Regenerating zebrafish scales express a subset of evolutionary conserved genes involved in human skeletal disease

Abstract

Background Scales are mineralised exoskeletal structures that are part of the dermal skeleton. Scales have been mostly lost during evolution of terrestrial vertebrates whilst bony fish have retained a mineralised dermal skeleton in the form of fin rays and scales. Each scale is a mineralised collagen plate that is decorated with both matrix-building and resorbing cells. When removed, an ontogenetic scale is quickly replaced following differentiation of the scale pocket-lining cells that regenerate a scale. Processes promoting de novo matrix formation and mineralisation initiated during scale regeneration are poorly understood. Therefore, we performed transcriptomic analysis to determine gene networks and their pathways involved in dermal scale regeneration. Results We defined the transcriptomic profiles of ontogenetic and regenerating scales of zebrafish and identified 604 differentially expressed genes (DEGs). These were enriched for extracellular matrix, ossification, and cell adhesion pathways, but not in enamel or dentin formation processes indicating that scales are reminiscent to bone. Hypergeometric tests involving monogenetic skeletal disorders showed that DEGs were strongly enriched for human orthologues that are mutated in low bone mass and abnormal bone mineralisation diseases (P< 2× 10−3). The DEGs were also enriched for human orthologues associated with polygenetic skeletal traits, including height (P< 6× 10−4), and estimated bone mineral density (eBMD, P< 2× 10−5). Zebrafish mutants of two human orthologues that were robustly associated with height (COL11A2, P=6× 10−24) or eBMD (SPP1, P=6× 10−20) showed both exo- and endo- skeletal abnormalities as predicted by our genetic association analyses; col11a2Y228X/Y228X mutants showed exoskeletal and endoskeletal features consistent with abnormal growth, whereas spp1P160X/P160X mutants predominantly showed mineralisation defects. Conclusion We show that scales have a strong osteogenic expression profile comparable to other elements of the dermal skeleton, enriched in genes that favour collagen matrix growth. Despite the many differences between scale and endoskeletal developmental processes, we also show that zebrafish scales express an evolutionarily conserved sub-population of genes that are relevant to human skeletal disease.

Precise targeting for 3D cryo-correlative light and electron microscopy volume imaging of tissues using a FinderTOP

“…Zebrafish and scale harvesting Zebrafish were maintained under normal husbandry conditions as described previously 37 . Transgenic line Tg(Ola.sp7:NLS-GFP) were anaesthetized using 0.1% (v/v) 2-phenoxyethanol and placed on a wet tissue containing system water and anesthetic.…”

Zebrafish models for glucocorticoid-induced osteoporosis

“…The episquamal side (external layer) is mineralized, containing a network of collagen fibrils[ 47 ]. Scleroblasts, the scale-forming osteoblasts derived from mesenchymal cells, are highly functionally similar to mammalian osteoblasts and express canonical osteoblastic markers[ 33 48 49 ]. The scale also contains osteoclasts required to maintain the homeostasis of scale growth[ 39 ].…”

High Bone Mass Disorders: New Insights From Connecting the Clinic and the Bench

“…(126) To date, there is a vast library of transgenic reporter and mutant zebrafish available that has been shown to accurately model various skeletal diseases (Zebrafish Information Network [ZFIN]; www.zfin.org), allowing bone cell populations to be imaged, FACS isolated, or manipulated. (124,127,128) Zebrafish can also model high BMD (84,(129)(130)(131) ; eg, an osteopetrosis-like phenotype in mmp9 À/À ;mmp13b À/À double mutant fish, (78) CSF1R-related dysosteosclerosis (OMIM 618476), (132,133) and PTDSS1-related Lenz-Majewski hyperostotic dysplasia (OMIM 151050). (134) Exploiting the zebrafish lifespan to understand spatiotemporal and molecular causes of HBM Initial zebrafish development is rapid with the first skeletal progenitor cells in the form of neural crest cells appear around the first day of development.…”

“…A recent study using bulk RNAseq on regenerating scales showed an enrichment of differentially expressed genes linked to ossification (GO:0001503), hedgehog/smoothened signaling pathway (GO:0007224), insulin-like growth factor signaling (GO:0048009), and cell adhesion (GO:0007155). (84) Moreover, many genes involved in a regenerating scale were enriched for human orthologues that cause monogenic skeletal diseases (eg, COL1A1-, SP7-, ANO5-related osteogenesis imperfecta) and/or are in loci associated with polygenic bone traits (eg BMD, height). (84) Shortening the Diagnostic Timeframe for HBM Disorders in the Future…”

“…(84) Moreover, many genes involved in a regenerating scale were enriched for human orthologues that cause monogenic skeletal diseases (eg, COL1A1-, SP7-, ANO5-related osteogenesis imperfecta) and/or are in loci associated with polygenic bone traits (eg BMD, height). (84) Shortening the Diagnostic Timeframe for HBM Disorders in the Future…”