| 1. |
- Leyhr, Jake, et al.
(författare)
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A novel cis-regulatory element drives early expression of Nkx3.2 in the gnathostome primary jaw joint
- 2022
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Ingår i: eLIFE. - : eLife Sciences Publications Ltd. - 2050-084X. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- The acquisition of movable jaws was a major event during vertebrate evolution. The role of NK3 homeobox 2 (Nkx3.2) transcription factor in patterning the primary jaw joint of gnathostomes (jawed vertebrates) is well known, however knowledge about its regulatory mechanism is lacking. In this study, we report a proximal enhancer element of Nkx3.2 that is deeply conserved in most gnathostomes but undetectable in the jawless hagfish and lamprey. This enhancer is active in the developing jaw joint region of the zebrafish Danio rerio, and was thus designated as jaw joint regulatory sequence 1 (JRS1). We further show that JRS1 enhancer sequences from a range of gnathostome species, including a chondrichthyan and mammals, have the same activity in the jaw joint as the native zebrafish enhancer, indicating a high degree of functional conservation despite the divergence of cartilaginous and bony fish lineages or the transition of the primary jaw joint into the middle ear of mammals. Finally, we show that deletion of JRS1 from the zebrafish genome using CRISPR/Cas9 results in a significant reduction of early gene expression of nkx3.2 and leads to a transient jaw joint deformation and partial fusion. Emergence of this Nkx3.2 enhancer in early gnathostomes may have contributed to the origin and shaping of the articulating surfaces of vertebrate jaws.
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| 2. |
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| 3. |
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| 4. |
- Waldmann, Laura, et al.
(författare)
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The Broad Role of Nkx3.2 in the Development of the Zebrafish Axial Skeleton
- 2024
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- The transcription factor Nkx3.2 (Bapx1) is an important chondrocyte maturation inhibitor. Previous Nkx3.2 knock-down and overexpression studies in non-mammalian gnathostomes have focused on its role in primary jaw joint development, while little is known about the function of this gene in broader skeletal development. We generated CRISPR/Cas9 knockout of nkx3.2 in zebrafish and applied a range of techniques to characterize skeletal phenotypes at developmental stages from larva to adult, revealing fusions in bones of the occiput, the loss or deformation of bony elements derived from basiventral cartilages of the vertebrae, and an increased length of the proximal radials of the dorsal and anal fins. These phenotypes are reminiscent of Nkx3.2 knockout phenotypes in mammals, suggesting that the function of this gene in axial skeletal development is ancestral to osteichthyans. Our results highlight the broad role of nkx3.2 in zebrafish skeletal development and its context-specific functions in different skeletal elements.
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| 5. |
- Waldmann, Laura, et al.
(författare)
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The broad role of Nkx3.2 in the development of the zebrafish axial skeleton
- 2021
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 16:8
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Tidskriftsartikel (refereegranskat)abstract
- The transcription factor Nkx3.2 (Bapx1) is an important chondrocyte maturation inhibitor. Previous Nkx3.2 knockdown and overexpression studies in non-mammalian gnathostomes have focused on its role in primary jaw joint development, while the function of this gene in broader skeletal development is not fully described. We generated a mutant allele of nkx3.2 in zebrafish with CRISPR/Cas9 and applied a range of techniques to characterize skeletal phenotypes at developmental stages from larva to adult, revealing loss of the jaw joint, fusions in bones of the occiput, morphological changes in the Weberian apparatus, and the loss or deformation of bony elements derived from basiventral cartilages of the vertebrae. Axial phenotypes are reminiscent of Nkx3.2 knockout in mammals, suggesting that the function of this gene in axial skeletal development is ancestral to osteichthyans. Our results highlight the broad role of nkx3.2 in zebrafish skeletal development and its context-specific functions in different skeletal elements.
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| 6. |
- Waldmann, Laura, et al.
(författare)
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The Role of Gdf5 in the Development of the Zebrafish Fin Endoskeleton
- 2022
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Ingår i: Developmental Dynamics. - : John Wiley & Sons. - 1058-8388 .- 1097-0177. ; 251:9, s. 1535-1549
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Tidskriftsartikel (refereegranskat)abstract
- The development of the vertebrate skeleton requires a complex interaction of multiple factors to facilitate correct shaping and positioning of bones and joints. Growth and differentiation factor 5 (Gdf5), a member of the transforming growth factor-beta family (TGF-beta) is involved in patterning appendicular skeletal elements including joints. Expression of gdf5 in zebrafish has been detected within the first pharyngeal arch jaw joint, fin mesenchyme condensations and segmentation zones in median fins, however little is known about the functional role of Gdf5 outside of Amniota. We generated CRISPR/Cas9 knockout of gdf5 in zebrafish and analysed the resulting phenotype at different developmental stages. Homozygous gdf5 mutant zebrafish display truncated median fin endoskeletal elements and loss of posterior radials in the pectoral fins. These findings are consistent with phenotypes observed in human and mouse appendicular skeleton in response to Gdf5 knockout, suggesting a broadly conserved role for Gdf5 in Osteichthyes.
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| 7. |
- Zhang, Hanqing, et al.
(författare)
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zOPT: an open source optical projection tomography system and methods for rapid 3D zebrafish imaging
- 2020
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Ingår i: Biomedical Optics Express. - : The Optical Society. - 2156-7085. ; 11:8, s. 4290-4305
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Tidskriftsartikel (refereegranskat)abstract
- Optical projection tomography (OPT) is a 3D imaging alternative to conventional microscopy which allows imaging of millimeter-sized object with isotropic micrometer resolution. The zebrafish is an established model organism and an important tool used in genetic and chemical screening. The size and optical transparency of the embryo and larva makes them well suited for imaging using OPT. Here, we present an open-source implementation of an OPT platform, built around a customized sample stage, 3D-printed parts and open source algorithms optimized for the system. We developed a versatile automated workflow including a two-step image processing approach for correcting the center of rotation and generating accurate 3D reconstructions. Our results demonstrate high-quality 3D reconstruction using synthetic data as well as real data of live and fixed zebrafish. The presented 3D-printable OPT platform represents a fully open design, low-cost and rapid loading and unloading of samples. Our system offers the opportunity for researchers with different backgrounds to setup and run OPT for large scale experiments, particularly in studies using zebrafish larvae as their key model organism.
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| 8. |
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| 9. |
- Debiais-Thibaud, Melanie, et al.
(författare)
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Skeletal Mineralization in Association with Type X Collagen Expression Is an Ancestral Feature for Jawed Vertebrates
- 2019
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Ingår i: Molecular biology and evolution. - : Oxford University Press (OUP). - 0737-4038 .- 1537-1719. ; 36:10, s. 2265-2276
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Tidskriftsartikel (refereegranskat)abstract
- In order to characterize the molecular bases of mineralizing cell evolution, we targeted type X collagen, a nonfibrillar network forming collagen encoded by the Col10a1 gene. It is involved in the process of endochondral ossification in ray-finned fishes and tetrapods (Osteichthyes), but until now unknown in cartilaginous fishes (Chondrichthyes). We show that holocephalans and elasmobranchs have respectively five and six tandemly duplicated Col10a1 gene copies that display conserved genomic synteny with osteichthyan Col10a1 genes. All Col10a1 genes in the catshark Scyliorhinus canicula are expressed in ameloblasts and/or odontoblasts of teeth and scales, during the stages of extracellular matrix protein secretion and mineralization. Only one duplicate is expressed in the endoskeletal (vertebral) mineralizing tissues. We also show that the expression of type X collagen is present in teeth of two osteichthyans, the zebrafish Danio rerio and the western clawed frog Xenopus tropicalis, indicating an ancestral jawed vertebrate involvement of type X collagen in odontode formation. Our findings push the origin of Col10a1 gene prior to the divergence of osteichthyans and chondrichthyans, and demonstrate its ancestral association with mineralization of both the odontode skeleton and the endoskeleton.
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| 10. |
- Haitina, Tatjana, Docent, 1981-, et al.
(författare)
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Evolutionary Genomics of Odontode Tissues
- 2023. - 1
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Ingår i: Odontodes. - Boca Raton : CRC Press. - 9781003439653 ; , s. 41-
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Bokkapitel (refereegranskat)abstract
- Teeth, scales, fin spines, and other odontodes are hot topics in evolutionary biology due to their great variation in number, shape, and anatomical localization in vertebrates as a result of different types of adaptations. Structural adaptation arose through variations in the composition of the enamel/enameloid and dentin layers that build up the mineralized components of teeth and other odontodes. These variations are known from both fossil and extant groups of vertebrates and rely on differences in the proteins secreted by the specialized odontode cells, ameloblasts, and odontoblasts.In this chapter, we focus on recent genomic and gene expression data to better understand the evolution of gene families engaged in the processes of secretion, mineralization, and maturation of odontode matrices. Our focus will be structural matrix proteins (collagenous and non-collagenous), calcium-interacting proteins necessary for mineralization, and proteins regulating matrix degradation over the course of tissue maturation.
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| 11. |
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| 12. |
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| 15. |
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| 16. |
- Jerve, Anna, et al.
(författare)
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Vascularization and odontode structure of a dorsal ridge spine of Romundina stellina Ørvig 1975
- 2017
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Ingår i: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 12:12
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Tidskriftsartikel (refereegranskat)abstract
- There are two types of dermal skeletons in jawed vertebrates: placoderms and osteichthyans carry large bony plates (macromery), whereas chondrichthyans and acanthodians are covered by small scales (micromery). Fin spines are one of the last large dermal structures found on micromeric taxa and offer a potential source of histology and morphology that can be compared to those found on macromeric groups. Dermal fin spines offer a variety of morphology but aspects of their growth modes and homology are unclear. Here, we provide detailed descriptions of the microstructure and growth of a dorsal ridge spine from the acanthothoracid placoderm, Romundina stellina, using virtual three-dimensional paleohistological datasets. From these data we identify several layers of dentine ornamentation covering the lateral surfaces of the spine and reconstructed their growth pattern. We show that this spine likely grew posteriorly and proximally from a narrow portion of bone located along the leading edge of the spine. The spine is similarly constructed to the scales with a few exceptions, including the absence of polarized fibers distributed throughout the bone and the presence of a thin layer of perichondral bone. The composition of the spine (semidentine odontodes, dermal bone, perichondral bone) is identical to that of the Romundina dermal plates. These results illustrate the similarities and differences between the dermal tissues in Romundina and indicate that the spine grew differently from the dentinous fin spines from extant and fossil chondrichthyans. The morphology and histology of Romundina is most similar to the fin spine of the probable stem osteichthyan Lophosteus, with a well-developed inner cellular bony base and star-shaped odontodes on the surface. Results from these studies will undoubtedly have impact on our understanding of fossil fin spine histology and evolution, contributing to the on-going revision of early gnathostome phylogeny.
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| 17. |
- Leurs, Nicolas, et al.
(författare)
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Evolution of Matrix Gla and Bone Gla Protein Genes in Jawed Vertebrates
- 2021
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Ingår i: Frontiers in Genetics. - : Frontiers Media S.A.. - 1664-8021. ; 12
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Tidskriftsartikel (refereegranskat)abstract
- Matrix Gla protein (Mgp) and bone Gla protein (Bgp) are vitamin-K dependent proteins that bind calcium in their gamma-carboxylated versions in mammals. They are recognized as positive (Bgp) or negative (Mgp and Bgp) regulators of biomineralization in a number of tissues, including skeletal tissues of bony vertebrates. The Mgp/Bgp gene family is poorly known in cartilaginous fishes, which precludes the understanding of the evolution of the biomineralization toolkit at the emergence of jawed vertebrates. Here we took advantage of recently released genomic and transcriptomic data in cartilaginous fishes and described the genomic loci and gene expression patterns of the Mgp/Bgp gene family. We identified three genes, Mgp1, Mgp2, and Bgp, in cartilaginous fishes instead of the single previously reported Mgp gene. We describe their genomic loci, resulting in a dynamic evolutionary scenario for this gene family including several events of local (tandem) duplications, but also of translocation events, along jawed vertebrate evolution. We describe the expression patterns of Mgp1, Mgp2, and Bgp in embryonic stages covering organogenesis in the small-spotted catshark Scyliorhinus canicula and present a comparative analysis with Mgp/Bgp family members previously described in bony vertebrates, highlighting ancestral features such as early embryonic, soft tissues, and neuronal expressions, but also derived features of cartilaginous fishes such as expression in fin supporting fibers. Our results support an ancestral function of Mgp in skeletal mineralization and a later derived function of Bgp in skeletal development that may be related to the divergence of bony vertebrates.
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| 18. |
- Leyhr, Jake, et al.
(författare)
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Enhanced contrast synchrotron X-ray microtomography for describing skeleton-associated soft tissue defects in zebrafish mutants
- 2023
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Ingår i: Frontiers in Endocrinology. - : Frontiers Media S.A.. - 1664-2392. ; 14
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Tidskriftsartikel (refereegranskat)abstract
- Detailed histological analyses are desirable for zebrafish mutants that are models for human skeletal diseases, but traditional histological techniques are limited to two-dimensional thin sections with orientations highly dependent on careful sample preparation. On the other hand, techniques that provide three-dimensional (3D) datasets including mu CT scanning are typically limited to visualizing the bony skeleton and lack histological resolution. We combined diffusible iodine-based contrast enhancement (DICE) and propagation phase-contrast synchrotron radiation micro-computed tomography (PPC-SR mu CT) to image late larval and juvenile zebrafish, obtaining high-quality 3D virtual histology datasets of the mineralized skeleton and surrounding soft tissues. To demonstrate this technique, we used virtual histological thin sections and 3D segmentation to qualitatively and quantitatively compare wild-type zebrafish and nkx3.2(-/-) mutants to characterize novel soft-tissue phenotypes in the muscles and tendons of the jaw and ligaments of the Weberian apparatus, as well as the sinus perilymphaticus associated with the inner ear. We could observe disrupted fiber organization and tendons of the adductor mandibulae and protractor hyoideus muscles associated with the jaws, and show that despite this, the overall muscle volumes appeared unaffected. Ligaments associated with the malformed Weberian ossicles were mostly absent in nkx3.2(-/-) mutants, and the sinus perilymphaticus was severely constricted or absent as a result of the fused exoccipital and basioccipital elements. These soft-tissue phenotypes have implications for the physiology of nkx3.2(-/-) zebrafish, and demonstrate the promise of DICE-PPC-SR mu CT for histopathological investigations of bone-associated soft tissues in small-fish skeletal disease models and developmental studies more broadly.
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| 19. |
- Medina Jimenez, Brenda Irene, 1986-
(författare)
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Single-cell RNA sequencing as a tool to study panarthropod evolution
- 2021
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Panarthropoda is a monophyletic group comprised of arthropods and lobopods, molting animals with a segmented body, paired appendages, dorsal brain, and ventral nerve cords. Evolutionary Developmental Biology (EvoDevo) is an interdisciplinary field that seeks to understand how changes in development form the basis for variations in morphology and phenotypic evolution, including the genetic network underlying these processes. To study the evolution of panarthropods from such an EvoDevo perspective, one typically uses standard molecular techniques. A first step here is to investigate the expression of a gene of interest in order to find out where and when it is transcribed during development. A hallmark of EvoDevo studies is its comparative character, often with respect to model organisms such as the fruit fly Drosophila melanogaster.Recently developed single-cell RNA sequencing technologies allow the profiling of a plethora of gene expression on the level of individual cells, and thus provide a much more detailed insight into gene expression.In Paper I, I applied standard molecular techniques used in EvoDevo research such as PCR, gene cloning, probe synthesis and whole mount in situ hybridization, to investigate the embryonic expression patterns of the tiptop/teashirt (tio/tsh) and spalt (sal) genes in a range of arthropods representing all main groups of this phylum, and an onychophoran. In the arthropod model Drosophila, these genes act as trunk-specifiers, and the objective of my work was to find out if this is conserved in Arthropoda or even Panarthropoda as a whole. I provide comprehensive data on arthropod tio/tsh and sal expression, including the first data from an onychophoran. The results support the idea that tio/tsh genes are involved in the development of ‘trunk’ segments by regulating limb development. In addition, my data suggest that the function of Sal is unlikely to be conserved in trunk vs head development. Early expression of sal, however, is in line with a potential homeotic function of this gene, at least in Arthropoda.In Paper II, I provide an embryonic tissue dissociation protocol for embryos of the common house spider Parasteatoda tepidariorum that I developed and that I successfully applied for single-cell RNA sequencing. In addition, I report on the progress of this experiment, and provide and discuss preliminary results.
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| 20. |
- Ocampo Daza, Daniel, 1984-, et al.
(författare)
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Reconstruction of the Carbohydrate 6-O Sulfotransferase Gene Family Evolution in Vertebrates Reveals Novel Member, CHST16, Lost in Amniotes
- 2020
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Ingår i: Genome Biology and Evolution. - : OXFORD UNIV PRESS. - 1759-6653. ; 12:7, s. 993-1012
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Tidskriftsartikel (refereegranskat)abstract
- Glycosaminoglycans are sulfated polysaccharide molecules, essential for many biological processes. The 6-O sulfation of glycosaminoglycans is carried out by carbohydrate 6-O sulfotransferases (C6OSTs), previously named Gal/GalNAc/GlcNAc 6-O sulfotransferases. Here, for the first time, we present a detailed phylogenetic reconstruction, analysis of gene synteny conservation and propose an evolutionary scenario for the C6OST family in major vertebrate groups, including mammals, birds, nonavian reptiles, amphibians, lobe-finned fishes, ray-finned fishes, cartilaginous fishes, and jawless vertebrates. The C6OST gene expansion likely started early in the chordate lineage, giving rise to four ancestral genes after the divergence of tunicates and before the emergence of extant vertebrates. The two rounds of whole-genome duplication in early vertebrate evolution (1R/2R) only contributed two additional C6OST subtype genes, increasing the vertebrate repertoire from four genes to six, divided into two branches. The first branch includes CHST1 and CHST3 as well as a previously unrecognized subtype, CHST16 that was lost in amniotes. The second branch includes CHST2, CHST7, and CHST5. Subsequently, local duplications of CHST5 gave rise to CHST4 in the ancestor of tetrapods, and to CHST6 in the ancestor of primates. The teleost-specific gene duplicates were identified for CHST1, CHST2, and CHST3 and are result of whole-genome duplication (3R) in the teleost lineage. We could also detect multiple, more recent lineage-specific duplicates. Thus, the vertebrate repertoire of C6OST genes has been shaped by gene duplications and gene losses at several stages of vertebrate evolution, with implications for the evolution of skeleton, nervous system, and cell-cell interactions.
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| 21. |
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| 22. |
- Stundl, Jan, et al.
(författare)
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Ancient vertebrate dermal armor evolved from trunk neural crest
- 2023
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Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences (PNAS). - 0027-8424 .- 1091-6490. ; 120:30
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Tidskriftsartikel (refereegranskat)abstract
- Bone is an evolutionary novelty of vertebrates, likely to have first emerged as part of ancestral dermal armor that consisted of osteogenic and odontogenic components. Whether these early vertebrate structures arose from mesoderm or neural crest cells has been a matter of considerable debate. To examine the developmental origin of the bony part of the dermal armor, we have performed in vivo lineage tracing in the sterlet sturgeon, a representative of nonteleost ray- finned fish that has retained an extensive postcranial dermal skeleton. The results definitively show that sterlet trunk neural crest cells give rise to osteoblasts of the scutes. Transcriptional profiling further reveals neural crest gene signature in sterlet scutes as well as bichir scales. Finally, histological and microCT analyses of ray- finned fish dermal armor show that their scales and scutes are formed by bone, dentin, and hypermineralized covering tissues, in various combinations, that resemble those of the first armored vertebrates. Taken together, our results support a primitive skeletogenic role for the neural crest along the entire body axis, that was later progressively restricted to the cranial region during vertebrate evolution. Thus, the neural crest was a crucial evolutionary innovation driving the origin and diversification of dermal armor along the entire body axis.
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| 23. |
- Tsakoumis, Emmanouil, MSc, 1990-
(författare)
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The role of leptin in zebrafish (Danio rerio) : Novel insights into appetite regulation and reproduction
- 2021
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The hormone leptin is a peripheral metabolic signal and an important regulator of energy balance. In mammals, leptin acts on the appetite centers in the hypothalamus, causing anorexigenic functions by inhibiting food intake. It is also considered as a link between the nutritional status and the endocrine reproductive axis. However, the actions of leptin in teleosts are not fully understood. This thesis investigated the possible role of leptin in the regulation of appetite and reproduction in teleosts, using a loss of function leptin receptor zebrafish strain (lepr sa12953).Under different feeding conditions (normal feeding, 7-day fasting, 2- and 6-hours post refeeding) the transcription of orexigenic and anorexigenic genes was influenced by leptin in the zebrafish brain. Leptin signaling inhibited the transcription of orexigenic genes, during short-term fasting and refeeding, and stimulated the transcription of anorexigenic genes under normal feeding in wild-types, indicating an anorexigenic role of leptin in appetite regulation in zebrafish. Moreover, a leptin-dependent gene regulatory network (GRN), involved in the behavioral and metabolic control of appetite was suggested in the brain, including the cart/crhb/gnrh2 genes and their respective co-expressed modules, mediated by the transcription factor sp3a.Furthermore, impaired leptin signaling resulted in severe reproductive deficiencies in female zebrafish. Folliculogenesis was not affected, but oocyte maturation and ovulation were disrupted in lepr mutant females, resulting in low number of ovulated eggs. Moreover, the transcripts of luteinizing hormone beta (lhb) in the pituitary were significantly lower in the mutant females. Analysis of candidate genes revealed differential expression of genes involved in steroidogenesis, oocyte maturation and ovulation in the ovaries of the lepr mutants. Transcriptomic analysis of isolated fully grown follicles linked the reproductive deficiencies to the suppression of essential metabolic pathways during oocyte maturation and ovulation in teleosts, such as estrogen regulation, ribosome biogenesis, mRNA translation and lipid metabolism.Overall, the results from the present thesis provided, for the first time in zebrafish, evidence that leptin is involved in appetite regulation, by mediating the transcription of appetite-regulating genes and a GRN in the brain, as well as that leptin consists a critical regulator of female reproduction, especially during oocyte maturation and ovulation.
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| 24. |
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