Abstract
The epimorphic regeneration of zebrafish caudal fin is rapid and complete. We have analyzed the biomechanism of zebrafish caudal fin regeneration at various time points based on differential proteomics approaches. The spectrum of proteome changes caused by regeneration were analyzed among controls (0 h) and 1, 12, 24, 48, and 72 h postamputation involving quantitative differential proteomics analysis based on two-dimensional gel electrophoresis matrix-assisted laser desorption/ionization and differential in-gel electrophoresis Orbitrap analysis. A total of 96 proteins were found differentially regulated between the control nonregenerating and regenerating tissues of different time points for having at least 1.5-fold changes. 90 proteins were identified as differentially regulated for regeneration based on differential in-gel electrophoresis analysis between the control and regenerating tissues. 35 proteins were characterized for its expression in all of the five regenerating time points against the control samples. The proteins identified and associated with regeneration were found to be directly allied with various molecular, biological, and cellular functions. Based on network pathway analysis, the identified proteome data set for regeneration was majorly associated in maintaining cellular structure and architecture. Also the proteins were found associated for the cytoskeleton remodeling pathway and cellular immune defense mechanism. The major proteins that were found differentially regulated during zebrafish caudal fin regeneration includes keratin and its 10 isoforms, cofilin 2, annexin a1, skeletal α1 actin, and structural proteins. Annexin A1 was found to be exclusively undergoing phosphorylation during regeneration. The obtained differential proteome and the direct association of the various proteins might lead to a new understanding of the regeneration mechanism.
Highlights
Regeneration is an important mechanism found among most of the animals including humans in various tissues and organs towards growth, regrowth, repair, reproduction, and survival
Keratin and Regeneration—In this study we have identified an idiosyncratic association of keratin and its isoforms for regeneration
Proteomics-based analysis of regeneration has been extensively studied among amphibians for its blastema formation in regenerating axolotl limbs (15) and proteomic changes during onset of regeneration (16)
Summary
Animals and Regeneration Experiments—Wild zebrafish obtained from local farmers were housed and maintained under standard conditions in sterile water (5, 6). Differential proteomic analyses were performed between the regenerating samples and the nonregenerating control based on 2DE and DIGE analysis. MS AND MSMS Analysis—Protein spots having more than 1.5-fold change between the nonregenerating (zero hour sample) and regenerating tissues (all five time points) based on 2DE and DIGE gel analysis were selected for the MS and MSMS analyses. One- and Two-dimensional Western Blot Analysis—Proteins obtained from the control and regenerating tissues were analyzed for its expression level based on both one-dimensional gel electrophoresis and 2DE Western blot analysis. For one-dimensional gel electrophoresis Western blot analysis, 40 g of total proteins of each time point were electrophoresed on a 10% SDS-PAGE and transferred using wet transfer method. Real Time PCR Analysis—Total RNA was extracted from the control and regenerating caudal fin tissues using TRI reagent (Sigma) according to the manufacturer’s protocol. The differentially displayed proteins were analyzed for different processes, networks and pathway maps using the GeneGo software (www.genego.com) analysis
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
