@article {126, title = {Profiling subcellular localization of nuclear-encoded mitochondrial gene products in zebrafish.}, journal = {Life Sci Alliance}, volume = {6}, year = {2023}, month = {2023 Jan}, abstract = {

Most mitochondrial proteins are encoded by nuclear genes, synthetized in the cytosol and targeted into the organelle. To characterize the spatial organization of mitochondrial gene products in zebrafish (), we sequenced RNA from different cellular fractions. Our results confirmed the presence of nuclear-encoded mRNAs in the mitochondrial fraction, which in unperturbed conditions, are mainly transcripts encoding large proteins with specific properties, like transmembrane domains. To further explore the principles of mitochondrial protein compartmentalization in zebrafish, we quantified the transcriptomic changes for each subcellular fraction triggered by the mutation, causing the disorders in the mitochondrial protein import. Our results indicate that the proteostatic stress further restricts the population of transcripts on the mitochondrial surface, allowing only the largest and the most evolutionary conserved proteins to be synthetized there. We also show that many nuclear-encoded mitochondrial transcripts translated by the cytosolic ribosomes stay resistant to the global translation shutdown. Thus, vertebrates, in contrast to yeast, are not likely to use localized translation to facilitate synthesis of mitochondrial proteins under proteostatic stress conditions.

}, keywords = {Animals, Genes, Mitochondrial, Mitochondria, Mitochondrial Proteins, Nuclear Proteins, RNA, Messenger, Saccharomyces cerevisiae, Zebrafish}, issn = {2575-1077}, doi = {10.26508/lsa.202201514}, author = {Uszczynska-Ratajczak, Barbara and Sugunan, Sreedevi and Kwiatkowska, Monika and Migdal, Maciej and Carbonell-Sala, Silvia and Sokol, Anna and Winata, Cecilia L and Chacinska, Agnieszka} } @article {127, title = {xcore: an R package for inference of gene expression regulators.}, journal = {BMC Bioinformatics}, volume = {24}, year = {2023}, month = {2023 Jan 11}, pages = {14}, abstract = {

BACKGROUND: Elucidating the Transcription Factors (TFs) that drive the gene expression changes in a given experiment is a common question asked by researchers. The existing methods rely on the predicted Transcription Factor Binding Site (TFBS) to model the changes in the motif activity. Such methods only work for TFs that have a motif and assume the TF binding profile is the same in all cell types.

RESULTS: Given the wealth of the ChIP-seq data available for a wide range of the TFs in various cell types, we propose that gene expression modeling can be done using ChIP-seq "signatures" directly, effectively skipping the motif finding and TFBS prediction steps. We present xcore, an R package that allows TF activity modeling based on ChIP-seq signatures and the user{\textquoteright}s gene expression data. We also provide xcoredata a companion data package that provides a collection of preprocessed ChIP-seq signatures. We demonstrate that xcore leads to biologically relevant predictions using transforming growth factor beta induced epithelial-mesenchymal transition time-courses, rinderpest infection time-courses, and embryonic stem cells differentiated to cardiomyocytes time-course profiled with Cap Analysis Gene Expression.

CONCLUSIONS: xcore provides a simple analytical framework for gene expression modeling using linear models that can be easily incorporated into differential expression analysis pipelines. Taking advantage of public ChIP-seq databases, xcore can identify meaningful molecular signatures and relevant ChIP-seq experiments.

}, keywords = {Animals, Binding Sites, Chromatin Immunoprecipitation, Chromatin Immunoprecipitation Sequencing, Gene Expression, Protein Binding, Transcription Factors}, issn = {1471-2105}, doi = {10.1186/s12859-022-05084-0}, author = {Migda{\l}, Maciej and Arakawa, Takahiro and Takizawa, Satoshi and Furuno, Masaaki and Suzuki, Harukazu and Arner, Erik and Winata, Cecilia Lanny and Kaczkowski, Bogumi{\l}} } @article {124, title = {Adar-mediated A-to-I editing is required for embryonic patterning and innate immune response regulation in zebrafish.}, journal = {Nat Commun}, volume = {13}, year = {2022}, month = {2022 Sep 20}, pages = {5520}, abstract = {

Adenosine deaminases (ADARs) catalyze the deamination of adenosine to inosine, also known as A-to-I editing, in RNA. Although A-to-I editing occurs widely across animals and is well studied, new biological roles are still being discovered. Here, we study the role of A-to-I editing in early zebrafish development. We demonstrate that Adar, the zebrafish orthologue of mammalian ADAR1, is essential for establishing the antero-posterior and dorso-ventral axes and patterning. Genome-wide editing discovery reveals pervasive editing in maternal and the earliest zygotic transcripts, the majority of which occurred in the 3{\textquoteright}-UTR. Interestingly, transcripts implicated in gastrulation as well as dorso-ventral and antero-posterior patterning are found to contain multiple editing sites. Adar knockdown or overexpression affect gene expression by 12 hpf. Analysis of adar-/- zygotic mutants further reveals that the previously described role of Adar in mammals in regulating the innate immune response is conserved in zebrafish. Our study therefore establishes distinct maternal and zygotic functions of RNA editing by Adar in embryonic patterning along the zebrafish antero-posterior and dorso-ventral axes, and in the regulation of the\ innate immune response, respectively.

}, keywords = {Adenosine, Animals, Immunity, Innate, Inosine, Mammals, RNA, RNA-Binding Proteins, Zebrafish}, issn = {2041-1723}, doi = {10.1038/s41467-022-33260-6}, author = {Niescierowicz, Katarzyna and Pryszcz, Leszek and Navarrete, Cristina and Tralle, Eugeniusz and Sulej, Agata and Abu Nahia, Karim and Kasprzyk, Marta El{\.z}bieta and Misztal, Katarzyna and Pateria, Abhishek and Paku{\l}a, Adrianna and Bochtler, Matthias and Winata, Cecilia} } @article {125, title = {Multiomic atlas with functional stratification and developmental dynamics of zebrafish cis-regulatory elements.}, journal = {Nat Genet}, volume = {54}, year = {2022}, month = {2022 Jul}, pages = {1037-1050}, abstract = {

Zebrafish, a popular organism for studying embryonic development and for modeling human diseases, has so far lacked a systematic functional annotation program akin to those in other animal models. To address this, we formed the international DANIO-CODE consortium and created a central repository to store and process zebrafish developmental functional genomic data. Our data coordination center ( https://danio-code.zfin.org ) combines a total of 1,802 sets of unpublished and re-analyzed published genomic data, which we used to improve existing annotations and show its utility in experimental design. We identified over 140,000 cis-regulatory elements throughout development, including classes with distinct features dependent on their activity in time and space. We delineated the distinct distance topology and chromatin features between regulatory elements active during zygotic genome activation and those active during organogenesis. Finally, we matched regulatory elements and epigenomic landscapes between zebrafish and mouse and predicted functional relationships between them beyond sequence similarity, thus extending the utility of zebrafish developmental genomics to mammals.

}, keywords = {Animals, Chromatin, Databases, Genetic, Gene Expression Regulation, Developmental, Genome, Genomics, Humans, Mice, Molecular Sequence Annotation, Organogenesis, Regulatory Sequences, Nucleic Acid, Zebrafish, Zebrafish Proteins}, issn = {1546-1718}, doi = {10.1038/s41588-022-01089-w}, author = {Baranasic, Damir and H{\"o}rtenhuber, Matthias and Balwierz, Piotr J and Zehnder, Tobias and Mukarram, Abdul Kadir and Nepal, Chirag and V{\'a}rnai, Csilla and Hadzhiev, Yavor and Jimenez-Gonzalez, Ada and Li, Nan and Wragg, Joseph and D{\textquoteright}Orazio, Fabio M and Relic, Dorde and Pachkov, Mikhail and D{\'\i}az, Noelia and Hern{\'a}ndez-Rodr{\'\i}guez, Benjam{\'\i}n and Chen, Zelin and Stoiber, Marcus and Dong, Micha{\"e}l and Stevens, Irene and Ross, Samuel E and Eagle, Anne and Martin, Ryan and Obasaju, Oluwapelumi and Rastegar, Sepand and McGarvey, Alison C and Kopp, Wolfgang and Chambers, Emily and Wang, Dennis and Kim, Hyejeong R and Acemel, Rafael D and Naranjo, Silvia and {\L}api{\'n}ski, Maciej and Chong, Vanessa and Mathavan, Sinnakaruppan and Peers, Bernard and Sauka-Spengler, Tatjana and Vingron, Martin and Carninci, Piero and Ohler, Uwe and Lacadie, Scott Allen and Burgess, Shawn M and Winata, Cecilia and van Eeden, Freek and Vaquerizas, Juan M and G{\'o}mez-Skarmeta, Jos{\'e} Luis and Onichtchouk, Daria and Brown, Ben James and Bogdanovic, Ozren and van Nimwegen, Erik and Westerfield, Monte and Wardle, Fiona C and Daub, Carsten O and Lenhard, Boris and M{\"u}ller, Ferenc} } @article {119, title = {Cardiac-specific β-catenin deletion dysregulates energetic metabolism and mitochondrial function in perinatal cardiomyocytes.}, journal = {Mitochondrion}, volume = {60}, year = {2021}, month = {2021 09}, pages = {59-69}, abstract = {

β-Catenin signaling pathway regulates cardiomyocytes proliferation and differentiation, though its involvement in metabolic regulation of cardiomyocytes remains unknown. We used one-day-old mice with cardiac-specific knockout of β-catenin and neonatal rat ventricular myocytes treated with β-catenin inhibitor to investigate the role of β-catenin metabolism regulation in perinatal cardiomyocytes. Transcriptomics of perinatal β-catenin-ablated hearts revealed a dramatic shift in the expression of genes involved in metabolic processes. Further analysis indicated an inhibition of lipolysis and glycolysis in both in vitro and in vivo models. Finally, we showed that β-catenin deficiency leads to mitochondria dysfunction via the downregulation of Sirt1/PGC-1α pathway. We conclude that cardiac-specific β-catenin ablation disrupts the energy substrate shift that is essential for postnatal heart maturation, leading to perinatal lethality of homozygous β-catenin knockout mice.

}, issn = {1872-8278}, doi = {10.1016/j.mito.2021.07.005}, author = {Balatskyi, Volodymyr V and Vaskivskyi, Vasyl O and Myronova, Anna and Avramets, Diana and Nahia, Karim Abu and Macewicz, Larysa L and Ruban, Tetiana P and Kucherenko, Dar{\textquoteright}ya Yu and Soldatkin, Oleksandr O and Lushnikova, Iryna V and Skibo, Galyna G and Winata, Cecilia L and Dobrzyn, Pawel and Piven, Oksana O} } @article {117, title = {Exploring Translational Control of Maternal mRNAs in Zebrafish.}, journal = {Methods Mol Biol}, volume = {2218}, year = {2021}, month = {2021}, pages = {367-380}, abstract = {

The study of translational regulation requires reliable measurement of both mRNA levels and protein synthesis. Cytoplasmic polyadenylation is a prevalent mode of translational regulation during oogenesis and early embryogenesis. Here the length of the poly(A) tail of an mRNA is coupled to its translatability. We describe a protocol to identify translationally regulated genes and measure their translation rate in the early zebrafish embryo using genome-wide polysome profiling. This protocol relies on the isolation of mRNA by means of an rRNA depletion strategy, which avoids capture bias due to short poly(A) tail that can occur when using conventional oligo(dT)-based methods. We also present a simple PCR-based method to measure the poly(A) tail length of selected mRNAs.

}, keywords = {Animals, Cytoplasm, Embryo, Nonmammalian, Embryonic Development, Oocytes, Oogenesis, Poly A, Polyadenylation, Protein Biosynthesis, RNA, Messenger, Stored, Zebrafish}, issn = {1940-6029}, doi = {10.1007/978-1-0716-0970-5_29}, author = {Winata, Cecilia Lanny and {\L}api{\'n}ski, Maciej and Ismail, Hisyam and Mathavan, Sinnakaruppan and Sampath, Prabha} } @article {120, title = {Genomic and physiological analyses of the zebrafish atrioventricular canal reveal molecular building blocks of the secondary pacemaker region.}, journal = {Cell Mol Life Sci}, volume = {78}, year = {2021}, month = {2021 Oct}, pages = {6669-6687}, abstract = {

The atrioventricular canal (AVC) is the site where key structures responsible for functional division between heart regions are established, most importantly, the atrioventricular (AV) conduction system and cardiac valves. To elucidate the mechanism underlying AVC development and function, we utilized transgenic zebrafish line sqet31Et expressing EGFP in the AVC to isolate this cell population and profile its transcriptome at 48 and 72 hpf. The zebrafish AVC transcriptome exhibits hallmarks of mammalian AV node, including the expression of genes implicated in its development and those encoding connexins forming low conductance gap junctions. Transcriptome analysis uncovered protein-coding and noncoding transcripts enriched in AVC, which have not been previously associated with this structure, as well as dynamic expression of epithelial-to-mesenchymal transition markers and components of TGF-β, Notch, and Wnt signaling pathways likely reflecting ongoing AVC and valve development. Using transgenic line Tg(myl7:mermaid) encoding voltage-sensitive fluorescent protein, we show that abolishing the pacemaker-containing sinoatrial ring (SAR) through Isl1 loss of function resulted in spontaneous activation in the AVC region, suggesting that it possesses inherent automaticity although insufficient to replace the SAR. The SAR and AVC transcriptomes express partially overlapping species of ion channels and gap junction proteins, reflecting their distinct roles. Besides identifying conserved aspects between zebrafish and mammalian conduction systems, our results established molecular hallmarks of the developing AVC which underlies its role in structural and electrophysiological separation between heart chambers. This data constitutes a valuable resource for studying AVC development and function, and identification of novel candidate genes implicated in these processes.

}, keywords = {Animals, Animals, Genetically Modified, Embryo, Nonmammalian, Gene Expression Regulation, Developmental, Genome, Genomics, Heart Septal Defects, Heart Valves, Myocardium, Organogenesis, Pacemaker, Artificial, Wnt Signaling Pathway, Zebrafish, Zebrafish Proteins}, issn = {1420-9071}, doi = {10.1007/s00018-021-03939-y}, author = {Abu Nahia, Karim and Migda{\l}, Maciej and Quinn, T Alexander and Poon, Kar-Lai and {\L}api{\'n}ski, Maciej and Sulej, Agata and Liu, Jiandong and Mondal, Shamba S and Pawlak, Micha{\l} and Bugajski, {\L}ukasz and Piwocka, Katarzyna and Brand, Thomas and Kohl, Peter and Korzh, Vladimir and Winata, Cecilia} } @article {122, title = {Multi-omics analyses of early liver injury reveals cell-type-specific transcriptional and epigenomic shift.}, journal = {BMC Genomics}, volume = {22}, year = {2021}, month = {2021 Dec 18}, pages = {904}, abstract = {

BACKGROUND: Liver fibrosis is a wound-healing response to tissue injury and inflammation hallmarked by the extracellular matrix (ECM) protein deposition in the liver parenchyma and tissue remodelling. Different cell types of the liver are known to play distinct roles in liver injury response. Hepatocytes and liver endothelial cells receive molecular signals indicating tissue injury and activate hepatic stellate cells which produce ECM proteins upon their activation. Despite the growing knowledge on the molecular mechanism underlying hepatic fibrosis in general, the cell-type-specific gene regulatory network associated with the initial response to hepatotoxic injury is still poorly characterized.

RESULTS: In this study, we used thioacetamide (TAA) to induce hepatic injury in adult zebrafish. We isolated three major liver cell types - hepatocytes, endothelial cells and hepatic stellate cells - and identified cell-type-specific chromatin accessibility and transcriptional changes in an early stage of liver injury. We found that TAA induced transcriptional shifts in all three cell types hallmarked by significant alterations in the expression of genes related to fatty acid and carbohydrate metabolism, as well as immune response-associated and vascular-specific genes. Interestingly, liver endothelial cells exhibit the most pronounced response to liver injury at the transcriptome and chromatin level, hallmarked by the loss of their angiogenic phenotype.

CONCLUSION: Our results uncovered cell-type-specific transcriptome and epigenome responses to early stage liver injury, which provide valuable insights into understanding the molecular mechanism implicated in the early response of the liver to pro-fibrotic signals.

}, keywords = {Animals, Endothelial Cells, Epigenomics, Liver, Zebrafish}, issn = {1471-2164}, doi = {10.1186/s12864-021-08173-1}, author = {Migda{\l}, Maciej and Tralle, Eugeniusz and Nahia, Karim Abu and Bugajski, {\L}ukasz and K{\k e}dzierska, Katarzyna Zofia and Garbicz, Filip and Piwocka, Katarzyna and Winata, Cecilia Lanny and Pawlak, Micha{\l}} } @article {121, title = {Transcriptome profile of the sinoatrial ring reveals conserved and novel genetic programs of the zebrafish pacemaker.}, journal = {BMC Genomics}, volume = {22}, year = {2021}, month = {2021 Oct 02}, pages = {715}, abstract = {

BACKGROUND: Sinoatrial Node (SAN) is part of the cardiac conduction system, which controls the rhythmic contraction of the vertebrate heart. The SAN consists of a specialized pacemaker cell population that has the potential to generate electrical impulses. Although the SAN pacemaker has been extensively studied in mammalian and teleost models, including the zebrafish, their molecular nature remains inadequately comprehended.

RESULTS: To characterize the molecular profile of the zebrafish sinoatrial ring (SAR) and elucidate the mechanism of pacemaker function, we utilized the transgenic line sqet33mi59BEt to isolate cells of the SAR of developing zebrafish embryos and profiled their transcriptome. Our analyses identified novel candidate genes and well-known conserved signaling pathways involved in pacemaker development. We show that, compared to the rest of the heart, the zebrafish SAR overexpresses several mammalian SAN pacemaker signature genes, which include hcn4 as well as those encoding calcium- and potassium-gated channels. Moreover, genes encoding components of the BMP and Wnt signaling pathways, as well as members of the Tbx family, which have previously been implicated in pacemaker development, were also overexpressed in the SAR. Among SAR-overexpressed genes, 24 had human homologues implicated in 104 different ClinVar phenotype entries related to various forms of congenital heart diseases, which suggest the relevance of our transcriptomics resource to studying human heart conditions. Finally, functional analyses of three SAR-overexpressed genes, pard6a, prom2, and atp1a1a.2, uncovered their novel role in heart development and physiology.

CONCLUSION: Our results established conserved aspects between zebrafish and mammalian pacemaker function and revealed novel factors implicated in maintaining cardiac rhythm. The transcriptome data generated in this study represents a unique and valuable resource for the study of pacemaker function and associated heart diseases.

}, keywords = {Animals, Heart Rate, Humans, Sinoatrial Node, Transcriptome, Zebrafish}, issn = {1471-2164}, doi = {10.1186/s12864-021-08016-z}, author = {Minhas, Rashid and Loeffler-Wirth, Henry and Siddiqui, Yusra H and Obr{\k e}bski, Tomasz and Vashisht, Shikha and Nahia, Karim Abu and Paterek, Alexandra and Brzozowska, Angelika and Bugajski, Lukasz and Piwocka, Katarzyna and Korzh, Vladimir and Binder, Hans and Winata, Cecilia Lanny} } @article {112, title = {Dynamics of cardiomyocyte transcriptome and chromatin landscape demarcates key events of heart development.}, journal = {Genome Res}, volume = {29}, year = {2019}, month = {2019 03}, pages = {506-519}, abstract = {

Organogenesis involves dynamic regulation of gene transcription and complex multipathway interactions. Despite our knowledge of key factors regulating various steps of heart morphogenesis, considerable challenges in understanding its mechanism still exist because little is known about their downstream targets and interactive regulatory network. To better understand transcriptional regulatory mechanism driving heart development and the consequences of its disruption in vivo, we performed time-series analyses of the transcriptome and genome-wide chromatin accessibility in isolated cardiomyocytes (CMs) from wild-type zebrafish embryos at developmental stages corresponding to heart tube morphogenesis, looping, and maturation. We identified genetic regulatory modules driving crucial events of heart development that contained key cardiac TFs and are associated with open chromatin regions enriched for DNA sequence motifs belonging to the family of the corresponding TFs. Loss of function of cardiac TFs Gata5, Tbx5a, and Hand2 affected the cardiac regulatory networks and caused global changes in chromatin accessibility profile, indicating their role in heart development. Among regions with differential chromatin accessibility in mutants were highly conserved noncoding elements that represent putative enhancers driving heart development. The most prominent gene expression changes, which correlated with chromatin accessibility modifications within their proximal promoter regions, occurred between heart tube morphogenesis and looping, and were associated with metabolic shift and hematopoietic/cardiac fate switch during CM maturation. Our results revealed the dynamic regulatory landscape throughout heart development and identified interactive molecular networks driving key events of heart morphogenesis.

}, keywords = {Animals, Cells, Cultured, Chromatin, Chromatin Assembly and Disassembly, Gene Expression Regulation, Developmental, Gene Regulatory Networks, Heart, Myocytes, Cardiac, Transcription Factors, Transcriptome, Zebrafish, Zebrafish Proteins}, issn = {1549-5469}, doi = {10.1101/gr.244491.118}, author = {Pawlak, Michal and Kedzierska, Katarzyna Z and Migdal, Maciej and Nahia, Karim Abu and Ramilowski, Jordan A and Bugajski, Lukasz and Hashimoto, Kosuke and Marconi, Aleksandra and Piwocka, Katarzyna and Carninci, Piero and Winata, Cecilia L} } @article {113, title = {A novel conserved enhancer at zebrafish zic3 and zic6 loci drives neural expression.}, journal = {Dev Dyn}, volume = {248}, year = {2019}, month = {2019 09}, pages = {837-849}, abstract = {

BACKGROUND: Identifying enhancers and deciphering their putative roles represent a major step to better understand the mechanism of metazoan gene regulation, development, and the role of regulatory elements in disease. Comparative genomics and transgenic assays have been used with some success to identify critical regions that are involved in regulating the spatiotemporal expression of genes during embryogenesis.

RESULTS: We identified two novel tetrapod-teleost conserved noncoding elements within the vicinity of the zic3 and zic6 loci in the zebrafish genome and demonstrated their ability to drive tissue-specific expression in a transgenic zebrafish assay. The syntenic analysis and robust green fluorescent expression in the developing habenula in the stable transgenic line were correlated with known sites of endogenous zic3 and zic6 expression.

CONCLUSION: This transgenic line that expresses green fluorescent protein in the habenula is a valuable resource for studying a specific population of cells in the zebrafish central nervous system. Our observations indicate that a genomic sequence that is conserved between humans and zebrafish acts as an enhancer that likely controls zic3 and zic6 expression.

}, keywords = {Animals, Animals, Genetically Modified, Conserved Sequence, Embryonic Development, Enhancer Elements, Genetic, Gene Expression Regulation, Developmental, Green Fluorescent Proteins, Habenula, Homeodomain Proteins, Humans, Nervous System, Repressor Proteins, Transcription Factors, Zebrafish, Zebrafish Proteins}, issn = {1097-0177}, doi = {10.1002/dvdy.69}, author = {Minhas, Rashid and Paterek, Aleksandra and {\L}api{\'n}ski, Maciej and Baza{\l}a, Micha{\l} and Korzh, Vladimir and Winata, Cecilia L} } @article {pmid29229769, title = {Cytoplasmic polyadenylation-mediated translational control of maternal mRNAs directs maternal-to-zygotic transition}, journal = {Development}, volume = {145}, year = {2018}, month = {01/2018}, abstract = {

In the earliest stages of animal development following fertilization, maternally deposited mRNAs direct biological processes to the point of zygotic genome activation (ZGA). These maternal mRNAs undergo cytoplasmic polyadenylation (CPA), suggesting translational control of their activation. To elucidate the biological role of CPA during embryogenesis, we performed genome-wide polysome profiling at several stages of zebrafish development. Our analysis revealed a correlation between CPA and polysome-association dynamics, demonstrating a coupling of translation to the CPA of maternal mRNAs. Pan-embryonic CPA inhibition disrupted the maternal-to-zygotic transition (MZT), causing a failure of developmental progression beyond the mid-blastula transition and changes in global gene expression that indicated a failure of ZGA and maternal mRNA clearance. Among the genes that were differentially expressed were those encoding chromatin modifiers and key transcription factors involved in ZGA, including nanog, pou5f3 and sox19b, which have distinct CPA dynamics. Our results establish the necessity of CPA for ensuring progression of the MZT. The RNA-seq data generated in this study represent a valuable zebrafish resource for the discovery of novel elements of the early embryonic transcriptome.

}, author = {Winata, C. L. and {\L}api{\'n}ski, M. and Pryszcz, L. and Vaz, C. and Bin Ismail, M. H. and Nama, S. and Hajan, H. S. and Lee, S. G. P. and Korzh, V. and Sampath, P. and Tanavde, V. and Mathavan, S.} } @article {74, title = {Impaired development of neural-crest cell-derived organs and intellectual disability caused by MED13L haploinsufficiency.}, journal = {Hum Mutat}, volume = {35}, year = {2014}, month = {2014 Nov}, pages = {1311-20}, abstract = {

MED13L is a component subunit of the Mediator complex, an important regulator of transcription that is highly conserved across eukaryotes. Here, we report MED13L disruption in a translocation t(12;19) breakpoint of a patient with Pierre-Robin syndrome, moderate intellectual disability, craniofacial anomalies, and muscular defects. The phenotype is similar to previously described patients with MED13L haploinsufficiency. Knockdown of MED13L orthologue in zebrafish, med13b, showed early defective migration of cranial neural crest cells (NCCs) that contributed to cartilage structure deformities in the later stage, recapitulating craniofacial anomalies seen in human patients. Notably, we observed abnormal distribution of developing neurons in different brain regions of med13b morphant embryos, which could be rescued upon introduction of full-length human MED13L mRNA. To compare with mammalian system, we suppressed MED13L expression by short-hairpin RNA in ES-derived human neural progenitors, and differentiated them into neurons. Transcriptome analysis revealed differential expression of components of Wnt and FGF signaling pathways in MED13L-deficient neurons. Our finding provides a novel insight into the mechanism of overlapping phenotypic outcome targeting NCCs derivatives organs in patients with MED13L haploinsufficiency, and emphasizes a clinically recognizable syndromic phenotype in these patients.

}, keywords = {Animals, Cell Differentiation, Cell Movement, Child, Preschool, Chromosome Breakpoints, Disease Models, Animal, Embryonic Stem Cells, Female, Gene Expression, Gene Expression Profiling, Gene Expression Regulation, Developmental, Gene Knockdown Techniques, Genetic Association Studies, Haploinsufficiency, Humans, Intellectual Disability, Mediator Complex, Neural Crest, Neurons, Phenotype, RNA, Messenger, Sequence Analysis, DNA, Transcriptome, Translocation, Genetic, Zebrafish}, issn = {1098-1004}, doi = {10.1002/humu.22636}, author = {Utami, Kagistia Hana and Winata, Cecilia L and Hillmer, Axel M and Aksoy, Irene and Long, Hoang Truong and Liany, Herty and Chew, Elaine G Y and Mathavan, Sinnakaruppan and Tay, Stacey K H and Korzh, Vladimir and Sarda, Pierre and Davila, Sonia and Cacheux, Valere} } @article {32, title = {Normalization of RNA-sequencing data from samples with varying mRNA levels.}, journal = {PLoS One}, volume = {9}, year = {2014}, month = {2014}, pages = {e89158}, abstract = {

Methods for normalization of RNA-sequencing gene expression data commonly assume equal total expression between compared samples. In contrast, scenarios of global gene expression shifts are many and increasing. Here we compare the performance of three normalization methods when polyA(+) RNA content fluctuates significantly during zebrafish early developmental stages. As a benchmark we have used reverse transcription-quantitative PCR. The results show that reads per kilobase per million (RPKM) and trimmed mean of M-values (TMM) normalization systematically leads to biased gene expression estimates. Biological scaling normalization (BSN), designed to handle differences in total expression, showed improved accuracy compared to the two other methods in estimating transcript level dynamics. The results have implications for past and future studies using RNA-sequencing on samples with different levels of total or polyA(+) RNA.

}, keywords = {Animals, Base Sequence, Gene Expression, Gene Expression Profiling, Polymerase Chain Reaction, RNA, Messenger, Sequence Analysis, RNA, Zebrafish}, issn = {1932-6203}, doi = {10.1371/journal.pone.0089158}, author = {Aanes, H{\r a}vard and Winata, Cecilia L and Moen, Lars F and {\O}strup, Olga and Mathavan, Sinnakaruppan and Collas, Philippe and Rognes, Torbj{\o}rn and Alestr{\"o}m, Peter} } @article {33, title = {Genome wide analysis reveals Zic3 interaction with distal regulatory elements of stage specific developmental genes in zebrafish.}, journal = {PLoS Genet}, volume = {9}, year = {2013}, month = {2013 Oct}, pages = {e1003852}, abstract = {

Zic3 regulates early embryonic patterning in vertebrates. Loss of Zic3 function is known to disrupt gastrulation, left-right patterning, and neurogenesis. However, molecular events downstream of this transcription factor are poorly characterized. Here we use the zebrafish as a model to study the developmental role of Zic3 in vivo, by applying a combination of two powerful genomics approaches--ChIP-seq and microarray. Besides confirming direct regulation of previously implicated Zic3 targets of the Nodal and canonical Wnt pathways, analysis of gastrula stage embryos uncovered a number of novel candidate target genes, among which were members of the non-canonical Wnt pathway and the neural pre-pattern genes. A similar analysis in zic3-expressing cells obtained by FACS at segmentation stage revealed a dramatic shift in Zic3 binding site locations and identified an entirely distinct set of target genes associated with later developmental functions such as neural development. We demonstrate cis-regulation of several of these target genes by Zic3 using in vivo enhancer assay. Analysis of Zic3 binding sites revealed a distribution biased towards distal intergenic regions, indicative of a long distance regulatory mechanism; some of these binding sites are highly conserved during evolution and act as functional enhancers. This demonstrated that Zic3 regulation of developmental genes is achieved predominantly through long distance regulatory mechanism and revealed that developmental transitions could be accompanied by dramatic changes in regulatory landscape.

}, issn = {1553-7404}, doi = {10.1371/journal.pgen.1003852}, author = {Winata, Cecilia L and Kondrychyn, Igor and Kumar, Vibhor and Srinivasan, Kandhadayar G and Orlov, Yuriy and Ravishankar, Ashwini and Prabhakar, Shyam and Stanton, Lawrence W and Korzh, Vladimir and Mathavan, Sinnakaruppan} } @article {34, title = {Prepatterning of developmental gene expression by modified histones before zygotic genome activation.}, journal = {Dev Cell}, volume = {21}, year = {2011}, month = {2011 Dec 13}, pages = {993-1004}, abstract = {

A hallmark of anamniote vertebrate development is a window of embryonic transcription-independent cell divisions before onset of zygotic genome activation (ZGA). Chromatin determinants of ZGA are unexplored; however, marking of developmental genes by modified histones in sperm suggests a predictive role of histone marks for ZGA. In zebrafish, pre-ZGA development for ten cell cycles provides an opportunity to examine whether genomic enrichment in modified histones is present before initiation of transcription. By profiling histone H3 trimethylation on all zebrafish promoters before and after ZGA, we demonstrate here an epigenetic prepatterning of developmental gene expression. This involves pre-ZGA marking of transcriptionally inactive genes involved in homeostatic and developmental regulation by permissive H3K4me3 with or without repressive H3K9me3 or H3K27me3. Our data suggest that histone modifications are instructive for the developmental gene expression program.

}, keywords = {Animals, Body Patterning, Chromatin, Epigenesis, Genetic, Female, Gene Expression Regulation, Developmental, Histones, Male, Methylation, Multigene Family, Promoter Regions, Genetic, Spermatozoa, Zebrafish, Zebrafish Proteins}, issn = {1878-1551}, doi = {10.1016/j.devcel.2011.10.008}, author = {Lindeman, Leif C and Andersen, Ingrid S and Reiner, Andrew H and Li, Nan and Aanes, H{\r a}vard and {\O}strup, Olga and Winata, Cecilia L and Mathavan, Sinnakaruppan and M{\"u}ller, Ferenc and Alestr{\"o}m, Peter and Collas, Philippe} } @article {36, title = {Zebrafish mRNA sequencing deciphers novelties in transcriptome dynamics during maternal to zygotic transition.}, journal = {Genome Res}, volume = {21}, year = {2011}, month = {2011 Aug}, pages = {1328-38}, abstract = {

Maternally deposited mRNAs direct early development before the initiation of zygotic transcription during mid-blastula transition (MBT). To study mechanisms regulating this developmental event in zebrafish, we applied mRNA deep sequencing technology and generated comprehensive information and valuable resources on transcriptome dynamics during early embryonic (egg to early gastrulation) stages. Genome-wide transcriptome analysis documented at least 8000 maternal genes and identified the earliest cohort of zygotic transcripts. We determined expression levels of maternal and zygotic transcripts with the highest resolution possible using mRNA-seq and clustered them based on their expression pattern. We unravel delayed polyadenylation in a large cohort of maternal transcripts prior to the MBT for the first time in zebrafish. Blocking polyadenylation of these transcripts confirms their role in regulating development from the MBT onward. Our study also identified a large number of novel transcribed regions in annotated and unannotated regions of the genome, which will facilitate reannotation of the zebrafish genome. We also identified splice variants with an estimated frequency of 50\%-60\%. Taken together, our data constitute a useful genomic information and valuable transcriptome resource for gene discovery and for understanding the mechanisms of early embryogenesis in zebrafish.

}, keywords = {Animals, Base Sequence, Genome, RNA, Messenger, RNA, Messenger, Stored, Sequence Analysis, RNA, Transcriptome, Zebrafish, Zebrafish Proteins, Zygote}, issn = {1549-5469}, doi = {10.1101/gr.116012.110}, author = {Aanes, H{\r a}vard and Winata, Cecilia L and Lin, Chi Ho and Chen, Jieqi P and Srinivasan, Kandhadayar G and Lee, Serene G P and Lim, Adrian Y M and Hajan, Hajira Shreen and Collas, Philippe and Bourque, Guillaume and Gong, Zhiyuan and Korzh, Vladimir and Alestr{\"o}m, Peter and Mathavan, Sinnakaruppan} } @article {69, title = {Chromatin states of developmentally-regulated genes revealed by DNA and histone methylation patterns in zebrafish embryos.}, journal = {Int J Dev Biol}, volume = {54}, year = {2010}, month = {2010}, pages = {803-13}, abstract = {

Embryo development proceeds from a cascade of gene activation and repression events controlled by epigenetic modifications of DNA and histones. Little is known about epigenetic states in the developing zebrafish, despite its importance as a model organism. We report here DNA methylation and histone modification profiles of promoters of developmentally-regulated genes (pou5f1, sox2, sox3, klf4, nnr, otx1b, nes, vasa), as well as tert and bactin2, in zebrafish embryos at the mid-late blastula transition, shortly after embryonic genome activation. We identify four classes of promoters based on the following profiles: (i) those enriched in marks of active genes (H3K9ac, H4ac, H3K4me3) without transcriptionally repressing H3K9me3 or H3K27me3; (ii) those enriched in H3K9ac, H4ac and H3K27me3, without H3K9me3; one such gene was klf4, shown by in situ hybridization to be mosaically expressed, likely accounting for the detection of both activating and repressive marks on its promoter; (iii) those enriched in H3K4me3 and H3K27me3 without acetylation; and (iv) those enriched in all histone modifications examined. Culture of embryo-derived cells under differentiation conditions leads to H3K9 and H4 deacetylation and H3K9 and H3K27 trimethylation on genes that are inactivated, yielding an epigenetic profile similar to those of fibroblasts or muscle. All promoters however retain H3K4me3, indicating an uncoupling of H3K4me3 occupancy and gene expression. All non-CpG island developmentally-regulated promoters are DNA unmethylated in embryos, but hypermethylated in fibroblasts. Our results suggest that differentially expressed embryonic genes are regulated by various patterns of histone modifications on unmethylated DNA, which create a developmentally permissive chromatin state.

}, keywords = {Animals, Blastula, Cell Line, Chromatin, Chromatin Immunoprecipitation, CpG Islands, DNA Methylation, Embryo, Nonmammalian, Fibroblasts, Gene Expression Profiling, Gene Expression Regulation, Developmental, Histones, In Situ Hybridization, Lysine, Methylation, Oligonucleotide Array Sequence Analysis, Promoter Regions, Genetic, Reverse Transcriptase Polymerase Chain Reaction, Zebrafish, Zebrafish Proteins}, issn = {1696-3547}, doi = {10.1387/ijdb.103081ll}, author = {Lindeman, Leif C and Winata, Cecilia L and Aanes, Hvard and Mathavan, Sinnakaruppan and Alestrom, Peter and Collas, Philippe} } @article {70, title = {Mercury-induced hepatotoxicity in zebrafish: in vivo mechanistic insights from transcriptome analysis, phenotype anchoring and targeted gene expression validation.}, journal = {BMC Genomics}, volume = {11}, year = {2010}, month = {2010}, pages = {212}, abstract = {

BACKGROUND: Mercury is a prominent environmental contaminant that causes detrimental effects to human health. Although the liver has been known to be a main target organ, there is limited information on in vivo molecular mechanism of mercury-induced toxicity in the liver. By using transcriptome analysis, phenotypic anchoring and validation of targeted gene expression in zebrafish, mercury-induced hepatotoxicity was investigated and a number of perturbed cellular processes were identified and compared with those captured in the in vitro human cell line studies.

RESULTS: Hepato-transcriptome analysis of mercury-exposed zebrafish revealed that the earliest deregulated genes were associated with electron transport chain, mitochondrial fatty acid beta-oxidation, nuclear receptor signaling and apoptotic pathway, followed by complement system and proteasome pathway, and thereafter DNA damage, hypoxia, Wnt signaling, fatty acid synthesis, gluconeogenesis, cell cycle and motility. Comparative meta-analysis of microarray data between zebrafish liver and human HepG2 cells exposed to mercury identified some common toxicological effects of mercury-induced hepatotoxicity in both models. Histological analyses of liver from mercury-exposed fish revealed morphological changes of liver parenchyma, decreased nucleated cell count, increased lipid vesicles, glycogen and apoptotic bodies, thus providing phenotypic evidence for anchoring of the transcriptome analysis. Validation of targeted gene expression confirmed deregulated gene-pathways from enrichment analysis. Some of these genes responding to low concentrations of mercury may serve as toxicogenomic-based markers for detection and health risk assessment of environmental mercury contaminations.

CONCLUSION: Mercury-induced hepatotoxicity was triggered by oxidative stresses, intrinsic apoptotic pathway, deregulation of nuclear receptor and kinase activities including Gsk3 that deregulates Wnt signaling pathway, gluconeogenesis, and adipogenesis, leading to mitochondrial dysfunction, endocrine disruption and metabolic disorders. This study provides important mechanistic insights into mercury-induced liver toxicity in a whole-animal physiology context, which will help in understanding the syndromes caused by mercury poisoning. The molecular conservation of mercury-induced hepatotoxicity between zebrafish and human cell line reveals the feasibility of using zebrafish to model molecular toxicity in human for toxicant risk assessments.

}, keywords = {Animals, Apoptosis, Arsenic, Cell Adhesion, Cell Line, Gene Expression Profiling, Hepatocytes, Humans, Liver, Mercury, Oligonucleotide Array Sequence Analysis, Zebrafish}, issn = {1471-2164}, doi = {10.1186/1471-2164-11-212}, author = {Ung, Choong Yong and Lam, Siew Hong and Hlaing, Mya Myintzu and Winata, Cecilia L and Korzh, Svetlana and Mathavan, Sinnakaruppan and Gong, Zhiyuan} } @article {65, title = {Transcriptome kinetics of arsenic-induced adaptive response in zebrafish liver.}, journal = {Physiol Genomics}, volume = {27}, year = {2006}, month = {2006 Nov 27}, pages = {351-61}, abstract = {

Arsenic is a prominent environmental toxicant and carcinogen; however, its molecular mechanism of toxicity and carcinogenicity remains poorly understood. In this study, we performed microarray-based expression profiling on liver of zebrafish exposed to 15 parts/million (ppm) arsenic [As(V)] for 8-96 h to identify global transcriptional changes and biological networks involved in arsenic-induced adaptive responses in vivo. We found that there was an increase of transcriptional activity associated with metabolism, especially for biosyntheses, membrane transporter activities, cytoplasm, and endoplasmic reticulum in the 96 h of arsenic treatment, while transcriptional programs for proteins in catabolism, energy derivation, and stress response remained active throughout the arsenic treatment. Many differentially expressed genes encoding proteins involved in heat shock proteins, DNA damage/repair, antioxidant activity, hypoxia induction, iron homeostasis, arsenic metabolism, and ubiquitin-dependent protein degradation were identified, suggesting strongly that DNA and protein damage as a result of arsenic metabolism and oxidative stress caused major cellular injury. These findings were comparable with those reported in mammalian systems, suggesting that the zebrafish liver coupled with the available microarray technology present an excellent in vivo toxicogenomic model for investigating arsenic toxicity. We proposed an in vivo, acute arsenic-induced adaptive response model of the zebrafish liver illustrating the relevance of many transcriptional activities that provide both global and specific information of a coordinated adaptive response to arsenic in the liver.

}, keywords = {Adaptation, Physiological, Animals, Arsenic, Down-Regulation, Gene Expression Profiling, Gene Expression Regulation, Genomics, Liver, Male, Metabolic Networks and Pathways, Oligonucleotide Array Sequence Analysis, Transcription, Genetic, Up-Regulation, Zebrafish}, issn = {1531-2267}, doi = {10.1152/physiolgenomics.00201.2005}, author = {Lam, Siew Hong and Winata, Cecilia L and Tong, Yan and Korzh, Svetlana and Lim, Wen San and Korzh, Vladimir and Spitsbergen, Jan and Mathavan, Sinnakarupan and Miller, Lance D and Liu, Edison T and Gong, Zhiyuan} }