| Heart is the first organ to perform function during the embryonic development process in vertebrates.Although the heart structure of zebrafish(Danio rerio)is simple compared to that of mammals,zebrafish has developed as an ideal model system to studying heart development and cardiac disease because of its advantages over traditional mammalian models including transparent embryo,external development,easy observation and manipulation.Although heart development is a continuous process,there are some important events,for example,linear heart tube formation,chamber formation,heart looping and valve formation,which occur at 24,30,36 and 48 hpf in zebrafish.In this study,the intact hearts at four time points mentioned above were isolated from the Tg(myl7:EGFP)zebrafish in which cardiomyocytes were labeled with EGPF and the whole bodies without hearts 24 hpf were also collected as the control.The transcriptome profiling was performed to identify novel genes and regulatory network controlling embryonic heart development.The result demonstrated that among 32,189 genes encoded in zebrafish genome,about 8.4-10.9%(2,696-3,523)genes in heart at each time point were significantly up-regulated compared to body.GO enrichment analysis showed that the up-regulated genes were significantly enriched in biological process of cardiovascular system development(GO:0072358),heart development(GO:0007507),cardiac muscle tissue development(GO:0048738).KEGG enrichment analysis showed the up-regulated genes were significantly enriched in the pathways including Wnt signaling pathway,Notch signaling pathway and regulation of actin cytoskeleton and so on.More genes enriched in signal transduction(GO:0007165)at 30 hpf compared to other time points,suggesting that more signal pathways be activated to promote the heart looping at 36 hpf.Similar number of genes expressed between adjacent time points may promote the heart morphogenesis Many well-known transcription factors that regulate heart development such as T-box family(tbx5a,tbx20,tbx2 b and tbx1),Nkx-homeobox family(nkx2.5 and nkx2.7),GATA family(gata4,gata5,and gata6)were presented within commonly up-regulated gene sets in hearts compared to body.Thirty-four extremely highly expressed novel genes will confirmed with qRT-PCR subsequently.Congenital heart disease(CHD)is the leading cause of child disability and death.In this study,we summed up 97 human CHD causal genes and identified the orthologous genes in mice and zebrafish,and the similarity of protein sequence were compared among these genes.According to the transcriptional data The transcriptomic profilling of these tissues were performed using the Illumina platform.About 50% of the CHD orthologous genes were significantly up-regulated during the early heart development stages compared with the body.This study has important theoretical significance and practical value for genetic diagnosis of CHD disease,and the study of CHD pathogenesis with zebrafish disease model.KCTD(potassium channel tetramerization domain)proteins comprise a large family containing 26 members in human with a conserved BTB domain also known as POZ domain at C-termial and a variable C-termial.KCTD are Mutations and abnormal expression of members of KCTD gene family are associated with many diseases.In this study,orthologous genes kctd1?kctd2 and kctd4 in zebrafish of the human SEN(scalp-ear-nipple)syndrome causal gene KCTD1 and other genes KCTD2 and KCTD4 were selected,and homozygous mutants were constructed by CRISPR/Cas9 technology.According to the results of the transcriptional analysis,combined with previous reports about human disease,the orthologous genes of human glycogen storage disease(GSD),gys1,gys2 and pygma in zebrafish were selected and knocked out successfully by CRISPR/Cas9 technology.These researches provide basic information for studying the function of KCTD genes. |
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