jianbo yao - Academia.edu (original) (raw)
Papers by jianbo yao
Human Genetics, 2000
We have studied the fibroblasts of three patients suffering from Leigh syndrome associated with c... more We have studied the fibroblasts of three patients suffering from Leigh syndrome associated with cytochrome c oxidase deficiency (LS-COX-). Their mitochondrial DNA was functional and all nuclear COX subunits had a normal sequence. The expression of transcripts encoding mitochondrial and nuclear COX subunits was normal or slightly increased. Similarly, the OXA1 transcript coding for a protein involved in COX assembly was increased. However, several COX-protein subunits were severely depressed, indicating deficient COX assembly. Surf1, a factor involved in COX biogenesis, was recently reported as mutated in LS-COX- patients, all mutations predicting a truncated protein. Sequence analysis of SURF1 gene in our three patients revealed seven heterozygous mutations, six of which were new : an insertion, a nonsense mutation, a splicing mutation of intron 7 in addition to three missense mutations. The mutation G385 A (Gly124-->Glu) changes a Gly that is strictly conserved in Surfl homologs of 12 species. The substitution G618 C (Asp202-->His), changing an Asp that is conserved only in mammals, appears to be a polymorphism. The mutation T751 C changes Ile246 to Thr, a position at which a hydrophobic amino acid is conserved in all eukaryotic and some bacterial species. Replacing Ile246 by Thr disrupts a predicted beta sheet structure present in all higher eukaryotes. COX activity could be restored in fibroblasts of the three patients by complementation with a retroviral vector containing normal SURF1 cDNA. These mutations identify domains essential to Surf1 protein structure and/or function.
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Biochemical and Biophysical Research Communications, 2000
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Journal of Medical Genetics, 2001
VON KLEIST-RETZOW, JÜRGEN-CHRISTOPH; YAO, JIANBO; TAANMAN, JAN-WILLEM; CHANTREL, KARINE; CHRETIEN... more VON KLEIST-RETZOW, JÜRGEN-CHRISTOPH; YAO, JIANBO; TAANMAN, JAN-WILLEM; CHANTREL, KARINE; CHRETIEN, DOMINIQUE; CORMIER-DAIRE, VALÉRIE; RÖTIG, AGNÈS; MUNNICH, ARNOLD; RUSTIN, PIERRE; SHOUBRIDGE, ERIC A.
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Genome, 1995
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Marine Biotechnology, 2010
MicroRNAs (miRNAs) are small, highly conserved, non-coding RNAs that regulate gene expression of ... more MicroRNAs (miRNAs) are small, highly conserved, non-coding RNAs that regulate gene expression of target mRNAs through cleavage or translational inhibition. miRNAs are most often identified through computational prediction from genome sequences. The rainbow trout genome sequence is not available yet, which does not allow miRNA prediction for this species which is of great economic interest for aquaculture and sport fisheries, and is a model research organism for studies related to carcinogenesis, toxicology, comparative immunology, disease ecology, physiology and nutrition. To identify miRNAs from rainbow trout, we constructed a miRNA library from a pool of nine somatic tissues. Analysis of the library identified 210 unique sequences representing 54 distinct miRNAs; 50 with conserved sequences matching previously identified miRNAs and four novel miRNAs. In addition, 13 miRNAs were computationally predicted from the rainbow trout transcriptome. Real-time PCR was used to measure miRNA expression patterns in adult somatic tissues and unfertilized eggs. The majority of the miRNAs showed characteristic tissue-specific expression patterns suggesting potential roles in maintaining tissue identity. Potential miRNA-target interactions were computationally predicted and single nucleotide polymorphisms (SNPs) were identified in the miRNAs and their target sites in the rainbow trout transcripts. The rainbow trout miRNAs identified and characterized in this study provide a new tool for functional genome research in salmonids. Tissue-specific miRNAs may serve as molecular markers, predictive of specific functional and diagnostic implications. The data on genetic polymorphisms in miRNA-target interactions is particularly useful for rainbow trout breeding programs.
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Comparative Biochemistry and Physiology D-genomics & Proteomics, 2006
Severe muscle deterioration is a physiological response to the energetic demands of fish spawning... more Severe muscle deterioration is a physiological response to the energetic demands of fish spawning. This response represents a suitable model to study mechanisms of muscle degradation in fish where typical tetrapod methods, such as muscle unloading, are not applicable. Enzyme activities and mRNA accumulations of genes in major proteolytic pathways, including cathepsins, calpains and the multi-catalytic proteasome, were measured in white muscles of rainbow trout during spawning and post-spawning seasons of gravid fish for comparisons to sterile fish. Fertile fish at spawning had less muscle tissue and less muscle protein compared to sterile fish and post-spawning fertile fish. Muscle deterioration of the fertile fish during spawning was associated with greater mRNA accumulation and elevated activity of cathepsin-L. Concurrently, muscle of spawning fish showed increased mRNA accumulations of cathepsin-D, the calpain regulatory subunit and the proteasome catalytic subunit alpha without corresponding increases in enzyme activities. In addition, elevated activity and increased mRNA accumulation of caspase-9, but not caspase-3, were observed in fertile fish during spawning. This study indicates that cathepsins mediate protein catabolism during spawning in rainbow trout and the catabolic process may involve activation of the apoptosis mediator, caspase-9, but not the apoptosis executioner, caspase-3.
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Comparative Biochemistry and Physiology A-molecular & Integrative Physiology, 2006
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Comparative Biochemistry and Physiology B-biochemistry & Molecular Biology, 2005
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BMC Genomics, 2011
Background Rainbow trout (Oncorhynchus mykiss) are the most-widely cultivated cold freshwater fis... more Background Rainbow trout (Oncorhynchus mykiss) are the most-widely cultivated cold freshwater fish in the world and an important model species for many research areas. Coupling great interest in this species as a research model with the need for genetic improvement of aquaculture production efficiency traits justifies the continued development of genomics research resources. Many quantitative trait loci (QTL) have been identified for production and life-history traits in rainbow trout. An integrated physical and genetic map is needed to facilitate fine mapping of QTL and the selection of positional candidate genes for incorporation in marker-assisted selection (MAS) programs for improving rainbow trout aquaculture production. Results The first generation integrated map of the rainbow trout genome is composed of 238 BAC contigs anchored to chromosomes of the genetic map. It covers more than 10% of the genome across segments from all 29 chromosomes. Anchoring of 203 contigs to chromosomes of the National Center for Cool and Cold Water Aquaculture (NCCCWA) genetic map was achieved through mapping of 288 genetic markers derived from BAC end sequences (BES), screening of the BAC library with previously mapped markers and matching of SNPs with BES reads. In addition, 35 contigs were anchored to linkage groups of the INRA (French National Institute of Agricultural Research) genetic map through markers that were not informative for linkage analysis in the NCCCWA mapping panel. The ratio of physical to genetic linkage distances varied substantially among chromosomes and BAC contigs with an average of 3,033 Kb/cM. Conclusions The integrated map described here provides a framework for a robust composite genome map for rainbow trout. This resource is needed for genomic analyses in this research model and economically important species and will facilitate comparative genome mapping with other salmonids and with model fish species. This resource will also facilitate efforts to assemble a whole-genome reference sequence for rainbow trout.
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BMC Genomics, 2007
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BMC Developmental Biology, 2008
Background Current literature and our previous results on expression patterns of oocyte-specific ... more Background Current literature and our previous results on expression patterns of oocyte-specific genes and transcription factors suggest a global but highly regulated maternal mRNA degradation at the time of embryonic genome activation (EGA). MicroRNAs (miRNAs) are small, non-coding regulatory RNAs (19–23 nucleotides) that regulate gene expression by guiding target mRNA cleavage or translational inhibition. These regulatory RNAs are potentially involved in the degradation of maternally inherited mRNAs during early embryogenesis. Results To identify miRNAs that might be important for early embryogenesis in rainbow trout, we constructed a miRNA library from a pool of unfertilized eggs and early stage embryos. Sequence analysis of random clones from the library identified 14 miRNAs, 4 of which are novel to rainbow trout. Real-time PCR was used to measure the expression of all cloned miRNAs during embryonic development. Four distinct expression patterns were observed and some miRNAs showed up-regulated expression during EGA. Analysis of tissue distribution of these miRNAs showed that some are present ubiquitously, while others are differentially expressed among different tissues. We also analyzed the expression patterns of Dicer, the enzyme required for the processing of miRNAs and Stat3, a transcription factor involved in activating the transcription of miR-21. Dicer is abundantly expressed during EGA and Stat3 is up-regulated before the onset of EGA. Conclusion This study led to the discovery of 14 rainbow trout miRNAs. Our data support the notion that Dicer processes miRNAs and Stat3 induces expression of miR-21 and possibly other miRNAs during EGA. These miRNAs in turn guide maternal mRNAs for degradation, which is required for normal embryonic development.
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BMC Genomics, 2006
Background Uncoupling protein 2 (UCP2) belongs to the superfamily of mitochondrial anion carriers... more Background Uncoupling protein 2 (UCP2) belongs to the superfamily of mitochondrial anion carriers that dissociate the respiratory chain from ATP synthesis. It has been determined that UCP2 plays a role in several physiological processes such as energy expenditure, body weight control and fatty acid metabolism in several vertebrate species. We report the first characterization of UCP2s in rainbow trout (Oncorhynchus mykiss). Results Two UCP2 genes were identified in the rainbow trout genome, UCP2A and UCP2B. These genes are 93% similar in their predicted amino acid sequences and display the same genomic structure as other vertebrates (8 exons and 7 introns) spanning 4.2 kb and 3.2 kb, respectively. UCP2A and UCP2B were widely expressed in all tissues of the study with a predominant level in macrophage-rich tissues and reproductive organs. In fry muscle we observed an increase in UCP2B expression in response to fasting and a decrease after refeeding in agreement with previous studies in human, mouse, rat, and marsupials. The converse expression pattern was observed for UCP2A mRNA which decreased during fasting, suggesting different metabolic roles for UCP2A and UCP2B in rainbow trout muscle. Phylogenetic analysis including other genes from the UCP core family located rainbow trout UCP2A and UCP2B with their orthologs and suggested an early divergence of vertebrate UCPs from a common ancestor gene. Conclusion We characterized two UCP2 genes in rainbow trout with similar genomic structures, amino acid sequences and distribution profiles. These genes appeared to be differentially regulated in response to fasting and refeeding in fry muscle. The genomic organization and phylogeny analysis support the hypothesis of a common ancestry between the vertebrate UCPs.
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Comparative Biochemistry and Physiology B-biochemistry & Molecular Biology, 2005
Calpastatin (CAST), the specific inhibitor of the calpain proteases, plays a role in muscle growt... more Calpastatin (CAST), the specific inhibitor of the calpain proteases, plays a role in muscle growth and meat quality. In rainbow trout (RBT), we identified cDNAs coding for two CAST isoforms, a long (CAST-L) and a short isoform (CAST-S), apparently derived from two different genes. Zebrafish and pufferfish CAST cDNA and genomic sequences were retrieved from GenBank and their exon/intron structures were characterized. Fish CASTs are novel in that they have fewer repetitive inhibitory domains as compared to their mammalian counterparts (one or two vs. four). The expressions of CAST mRNAs were measured in three RBT strains with different growth rates and fillet firmness that were fed either high energy or control diets. CAST-L and S expressions were significantly lower (p<0.01) in the strain that has the slowest growth rate and yielded the softest fillet. Strain or diet did not affect level of calpain mRNAs. However, the decrease in the CAST/calpain ratio at the mRNA level did not lead to a corresponding change in the calpain catalytic activity. Further investigation should reveal a potential use of the CAST gene as a tool to monitor fish muscle growth and fillet firmness.
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BMC Genomics, 2009
Background To enhance capabilities for genomic analyses in rainbow trout, such as genomic selecti... more Background To enhance capabilities for genomic analyses in rainbow trout, such as genomic selection, a large suite of polymorphic markers that are amenable to high-throughput genotyping protocols must be identified. Expressed Sequence Tags (ESTs) have been used for single nucleotide polymorphism (SNP) discovery in salmonids. In those strategies, the salmonid semi-tetraploid genomes often led to assemblies of paralogous sequences and therefore resulted in a high rate of false positive SNP identification. Sequencing genomic DNA using primers identified from ESTs proved to be an effective but time consuming methodology of SNP identification in rainbow trout, therefore not suitable for high throughput SNP discovery. In this study, we employed a high-throughput strategy that used pyrosequencing technology to generate data from a reduced representation library constructed with genomic DNA pooled from 96 unrelated rainbow trout that represent the National Center for Cool and Cold Water Aquaculture (NCCCWA) broodstock population. Results The reduced representation library consisted of 440 bp fragments resulting from complete digestion with the restriction enzyme HaeIII; sequencing produced 2,000,000 reads providing an average 6 fold coverage of the estimated 150,000 unique genomic restriction fragments (300,000 fragment ends). Three independent data analyses identified 22,022 to 47,128 putative SNPs on 13,140 to 24,627 independent contigs. A set of 384 putative SNPs, randomly selected from the sets produced by the three analyses were genotyped on individual fish to determine the validation rate of putative SNPs among analyses, distinguish apparent SNPs that actually represent paralogous loci in the tetraploid genome, examine Mendelian segregation, and place the validated SNPs on the rainbow trout linkage map. Approximately 48% (183) of the putative SNPs were validated; 167 markers were successfully incorporated into the rainbow trout linkage map. In addition, 2% of the sequences from the validated markers were associated with rainbow trout transcripts. Conclusion The use of reduced representation libraries and pyrosequencing technology proved to be an effective strategy for the discovery of a high number of putative SNPs in rainbow trout; however, modifications to the technique to decrease the false discovery rate resulting from the evolutionary recent genome duplication would be desirable.
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Animal Reproduction Science, 2010
Ovarian folliculogenesis and early embryogenesis are complex processes, which require tightly reg... more Ovarian folliculogenesis and early embryogenesis are complex processes, which require tightly regulated expression and interaction of a multitude of genes. Small endogenous RNA molecules, termed microRNAs (miRNAs), are involved in the regulation of gene expression during folliculogenesis and early embryonic development. To identify miRNAs in bovine oocytes/ovaries, a bovine fetal ovary miRNA library was constructed. Sequence analysis of random clones from the library identified 679 miRNA sequences, which represent 58 distinct bovine miRNAs. Of these distinct miRNAs, 42 are known bovine miRNAs present in the miRBase database and the remaining 16 miRNAs include 15 new bovine miRNAs that are homologous to miRNAs identified in other species, and one novel miRNA, which does not match any miRNAs in the database. The precursor sequences for 14 of the new 15 miRNAs as well as the novel miRNA were identified from the bovine genome database and their hairpin structures were predicted. Expression analysis of the 58 miRNAs in fetal ovaries in comparison to somatic tissue pools identified 8 miRNAs predominantly expressed in fetal ovaries. Further analysis of the eight miRNAs in germinal vesicle (GV) stage oocytes identified two miRNAs (bta-mir424 and bta-mir-10b), that are highly abundant in GV oocytes. Both miRNAs show similar expression patterns during oocyte maturation and preimplantation development of bovine embryos, being abundant in GV and MII stage oocytes, as well as in early stage embryos (until 16-cell stage). The amount of the novel miRNA is relatively small in oocytes and early cleavage embryos but greater in blastocysts, suggesting a role of this miRNA in blastocyst cell differentiation.
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Molecular Reproduction and Development, 2007
Genes specifically expressed in oocytes are important for the development of oocytes and early em... more Genes specifically expressed in oocytes are important for the development of oocytes and early embryos. By analyzing expressed sequence tags (ESTs) from a rainbow trout oocyte cDNA library, we identified a novel EST sequence that does not show homology to any sequences in the GenBank. Analysis of tissue distribution by RT-PCR revealed that this gene was only expressed in unfertilized oocytes. Sequencing of the EST clone identified a cDNA of 3,163 bp. Northern blot analysis showed the novel gene has a single transcript of 3.4 kb. Additional 5′ sequence was obtained by 5′ RACE, extending the novel cDNA to 3,333 bp. Analysis of the full-length cDNA identified an open reading frame (ORF) encoding a protein of 564 amino acids. The novel protein contains a conserved oxysterol binding protein (OSBP) domain at the C terminus that is characteristic of OSBP-related proteins (ORPs) implicated in lipid metabolism. Therefore, we named the novel gene as Oocyte-specific Oxysterol binding protein Related-Protein of Trout (OORP-T). In situ hybridization showed that the OORP-T mRNA appears to be confined to the cytoplasm of vitellogenic oocytes. Transcription of OORP-T appears to start during pre-vitellogenesis and increases steadily, reaching its peak in the late vitellogenic stage. OORP-T transcript is abundantly present in unfertilized eggs but the level drops significantly in day 2 embryos and continues to decline in day 7 embryos after which it remains low. We propose that OORP-T may play an important role in the utilization of yolk-derived lipid products during oocyte development and early stages of embryonic development in rainbow trout. Mol. Reprod. Dev. 74: 502–511, 2007. © 2006 Wiley-Liss, Inc.
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Animal Reproduction Science, 2008
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BMC Genomics, 2010
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Human Genetics, 2000
We have studied the fibroblasts of three patients suffering from Leigh syndrome associated with c... more We have studied the fibroblasts of three patients suffering from Leigh syndrome associated with cytochrome c oxidase deficiency (LS-COX-). Their mitochondrial DNA was functional and all nuclear COX subunits had a normal sequence. The expression of transcripts encoding mitochondrial and nuclear COX subunits was normal or slightly increased. Similarly, the OXA1 transcript coding for a protein involved in COX assembly was increased. However, several COX-protein subunits were severely depressed, indicating deficient COX assembly. Surf1, a factor involved in COX biogenesis, was recently reported as mutated in LS-COX- patients, all mutations predicting a truncated protein. Sequence analysis of SURF1 gene in our three patients revealed seven heterozygous mutations, six of which were new : an insertion, a nonsense mutation, a splicing mutation of intron 7 in addition to three missense mutations. The mutation G385 A (Gly124-->Glu) changes a Gly that is strictly conserved in Surfl homologs of 12 species. The substitution G618 C (Asp202-->His), changing an Asp that is conserved only in mammals, appears to be a polymorphism. The mutation T751 C changes Ile246 to Thr, a position at which a hydrophobic amino acid is conserved in all eukaryotic and some bacterial species. Replacing Ile246 by Thr disrupts a predicted beta sheet structure present in all higher eukaryotes. COX activity could be restored in fibroblasts of the three patients by complementation with a retroviral vector containing normal SURF1 cDNA. These mutations identify domains essential to Surf1 protein structure and/or function.
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Biochemical and Biophysical Research Communications, 2000
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Journal of Medical Genetics, 2001
VON KLEIST-RETZOW, JÜRGEN-CHRISTOPH; YAO, JIANBO; TAANMAN, JAN-WILLEM; CHANTREL, KARINE; CHRETIEN... more VON KLEIST-RETZOW, JÜRGEN-CHRISTOPH; YAO, JIANBO; TAANMAN, JAN-WILLEM; CHANTREL, KARINE; CHRETIEN, DOMINIQUE; CORMIER-DAIRE, VALÉRIE; RÖTIG, AGNÈS; MUNNICH, ARNOLD; RUSTIN, PIERRE; SHOUBRIDGE, ERIC A.
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Genome, 1995
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Marine Biotechnology, 2010
MicroRNAs (miRNAs) are small, highly conserved, non-coding RNAs that regulate gene expression of ... more MicroRNAs (miRNAs) are small, highly conserved, non-coding RNAs that regulate gene expression of target mRNAs through cleavage or translational inhibition. miRNAs are most often identified through computational prediction from genome sequences. The rainbow trout genome sequence is not available yet, which does not allow miRNA prediction for this species which is of great economic interest for aquaculture and sport fisheries, and is a model research organism for studies related to carcinogenesis, toxicology, comparative immunology, disease ecology, physiology and nutrition. To identify miRNAs from rainbow trout, we constructed a miRNA library from a pool of nine somatic tissues. Analysis of the library identified 210 unique sequences representing 54 distinct miRNAs; 50 with conserved sequences matching previously identified miRNAs and four novel miRNAs. In addition, 13 miRNAs were computationally predicted from the rainbow trout transcriptome. Real-time PCR was used to measure miRNA expression patterns in adult somatic tissues and unfertilized eggs. The majority of the miRNAs showed characteristic tissue-specific expression patterns suggesting potential roles in maintaining tissue identity. Potential miRNA-target interactions were computationally predicted and single nucleotide polymorphisms (SNPs) were identified in the miRNAs and their target sites in the rainbow trout transcripts. The rainbow trout miRNAs identified and characterized in this study provide a new tool for functional genome research in salmonids. Tissue-specific miRNAs may serve as molecular markers, predictive of specific functional and diagnostic implications. The data on genetic polymorphisms in miRNA-target interactions is particularly useful for rainbow trout breeding programs.
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Comparative Biochemistry and Physiology D-genomics & Proteomics, 2006
Severe muscle deterioration is a physiological response to the energetic demands of fish spawning... more Severe muscle deterioration is a physiological response to the energetic demands of fish spawning. This response represents a suitable model to study mechanisms of muscle degradation in fish where typical tetrapod methods, such as muscle unloading, are not applicable. Enzyme activities and mRNA accumulations of genes in major proteolytic pathways, including cathepsins, calpains and the multi-catalytic proteasome, were measured in white muscles of rainbow trout during spawning and post-spawning seasons of gravid fish for comparisons to sterile fish. Fertile fish at spawning had less muscle tissue and less muscle protein compared to sterile fish and post-spawning fertile fish. Muscle deterioration of the fertile fish during spawning was associated with greater mRNA accumulation and elevated activity of cathepsin-L. Concurrently, muscle of spawning fish showed increased mRNA accumulations of cathepsin-D, the calpain regulatory subunit and the proteasome catalytic subunit alpha without corresponding increases in enzyme activities. In addition, elevated activity and increased mRNA accumulation of caspase-9, but not caspase-3, were observed in fertile fish during spawning. This study indicates that cathepsins mediate protein catabolism during spawning in rainbow trout and the catabolic process may involve activation of the apoptosis mediator, caspase-9, but not the apoptosis executioner, caspase-3.
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Comparative Biochemistry and Physiology A-molecular & Integrative Physiology, 2006
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Comparative Biochemistry and Physiology B-biochemistry & Molecular Biology, 2005
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BMC Genomics, 2011
Background Rainbow trout (Oncorhynchus mykiss) are the most-widely cultivated cold freshwater fis... more Background Rainbow trout (Oncorhynchus mykiss) are the most-widely cultivated cold freshwater fish in the world and an important model species for many research areas. Coupling great interest in this species as a research model with the need for genetic improvement of aquaculture production efficiency traits justifies the continued development of genomics research resources. Many quantitative trait loci (QTL) have been identified for production and life-history traits in rainbow trout. An integrated physical and genetic map is needed to facilitate fine mapping of QTL and the selection of positional candidate genes for incorporation in marker-assisted selection (MAS) programs for improving rainbow trout aquaculture production. Results The first generation integrated map of the rainbow trout genome is composed of 238 BAC contigs anchored to chromosomes of the genetic map. It covers more than 10% of the genome across segments from all 29 chromosomes. Anchoring of 203 contigs to chromosomes of the National Center for Cool and Cold Water Aquaculture (NCCCWA) genetic map was achieved through mapping of 288 genetic markers derived from BAC end sequences (BES), screening of the BAC library with previously mapped markers and matching of SNPs with BES reads. In addition, 35 contigs were anchored to linkage groups of the INRA (French National Institute of Agricultural Research) genetic map through markers that were not informative for linkage analysis in the NCCCWA mapping panel. The ratio of physical to genetic linkage distances varied substantially among chromosomes and BAC contigs with an average of 3,033 Kb/cM. Conclusions The integrated map described here provides a framework for a robust composite genome map for rainbow trout. This resource is needed for genomic analyses in this research model and economically important species and will facilitate comparative genome mapping with other salmonids and with model fish species. This resource will also facilitate efforts to assemble a whole-genome reference sequence for rainbow trout.
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BMC Genomics, 2007
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BMC Developmental Biology, 2008
Background Current literature and our previous results on expression patterns of oocyte-specific ... more Background Current literature and our previous results on expression patterns of oocyte-specific genes and transcription factors suggest a global but highly regulated maternal mRNA degradation at the time of embryonic genome activation (EGA). MicroRNAs (miRNAs) are small, non-coding regulatory RNAs (19–23 nucleotides) that regulate gene expression by guiding target mRNA cleavage or translational inhibition. These regulatory RNAs are potentially involved in the degradation of maternally inherited mRNAs during early embryogenesis. Results To identify miRNAs that might be important for early embryogenesis in rainbow trout, we constructed a miRNA library from a pool of unfertilized eggs and early stage embryos. Sequence analysis of random clones from the library identified 14 miRNAs, 4 of which are novel to rainbow trout. Real-time PCR was used to measure the expression of all cloned miRNAs during embryonic development. Four distinct expression patterns were observed and some miRNAs showed up-regulated expression during EGA. Analysis of tissue distribution of these miRNAs showed that some are present ubiquitously, while others are differentially expressed among different tissues. We also analyzed the expression patterns of Dicer, the enzyme required for the processing of miRNAs and Stat3, a transcription factor involved in activating the transcription of miR-21. Dicer is abundantly expressed during EGA and Stat3 is up-regulated before the onset of EGA. Conclusion This study led to the discovery of 14 rainbow trout miRNAs. Our data support the notion that Dicer processes miRNAs and Stat3 induces expression of miR-21 and possibly other miRNAs during EGA. These miRNAs in turn guide maternal mRNAs for degradation, which is required for normal embryonic development.
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BMC Genomics, 2006
Background Uncoupling protein 2 (UCP2) belongs to the superfamily of mitochondrial anion carriers... more Background Uncoupling protein 2 (UCP2) belongs to the superfamily of mitochondrial anion carriers that dissociate the respiratory chain from ATP synthesis. It has been determined that UCP2 plays a role in several physiological processes such as energy expenditure, body weight control and fatty acid metabolism in several vertebrate species. We report the first characterization of UCP2s in rainbow trout (Oncorhynchus mykiss). Results Two UCP2 genes were identified in the rainbow trout genome, UCP2A and UCP2B. These genes are 93% similar in their predicted amino acid sequences and display the same genomic structure as other vertebrates (8 exons and 7 introns) spanning 4.2 kb and 3.2 kb, respectively. UCP2A and UCP2B were widely expressed in all tissues of the study with a predominant level in macrophage-rich tissues and reproductive organs. In fry muscle we observed an increase in UCP2B expression in response to fasting and a decrease after refeeding in agreement with previous studies in human, mouse, rat, and marsupials. The converse expression pattern was observed for UCP2A mRNA which decreased during fasting, suggesting different metabolic roles for UCP2A and UCP2B in rainbow trout muscle. Phylogenetic analysis including other genes from the UCP core family located rainbow trout UCP2A and UCP2B with their orthologs and suggested an early divergence of vertebrate UCPs from a common ancestor gene. Conclusion We characterized two UCP2 genes in rainbow trout with similar genomic structures, amino acid sequences and distribution profiles. These genes appeared to be differentially regulated in response to fasting and refeeding in fry muscle. The genomic organization and phylogeny analysis support the hypothesis of a common ancestry between the vertebrate UCPs.
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Comparative Biochemistry and Physiology B-biochemistry & Molecular Biology, 2005
Calpastatin (CAST), the specific inhibitor of the calpain proteases, plays a role in muscle growt... more Calpastatin (CAST), the specific inhibitor of the calpain proteases, plays a role in muscle growth and meat quality. In rainbow trout (RBT), we identified cDNAs coding for two CAST isoforms, a long (CAST-L) and a short isoform (CAST-S), apparently derived from two different genes. Zebrafish and pufferfish CAST cDNA and genomic sequences were retrieved from GenBank and their exon/intron structures were characterized. Fish CASTs are novel in that they have fewer repetitive inhibitory domains as compared to their mammalian counterparts (one or two vs. four). The expressions of CAST mRNAs were measured in three RBT strains with different growth rates and fillet firmness that were fed either high energy or control diets. CAST-L and S expressions were significantly lower (p<0.01) in the strain that has the slowest growth rate and yielded the softest fillet. Strain or diet did not affect level of calpain mRNAs. However, the decrease in the CAST/calpain ratio at the mRNA level did not lead to a corresponding change in the calpain catalytic activity. Further investigation should reveal a potential use of the CAST gene as a tool to monitor fish muscle growth and fillet firmness.
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BMC Genomics, 2009
Background To enhance capabilities for genomic analyses in rainbow trout, such as genomic selecti... more Background To enhance capabilities for genomic analyses in rainbow trout, such as genomic selection, a large suite of polymorphic markers that are amenable to high-throughput genotyping protocols must be identified. Expressed Sequence Tags (ESTs) have been used for single nucleotide polymorphism (SNP) discovery in salmonids. In those strategies, the salmonid semi-tetraploid genomes often led to assemblies of paralogous sequences and therefore resulted in a high rate of false positive SNP identification. Sequencing genomic DNA using primers identified from ESTs proved to be an effective but time consuming methodology of SNP identification in rainbow trout, therefore not suitable for high throughput SNP discovery. In this study, we employed a high-throughput strategy that used pyrosequencing technology to generate data from a reduced representation library constructed with genomic DNA pooled from 96 unrelated rainbow trout that represent the National Center for Cool and Cold Water Aquaculture (NCCCWA) broodstock population. Results The reduced representation library consisted of 440 bp fragments resulting from complete digestion with the restriction enzyme HaeIII; sequencing produced 2,000,000 reads providing an average 6 fold coverage of the estimated 150,000 unique genomic restriction fragments (300,000 fragment ends). Three independent data analyses identified 22,022 to 47,128 putative SNPs on 13,140 to 24,627 independent contigs. A set of 384 putative SNPs, randomly selected from the sets produced by the three analyses were genotyped on individual fish to determine the validation rate of putative SNPs among analyses, distinguish apparent SNPs that actually represent paralogous loci in the tetraploid genome, examine Mendelian segregation, and place the validated SNPs on the rainbow trout linkage map. Approximately 48% (183) of the putative SNPs were validated; 167 markers were successfully incorporated into the rainbow trout linkage map. In addition, 2% of the sequences from the validated markers were associated with rainbow trout transcripts. Conclusion The use of reduced representation libraries and pyrosequencing technology proved to be an effective strategy for the discovery of a high number of putative SNPs in rainbow trout; however, modifications to the technique to decrease the false discovery rate resulting from the evolutionary recent genome duplication would be desirable.
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Animal Reproduction Science, 2010
Ovarian folliculogenesis and early embryogenesis are complex processes, which require tightly reg... more Ovarian folliculogenesis and early embryogenesis are complex processes, which require tightly regulated expression and interaction of a multitude of genes. Small endogenous RNA molecules, termed microRNAs (miRNAs), are involved in the regulation of gene expression during folliculogenesis and early embryonic development. To identify miRNAs in bovine oocytes/ovaries, a bovine fetal ovary miRNA library was constructed. Sequence analysis of random clones from the library identified 679 miRNA sequences, which represent 58 distinct bovine miRNAs. Of these distinct miRNAs, 42 are known bovine miRNAs present in the miRBase database and the remaining 16 miRNAs include 15 new bovine miRNAs that are homologous to miRNAs identified in other species, and one novel miRNA, which does not match any miRNAs in the database. The precursor sequences for 14 of the new 15 miRNAs as well as the novel miRNA were identified from the bovine genome database and their hairpin structures were predicted. Expression analysis of the 58 miRNAs in fetal ovaries in comparison to somatic tissue pools identified 8 miRNAs predominantly expressed in fetal ovaries. Further analysis of the eight miRNAs in germinal vesicle (GV) stage oocytes identified two miRNAs (bta-mir424 and bta-mir-10b), that are highly abundant in GV oocytes. Both miRNAs show similar expression patterns during oocyte maturation and preimplantation development of bovine embryos, being abundant in GV and MII stage oocytes, as well as in early stage embryos (until 16-cell stage). The amount of the novel miRNA is relatively small in oocytes and early cleavage embryos but greater in blastocysts, suggesting a role of this miRNA in blastocyst cell differentiation.
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Molecular Reproduction and Development, 2007
Genes specifically expressed in oocytes are important for the development of oocytes and early em... more Genes specifically expressed in oocytes are important for the development of oocytes and early embryos. By analyzing expressed sequence tags (ESTs) from a rainbow trout oocyte cDNA library, we identified a novel EST sequence that does not show homology to any sequences in the GenBank. Analysis of tissue distribution by RT-PCR revealed that this gene was only expressed in unfertilized oocytes. Sequencing of the EST clone identified a cDNA of 3,163 bp. Northern blot analysis showed the novel gene has a single transcript of 3.4 kb. Additional 5′ sequence was obtained by 5′ RACE, extending the novel cDNA to 3,333 bp. Analysis of the full-length cDNA identified an open reading frame (ORF) encoding a protein of 564 amino acids. The novel protein contains a conserved oxysterol binding protein (OSBP) domain at the C terminus that is characteristic of OSBP-related proteins (ORPs) implicated in lipid metabolism. Therefore, we named the novel gene as Oocyte-specific Oxysterol binding protein Related-Protein of Trout (OORP-T). In situ hybridization showed that the OORP-T mRNA appears to be confined to the cytoplasm of vitellogenic oocytes. Transcription of OORP-T appears to start during pre-vitellogenesis and increases steadily, reaching its peak in the late vitellogenic stage. OORP-T transcript is abundantly present in unfertilized eggs but the level drops significantly in day 2 embryos and continues to decline in day 7 embryos after which it remains low. We propose that OORP-T may play an important role in the utilization of yolk-derived lipid products during oocyte development and early stages of embryonic development in rainbow trout. Mol. Reprod. Dev. 74: 502–511, 2007. © 2006 Wiley-Liss, Inc.
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Animal Reproduction Science, 2008
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