Project description:Among the Chordates tunicates demonstrate the highest capacity for regeneration, ranging from full-body regeneration in colonial ascidians to appendage regeneration in solitary ascidians. Here we present a parallel study of mRNA and microRNA expression at three stages of oral siphon regeneration in the solitary ascidian Ciona robusta (a.k.a., Ciona intestinalis), and the derived network of their interactions. In the process of identifying 248 mRNAs and 15 microRNAs as differentially expressed (DE) across the course of regeneration, we also identified 57 novel microRNAs, several of which are among the most highly DE. Analysis functional categories identified transcripts related to stress responses and apoptosis enriched at the wound healing stage, various signaling pathways including Wnt and TGF-β enriched during early regrowth, and negative regulation of extracellular remodeling proteases as enriched in late stage regeneration. Additionally, comprehensive 3’-UTR binding site prediction and probability of conserved targeting for all C. robusta microRNAs (including those identified here) was calculated using TargetScanS. A microRNA-target network was subsequently constructed by calculating Pearson correlation coefficients for all predicted microRNA-mRNA target pairs expressed during regeneration. Network based clustering associated one or more microRNAs and their targets into 22 non-overlapping groups. Functional analysis of mRNA targets in each network cluster showed that enrichment of stress response, signaling pathway and extracellular remodeling categories associated with specific stages could also be associated with specific microRNAs. Finally, predicted targets of the miR-9 network cluster suggest a role in regulating differentiation and proliferative state of neural progenitors through regulation of the cytoskeleton and cell cycle. This work represents a significant advance in the prediction of microRNA effects on appendage regeneration and provides a foundation for investigating evolutionary conservation of microRNAs during regeneration in chordates.
Project description:Among the Chordates tunicates demonstrate the highest capacity for regeneration, ranging from full-body regeneration in colonial ascidians to appendage regeneration in solitary ascidians. Here we present a parallel study of mRNA and microRNA expression at three stages of oral siphon regeneration in the solitary ascidian Ciona robusta (a.k.a., Ciona intestinalis), and the derived network of their interactions. In the process of identifying 248 mRNAs and 15 microRNAs as differentially expressed (DE) across the course of regeneration, we also identified 57 novel microRNAs, several of which are among the most highly DE. Analysis functional categories identified transcripts related to stress responses and apoptosis enriched at the wound healing stage, various signaling pathways including Wnt and TGF-β enriched during early regrowth, and negative regulation of extracellular remodeling proteases as enriched in late stage regeneration. Additionally, comprehensive 3’-UTR binding site prediction and probability of conserved targeting for all C. robusta microRNAs (including those identified here) was calculated using TargetScanS. A microRNA-target network was subsequently constructed by calculating Pearson correlation coefficients for all predicted microRNA-mRNA target pairs expressed during regeneration. Network based clustering associated one or more microRNAs and their targets into 22 non-overlapping groups. Functional analysis of mRNA targets in each network cluster showed that enrichment of stress response, signaling pathway and extracellular remodeling categories associated with specific stages could also be associated with specific microRNAs. Finally, predicted targets of the miR-9 network cluster suggest a role in regulating differentiation and proliferative state of neural progenitors through regulation of the cytoskeleton and cell cycle. This work represents a significant advance in the prediction of microRNA effects on appendage regeneration and provides a foundation for investigating evolutionary conservation of microRNAs during regeneration in chordates.
Project description:Microarray analysis indicated many changes in gene expression, including genes related the the Notch signaling system, during oral siphon regeneration in adult Ciona. Subsequent qPCR gene expression and inhibitor of experiments confirm a role of the Notch system, probably in the formation of a regeneration blastema.
Project description:Microarray analysis indicated many changes in gene expression, including genes related the the Notch signaling system, during oral siphon regeneration in adult Ciona. Subsequent qPCR gene expression and inhibitor of experiments confirm a role of the Notch system, probably in the formation of a regeneration blastema. To identify the genes defferentially expressed in degenerating siphone, gene expressions in intact siphones and excised siphones (after 3 days, 6 days and 9 days) were examined.
Project description:Ciona intestinalis is an invertebrate animal model system that is well characterized and has many advantages for the study of cardiovascular biology. The regulatory mechanisms of cardiac myocyte proliferation in Ciona are intriguing since Ciona are capable of regeneration throughout their lifespan. To identify important regeneration factors in Ciona, microarray analysis was conducted on RNA from adult Ciona hearts with normal or damaged myocardium using custom Affymetrix GeneChips. After a 24- or 48-hour recovery period, total RNA was isolated from damaged and control hearts. Initial results indicate significant changes in gene expression in hearts damaged by ligation in comparison to cryoinjured or control hearts. Ligation injury shows differential expression of 223 genes as compared to control (fold change >2, p<0.01, Student’s t-test) with limited false discovery (5.8%). Among these 223 genes, 117 have known human orthologs of which 68 were up-regulated and 49 were down-regulated. Notably, FGF 9/16/20 and Ras were significantly upregulated in injured hearts. Histological analyses of injured myocardium were conducted in parallel to the microarray study. Taken together, these studies will coordinate differences in gene expression to cellular changes in the regenerative myocardium of Ciona, which will help to elucidate the regulatory mechanisms of cardiac myocyte proliferation across chordates.