overlap with gonad-derived public zebrafish ESTs listed in GenBank are shown. For details on sequence pre-processing and clustering, refer to the Methods section. In overlapping sections, the total number of clusters, prefaced by ‘C’, is followed in parentheses by the total number of ESTs contained by the clusters. The total number of singletons is prefaced by ‘S’.
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pone-0001791-g001: Venn diagram depicting the clones used for in silico normalization for the generation of the Gonad Uniclone Microarray.The number of adult and differentiating gonad-derived zebrafish sequences present in our Gonad Uniclone EST collection and their extent of overlap with gonad-derived public zebrafish ESTs listed in GenBank are shown. For details on sequence pre-processing and clustering, refer to the Methods section. In overlapping sections, the total number of clusters, prefaced by ‘C’, is followed in parentheses by the total number of ESTs contained by the clusters. The total number of singletons is prefaced by ‘S’.

Mentions: In order to minimize redundancy, we performed in silico normalization of the above clone set by clustering their sequences together with our clone set published earlier [34], gonad-derived zebrafish ESTs from GenBank and gonadal zebrafish sequences from RefSeq (see Methods for details). Clustering of these sequences resulted in 11320 clusters and 21520 singletons (Fig. 1). Out of these, 542 clusters (representing 1893 ESTs) and 6325 singletons that were isolated by us have not been previously described from the zebrafish gonad by others. When we BLAST-searched our transcripts against GenBank at the nucleotide level (July 2007), 2018 transcripts did not hit any coding regions of the zebrafish genome. Out of these, 90 of them could be translated into peptides, 24 of which contained domains categorized by Gene Ontologies [35] (Table S2). Of the 2018 transcripts, 365 did not find any significant sequence identity to zebrafish ESTs, indicating that they are likely to be absent from commercial zebrafish microarrays. Of the 365 transcripts, 185 did not have hits to any characterized genes in GenBank. Therefore, these sequences represent entirely new zebrafish or vertebrate ESTs, justifying our extension of gene discovery efforts to developmental stages that have not been sampled previously.


Transcriptomic analyses reveal novel genes with sexually dimorphic expression in the zebrafish gonad and brain.

Sreenivasan R, Cai M, Bartfai R, Wang X, Christoffels A, Orban L - PLoS ONE (2008)

Venn diagram depicting the clones used for in silico normalization for the generation of the Gonad Uniclone Microarray.The number of adult and differentiating gonad-derived zebrafish sequences present in our Gonad Uniclone EST collection and their extent of overlap with gonad-derived public zebrafish ESTs listed in GenBank are shown. For details on sequence pre-processing and clustering, refer to the Methods section. In overlapping sections, the total number of clusters, prefaced by ‘C’, is followed in parentheses by the total number of ESTs contained by the clusters. The total number of singletons is prefaced by ‘S’.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2262149&req=5

pone-0001791-g001: Venn diagram depicting the clones used for in silico normalization for the generation of the Gonad Uniclone Microarray.The number of adult and differentiating gonad-derived zebrafish sequences present in our Gonad Uniclone EST collection and their extent of overlap with gonad-derived public zebrafish ESTs listed in GenBank are shown. For details on sequence pre-processing and clustering, refer to the Methods section. In overlapping sections, the total number of clusters, prefaced by ‘C’, is followed in parentheses by the total number of ESTs contained by the clusters. The total number of singletons is prefaced by ‘S’.
Mentions: In order to minimize redundancy, we performed in silico normalization of the above clone set by clustering their sequences together with our clone set published earlier [34], gonad-derived zebrafish ESTs from GenBank and gonadal zebrafish sequences from RefSeq (see Methods for details). Clustering of these sequences resulted in 11320 clusters and 21520 singletons (Fig. 1). Out of these, 542 clusters (representing 1893 ESTs) and 6325 singletons that were isolated by us have not been previously described from the zebrafish gonad by others. When we BLAST-searched our transcripts against GenBank at the nucleotide level (July 2007), 2018 transcripts did not hit any coding regions of the zebrafish genome. Out of these, 90 of them could be translated into peptides, 24 of which contained domains categorized by Gene Ontologies [35] (Table S2). Of the 2018 transcripts, 365 did not find any significant sequence identity to zebrafish ESTs, indicating that they are likely to be absent from commercial zebrafish microarrays. Of the 365 transcripts, 185 did not have hits to any characterized genes in GenBank. Therefore, these sequences represent entirely new zebrafish or vertebrate ESTs, justifying our extension of gene discovery efforts to developmental stages that have not been sampled previously.

Bottom Line: We found that five members of the GTP-binding septin gene family, from which only one member (septin 4) has previously been implicated in reproduction in mice, were all strongly expressed in the gonads.Our microarray would be useful for studying gonad development, differentiation and function not only in zebrafish but also in related teleosts via cross-species hybridizations.Since several genes have been shown to play similar roles in gonadogenesis in zebrafish and other vertebrates, our array may even provide information on genetic disorders affecting gonadal phenotypes and fertility in mammals.

View Article: PubMed Central - PubMed

Affiliation: Reproductive Genomics Group, Temasek Life Sciences Laboratory, Singapore, Singapore.

ABSTRACT

Background: Our knowledge on zebrafish reproduction is very limited. We generated a gonad-derived cDNA microarray from zebrafish and used it to analyze large-scale gene expression profiles in adult gonads and other organs.

Methodology/principal findings: We have identified 116638 gonad-derived zebrafish expressed sequence tags (ESTs), 21% of which were isolated in our lab. Following in silico normalization, we constructed a gonad-derived microarray comprising 6370 unique, full-length cDNAs from differentiating and adult gonads. Labeled targets from adult gonad, brain, kidney and 'rest-of-body' from both sexes were hybridized onto the microarray. Our analyses revealed 1366, 881 and 656 differentially expressed transcripts (34.7% novel) that showed highest expression in ovary, testis and both gonads respectively. Hierarchical clustering showed correlation of the two gonadal transcriptomes and their similarities to those of the brains. In addition, we have identified 276 genes showing sexually dimorphic expression both between the brains and between the gonads. By in situ hybridization, we showed that the gonadal transcripts with the strongest array signal intensities were germline-expressed. We found that five members of the GTP-binding septin gene family, from which only one member (septin 4) has previously been implicated in reproduction in mice, were all strongly expressed in the gonads.

Conclusions/significance: We have generated a gonad-derived zebrafish cDNA microarray and demonstrated its usefulness in identifying genes with sexually dimorphic co-expression in both the gonads and the brains. We have also provided the first evidence of large-scale differential gene expression between female and male brains of a teleost. Our microarray would be useful for studying gonad development, differentiation and function not only in zebrafish but also in related teleosts via cross-species hybridizations. Since several genes have been shown to play similar roles in gonadogenesis in zebrafish and other vertebrates, our array may even provide information on genetic disorders affecting gonadal phenotypes and fertility in mammals.

Show MeSH
Related in: MedlinePlus