Small RNA and mRNA expression profiling during zebrafish caudal fin regeneration
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ABSTRACT: Previous studies of zebrafish caudal fin regeneration have shown that multiple genetic programs are moduled through regulatory factors. MicroRNAs are short highly conserved non-coding genes that suppress expression of target genes and thereby control multiple genetic programs. Given their important regulatory roles and evolutionary conservation, we hypothesize that microRNAs define a conserved genetic regulatory circuit important for appendage regeneration. We characterized microRNA expression during zebrafish caudal fin regeneration using small RNA sequencing. The stages of caudal fin regeneration were assayed for mRNA expression using mRNA sequencing. Small RNA and mRNA gene expression profiling during 0 and 4 days post amputation.
Project description:Previous studies of vertebrate appendage regeneration have shown that multiple genetic programs are moduled through regulatory factors. MicroRNAs are short highly conserved non-coding genes that suppress expression of target genes and thereby control multiple genetic programs. Given their important regulatory roles and evolutionary conservation, we hypothesize that microRNAs define a conserved genetic regulatory circuit important for appendage regeneration. We characterized microRNA expression during Polypterus senegalus (bichir) pectoral fin regeneration using small RNA sequencing. The same samples were assayed for mRNA expression using mRNA sequencing. Small RNA and mRNA gene expression profiling during 0, 3, 7 and 14 days post amputation.
Project description:Previous studies of appendage regeneration in the axolotl have shown that multiple genetic programs are modulated through regulatory factors. MicroRNAs are short highly conserved non-coding genes that suppress expression of target genes and thereby control multiple genetic programs. Given their important regulatory roles and evolutionary conservation, we hypothesize that microRNAs define a conserved genetic regulatory circuit important for appendage regeneration. We characterized microRNA expression during Axolotl forelimb regeneration using small RNA sequencing. The same samples were assayed for mRNA expression using mRNA sequencing. Small RNA and mRNA gene expression profiling during 0, 3, 6 and 14 days post amputation.
Project description:Background: Adult zebrafish spontaneously regenerate their retinas after damage. Although a number of genes and signaling pathways involved in regeneration have been identified, the extent of mechanisms regulating regeneration is unclear. Small non-coding RNAs, microRNAs (miRNAs), that regulate regeneration of various tissues in lower vertebrates were examined for their potential roles in regulating zebrafish retinal regeneration. Results: To investigate the requirement of miRNAs during zebrafish retinal regeneration, we knocked down the expression of the miRNA-processing enzyme Dicer in retinas prior to light-induced damage. Dicer loss significantly reduced proliferation of Müller glia-derived neuronal progenitor cells during regeneration. To identify individual miRNAs with roles in retina regeneration, we collected retinas at different stages of light damage and performed small RNA high-throughput sequencing. We identified subsets of miRNAs that were differentially expressed during active regeneration but returned to basal levels once regeneration was completed. To validate the roles of differentially expressed miRNAs, we knocked down 6 different miRNAs that were upregulated in expression during regeneration and demonstrated that they have distinct effects on neuronal progenitor cell proliferation and migration during retina regeneration. Conclusions: miRNAs are necessary for retinal regeneration. miRNA expression is dynamic during regeneration. miRNAs function during initiation and progression of retinal regeneration. Identification of miRNAs before, during and after completion of zebrafish retinal regeneration
Project description:Previous studies of zebrafish caudal fin regeneration have shown that multiple genetic programs are moduled through regulatory factors. MicroRNAs are short highly conserved non-coding genes that suppress expression of target genes and thereby control multiple genetic programs. Given their important regulatory roles and evolutionary conservation, we hypothesize that microRNAs define a conserved genetic regulatory circuit important for appendage regeneration. We characterized microRNA expression during zebrafish caudal fin regeneration using small RNA sequencing. The stages of caudal fin regeneration were assayed for mRNA expression using mRNA sequencing.
Project description:Bone is a highly regenerative tissue but it can be permanently lost due to severe trauma and as a result of disease. In zebrafish, which are capable of complex organ regeneration, intramembranous bones of the fins and skull regenerate rapidly even after profound loss. The fin regenerate, albeit being composed of different tissues is a structure of modest complexity which, together with the fact that it is easy to access and monitor, makes it a useful system to study bone formation, wound healing and metabolic disease. Glucocorticoids are powerful drugs used in anti-inflammatory and immunosuppressive therapy. Despite their unmistakable advantages and usefulness, long term treatment with glucocorticoids can cause a variety of adverse effects such as insulin resistance, high blood pressure and bone fragility. Here we made use of proteomics to study the fin regeneration by sampling zebrafish caudal fins without prior injury and regenerated tissue 4 days post amputation at normal and perturbed conditions (prednisolone-treatment).
Project description:Adult zebrafish are able to regenerate many organs such as their caudal fin in only few days post amputation. To explore the landscape and dynamic of the genes involed in regeneration, we performed a global transcriptomic analysis using RNA-seq during zebrafish caudal fin regeneration.
Project description:Zic3 regulates early embryonic patterning in vertebrates. Its loss-of-function disrupts gastrulation, left-right patterning, and neurogenesis. We use the zebrafish as a model to study the developmental role of Zic3 in vivo. Using a combination of two genomics approaches – ChIP-seq and microarray, we identified Zic3 targets, which include genes from the Nodal and Wnt pathways, and show for the first time cis-regulation of these genes by Zic3 using in vivo enhancer assay. We uncovered a previously unrecognized link between Zic3 and the non-canonical Wnt pathway in gastrulation and left-right patterning, and identified neural pre-pattern genes as Zic3 targets during the early steps of neural induction. Zic3 preferably binds to distal intergenic regions, some of which contain evolutionarily conserved functional enhancers. Our study establishes the zebrafish as an excellent model for genome-wide study of a transcription factor in vivo. Zic3 ChIP of wild type and sqet33 transgenic
Project description:Teleost fish have the remarkable ability to regenerate their body parts including heart, spinal cord, and the caudal fin, while many higher vertebrates including us humans have only a limited ability. To facilitate molecular and genetic approaches for regeneration, we previously established an assay using the fin fold of early stage larvae, which regenerate their caudal fin folds as in adult regeneration. Here, we performed transcriptional profiling of regenerating larval fin folds and identified genes with differential expression during regeneration. Gene expression profiling of zebrafish larval fin-fold regeneration was performed by comparing amputated fin fold and uncut control. Keywords: Stress response, injury response.
Project description:Purpose: The goal of this study was to establish the first detailed cell atlas of the regenerating caudal fin of zebrafish larvae. Intact and regenerating caudal fin were used for single-cell RNA-sequencing with the aim to provide the first integrated model of epimorphic regeneration in zebrafish larvae and demonstrate the diversity of the cells required for blastema formation. Methods: 150 of regenerating caudal fin (cut) and intact caudal fine (uncut) samples were dissociated and loaded into the 10x Genomics Chromium Platform, and sequenced using Illumina NovaSeq 6000. Conclusion: Our study constitutes a resource of the gene expression profile in intact and regenerating caudal fin of zebrafish larvae. We report the application of single-molecule-based sequencing technology for high-throughput profiling of both intact (uncut) and regenerating caudal fin samples (cut) at 24hpA. We confirmed the presence of macrophage subsets, previously described by our group to govern zebrafish fin regeneration, and identified a novel blastemal cell population.
Project description:Zebrafish have the remarkable ability to regenerate body parts including the heart, spinal cord and fins by a process referred to as epimorphic regeneration. Recent studies have illustrated that similar to adult zebrafish, early life stage-larvae also possess the ability to regenerate the caudal fin. A comparative genomic analysis was used to determine the degree of conservation in gene expression among the regenerating adult caudal fin, adult heart and larval fin. Results indicate that these tissues respond to amputation/injury with strikingly similar genomic responses. Comparative analysis revealed raldh2, a rate-limiting enzyme for the synthesis of Retinoic acid (RA), as one of the highly induced genes across the three regeneration platforms. Experiment Overall Design: The caudal fin of zebrafish larvae at 2days post fertilization were amputated. Caudal fin tissue at 2dpf and regenerating fins were isolated at 1, 2and 3 days post amputation. Three replicates were collected at each time point. 150 fins were pooled to comprise one replicate.