Scale Morphogenesis Research Articles

Transcriptomic profiling of tissue environments critical for post-embryonic patterning and morphogenesis of zebrafish skin.

Pigment patterns and skin appendages are prominent features of vertebrate skin. In zebrafish, regularly patterned pigment stripes and an array of calcified scales form simultaneously in the skin during post-embryonic development. Understanding the mechanisms that regulate stripe patterning and scale morphogenesis may lead to the discovery of fundamental mechanisms that govern the development of animal form. To learn about cell types and signaling interactions that govern skin patterning and morphogenesis, we generated and analyzed single-cell transcriptomes of skin from wild-type fish as well as fish having genetic or transgenically induced defects in squamation or pigmentation. These data reveal a previously undescribed population of epidermal cells that express transcripts encoding enamel matrix proteins, suggest hormonal control of epithelial-mesenchymal signaling, clarify the signaling network that governs scale papillae development, and identify a critical role for the hypodermis in supporting pigment cell development. Additionally, these comprehensive single-cell transcriptomic data representing skin phenotypes of biomedical relevance should provide a useful resource for accelerating the discovery of mechanisms that govern skin development and homeostasis.

Transcriptomic profiling of tissue environments critical for post-embryonic patterning and morphogenesis of zebrafish skin

Pigment patterns and skin appendages are prominent features of vertebrate skin. In zebrafish, regularly patterned pigment stripes and an array of calcified scales form simultaneously in the skin during post-embryonic development. Understanding the mechanisms that regulate stripe patterning and scale morphogenesis may lead to the discovery of fundamental mechanisms that govern the development of animal form. To learn about cell types and signaling interactions that govern skin patterning and morphogenesis, we generated and analyzed single-cell transcriptomes of skin from wild-type fish as well as fish having genetic or transgenically induced defects in squamation or pigmentation. These data reveal a previously undescribed population of epidermal cells that express transcripts encoding enamel matrix proteins, suggest hormonal control of epithelial–mesenchymal signaling, clarify the signaling network that governs scale papillae development, and identify a critical role for the hypodermis in supporting pigment cell development. Additionally, these comprehensive single-cell transcriptomic data representing skin phenotypes of biomedical relevance should provide a useful resource for accelerating the discovery of mechanisms that govern skin development and homeostasis.

Identification of genes involved in regulating the development of feathered feet in chicken embryo

The genetic and developmental factors driving the diverse distribution and morphogenesis of feathers and scales on bird feet are yet unclear. Within a single species, Guangxi domestic chickens exhibit dramatic variety in feathered feet, making them an accessible model for research into the molecular basis of variations in skin appendages. In this study, we used H&E staining to observe the morphogenesis of feathered feet, scaled feet and wings skin at different embryonic stages in Longsheng-Feng chickens and Guangxi Partridge chickens. We selected 4 periods (E6, E7, E8, and E12) that play an important role in feather development and performed transcriptome sequencing to screen for candidate genes associated with feathered feet. Through comparison and analysis of transcriptome data, we identified a set of differently expressed genes (DGEs), which were enriched in appendage organ development, hindlimb morphogenesis, activation of transcription factor binding, and binding of sequence-specific DNA in the cis-regulatory region. In addition, we identified some feathered feet-related genes by analyzing the classical signaling pathways that regulate feather development. Finally, we identified candidate genes that regulate feathered feet formation, which include TBX5, PITX1, ZIC1, FGF20, WNT11, WNT7A, WNT16, and SHH. Interestingly, we found that TBX5 was significantly overexpressed in the skin of the feathered feet and had the highest expression at E7 (P < 0.01), whereas PITX1 expression was significantly reduced at E7(P < 0.01). It is hypothesized that TBX5 and PITX1 regulate the development of hair follicles through the Wnt/β-catenin signaling pathway at E7. Our results provide a theoretical basis for investigating the molecular regulatory mechanisms underlying the formation of chicken feathered feet.

Lateral adherens junctions mediate a supracellular actomyosin cortex in drosophila trachea

Drosophila trachea is a classical model for analyzing epithelial, especially tubular epithelial biology. We identify lateral E-cadherin mediated junctions that encircle the cells just basal to the zonula adherens in the larval trachea. The lateral junction is associated with downstream adapters, including catenins, and has a distinct junctional actin cortex. The lateral cortex contributes to the development of a supracellular actomyosin mesh in the late larvae. Establishment of this cytoskeletal structure depends on lateral junction associated Rho1 and Cdc42 GTPases, and Arp and WASP pathways. The supracellular network takes the character of stress fibers along the AP axis in the early hours of pupation. It contributes to the shortening of the epithelial tube albeit in a manner redundant to ECM-mediated compression mechanism. In conclusion, we show the invivo existence of functional lateral adherens junction and suggest a role for it in mediating dynamic cytoskeletal events during tissue scale morphogenesis.

Revealing the impacts on shaping scutate scales in goose skin

Feather follicles and scales are two types of skin appendages distributed on different parts of avian skin. The morphogenesis and development of scales in waterfowl remain largely unknown. Here, we used H&E staining, ISH and RNA sequencing to reveal the morphological and molecular variations at the early development of scutate scales in goose shank skin. Transcriptome analysis produced 1824 differentially expressed genes (DEGs) regulating the induction of scales and further enriched gene function in cell adhesion and Wnt signaling pathway, etc. A total of 8 candidate genes (ALDOC, CSRP2, KRT15, KRT75, LGALS1, S100A6, OGN and SFRP2) were further detected by RT-qPCR to show upregulated (6 genes) and downregulated (2 genes) from pre-placodal to placode stage during the induction of goose scales. The localization of 7 candidate genes (ALDOC, CSRP2, CD109, KRT15, KRT75, S100A6, and OGN) by ISH suggests the potential roles for dermal and epidermal development during the induction of scutate scales. The dynamic molecular changes and specific gene expression patterns revealed in this report provide general knowledge of scale development in waterfowl as well as skin appendage diversity.

Expression variations in ectodysplasin-A gene (eda) may contribute to morphological divergence of scales in haplochromine cichlids

BackgroundElasmoid scales are one of the most common dermal appendages and can be found in almost all species of bony fish differing greatly in their shape. Whilst the genetic underpinnings behind elasmoid scale development have been investigated, not much is known about the mechanisms involved in moulding of scales. To investigate the links between gene expression differences and morphological divergence, we inferred shape variation of scales from two different areas of the body (anterior and posterior) stemming from ten haplochromine cichlid species from different origins (Lake Tanganyika, Lake Malawi, Lake Victoria and riverine). Additionally, we investigated transcriptional differences of a set of genes known to be involved in scale development and morphogenesis in fish.ResultsWe found that scales from the anterior and posterior part of the body strongly differ in their overall shape, and a separate look on scales from each body part revealed similar trajectories of shape differences considering the lake origin of single investigated species. Above all, nine as well as 11 out of 16 target genes showed expression differences between the lakes for the anterior and posterior dataset, respectively. Whereas in posterior scales four genes (dlx5, eda, rankl and shh) revealed significant correlations between expression and morphological differentiation, in anterior scales only one gene (eda) showed such a correlation. Furthermore, eda displayed the most significant expression difference between species of Lake Tanganyika and species of the other two younger lakes. Finally, we found genetic differences in downstream regions of eda gene (e.g., in the eda-tnfsf13b inter-genic region) that are associated with observed expression differences. This is reminiscent of a genetic difference in the eda-tnfsf13b inter-genic region which leads to gain or loss of armour plates in stickleback.ConclusionThese findings provide evidence for cross-species transcriptional differences of an important morphogenetic factor, eda, which is involved in formation of ectodermal appendages. These expression differences appeared to be associated with morphological differences observed in the scales of haplochromine cichlids indicating potential role of eda mediated signal in divergent scale morphogenesis in fish.

Cell adhesion and junctional proteins in the developing skin of snakes indicate they coordinate the differentiation of the epidermis.

The development of scales and the sequence of epidermal layers during snake embryogenesis has been studied by immunofluorescence for the localization of cell adhesion, adherens, and communicating cell junctional proteins. At about 2nd/3rd of embryonic development in snakes the epidermis forms symmetric bumps at the beginning of scale formation, and they rapidly become asymmetric and elongate forming outer and inner surfaces of the very overlapped scales seen at hatching. The dermis separates a superficial loose from a deeper dense part; the latter is joined to segmental muscles and nerves, likely acting on scale orientation during snake movements. N-cam is present in the differentiating epidermis and mesenchyme of forming scales while L-cam is only/mainly detected in the periderm and epidermis. Mesenchymal N-cam is associated with the epidermis of the elongating dorsal scale surface and with the beta-differentiation that occurs in the overlapping outer surface of scales. Beta-catenin and Connexin-43 show a similar distribution, and they are mainly present in the periderm and differentiating suprabasal keratinocytes likely forming an intense connectivity during epidermal differentiation. Beta-catenin also shows nuclear localization in differentiating cells of the shedding and beta-layers at late stages of scale morphogenesis, before hatching. The study suggests that intensification of adhesion and gap junctions allows synchronization of the differentiation of suprabasal cells to produce the ordered sequence of epidermal layers of snake scales, starting from the shedding complex and the beta-layer.

Modeling scale morphogenesis in a Devonian chondrichthyan and scale growth patterns in crown gnathostomes

ABSTRACT Early chondrichthyans (cartilaginous fishes) possess a rich record of scale fossils that have impacted on taxonomic and phylogenetic studies of early jawed vertebrates, but the significance of these scales remains elusive due to the lack of well-defined characters. Gualepis, a scale-based genus from the Lower Devonian of Yunnan, China, was initially recognized as an early chondrichthyan of uncertain affinity. We investigate new material of Gualepis elegans from the type locality. By combining microtomographic data with morphological and histological examination of specimens at successive growth stages, we have reconstructed the 3D growth pattern of the scales. During growth, Gualepis scales added new odontodes appositionally, which has previously been documented in the dermal skeletons of Seretolepis, Kathemacanthus, Parexus, and other crownward members of the stem chondrichthyans. Phylogenetic analysis resolves Gualepis as a non-acanthodid stem chondrichthyan near the crown chondrichthyan node. Our 3D approach is potentially useful to render more reliable scale-based characters, especially regarding scale growth patterns. This study provides the first 3D virtual growth model of stem chondrichthyan scales with a Seretolepis-type morphogenesis, offering new insights into the early evolution of chondrichthyan dermal skeletons.

Transcriptome profiling towards understanding of the morphogenesis in the scale development of blunt snout bream (Megalobrama amblycephala)

Skin appendages in vertebrates have individual morphological differences, but share the same evolutionary origin. In this study, we used Megalobrama amblycephala as a fish model to study the developmental regulation mechanism of a common skin appendage in fish: scales. By combining in-toto live imaging method and transcriptomic analysis during the scale development, we elucidated core features of scale patterning containing three distinct regions and experiencing four stages. Differentially expressed genes in skin tissues at the initial site before and after scale development were analyzed and some key regulatory genes (Wnt3, Wnt6, Fgf8, Fgf10, Fgf16, Fgfr1a, Ihhb and BMP6) which are crucial for scale morphogenesis were selected. This study provides a strong reference for further exploration of the function of genes related to the molecular regulation mechanism of scale development in M. amblycephala, as well as in other fishes.

Actin bundles play a different role in shaping scales compared to bristles in the mosquito Aedes aegypti

Insect epithelial cells contain cellular extensions such as bristles, hairs, and scales. These cellular extensions are homologous structures that differ in morphology and function. They contain actin bundles that dictate their cellular morphology. While the organization, function, and identity of the major actin-bundling proteins in bristles and hairs are known, this information on scales is unknown. In this study, we characterized the development of scales and the role of actin bundles in the mosquito, Aedes aegypti. We show that scales undergo drastic morphological changes during development, from a cylindrical to flat shape with longer membrane invagination. Scale actin-bundle distribution changes from the symmetrical organization of actin bundles located throughout the bristle membrane to an asymmetrical organization. By chemically inhibiting actin polymerization and by knocking out the forked gene in the mosquito (Ae-Forked; a known actin-bundling protein) by CRISPR-Cas9 gene editing, we showed that actin bundles are required for shaping bristle, hair, and scale morphology. We demonstrated that actin bundles and Ae-Forked are required for bristle elongation, but not for that of scales. In scales, actin bundles are required for width formation. In summary, our results reveal, for the first time, the developmental process of mosquito scale formation and also the role of actin bundles and actin-bundle proteins in scale morphogenesis. Moreover, our results reveal that although scale and bristle are thought to be homologous structures, actin bundles have a differential requirement in shaping mosquito scales compared to bristles.

Regulation of Scale Morphogenesis through Wnt/PCP Signaling in Zebrafish

Regulation of Scale Morphogenesis through Wnt/PCP Signaling in Zebrafish

Mechanisms Governing Protein Clustering and Shape Changes in Biological Membranes

A characteristic feature of eukaryotic cells is the variety of membrane bound organelles, distinguished by their unique morphology and chemical composition. Despite the differences in membrane composition across organelles, ramified, tubular or sheet-like shapes are generic large scale morphologies observed in internal membranes, which suggest involvement of common underlying principles. While there is detailed knowledge of the molecular processes involved in membrane remodelling at short scales, our understanding of the underlying physical principles governing large scale morphogenesis is still rudimentary. One common aspect is that membranes are subject to the action of curvature sensing and curvature generating proteins which modulate local membrane shape and lipid composition. Even at relatively small values of surface coverage these proteins can cluster through to a variety or processes and trigger morphological changes which are important for singling. Another important feature, specific to organelle membranes, especially those in the trafficking pathways, is that they are subject to and driven by a continuous flux of membrane bound materials, on time scales comparable to membrane relaxation times. Here we present Monte Carlo Simulation models that tests the effect of the above processes on the morphology of the membranes. We find that the steady state shapes obtained as a result of such active processes, bear a striking resemblance to the ramified morphologies of organelles in vivo, pointing to the relevance of nonequilibrium fission-fusion in organelle morphogenesis. We show that, due to membrane curvature and composition interactions, the curvature-inducing membrane-nematogens can aggregate spontaneously, even at low concentrations, leading to a variety of membrane morphologies such as tubular and the sheet conformations. Strong lipid-protein interaction can result in fast protein clustering indicating a route to a lipid mediated signal amplification.

A living mesoscopic cellular automaton made of skin scales.

In vertebrates, skin colour patterns emerge from nonlinear dynamical microscopic systems of cell interactions. Here we show that in ocellated lizards a quasi-hexagonal lattice of skin scales, rather than individual chromatophore cells, establishes a green and black labyrinthine pattern of skin colour. We analysed time series of lizard scale colour dynamics over four years of their development and demonstrate that this pattern is produced by a cellular automaton (a grid of elements whose states are iterated according to a set of rules based on the states of neighbouring elements) that dynamically computes the colour states of individual mesoscopic skin scales to produce the corresponding macroscopic colour pattern. Using numerical simulations and mathematical derivation, we identify how a discrete von Neumann cellular automaton emerges from a continuous Turing reaction-diffusion system. Skin thickness variation generated by three-dimensional morphogenesis of skin scales causes the underlying reaction-diffusion dynamics to separate into microscopic and mesoscopic spatial scales, the latter generating a cellular automaton. Our study indicates that cellular automata are not merely abstract computational systems, but can directly correspond to processes generated by biological evolution.

Quantifying the relationship between cell division angle and morphogenesis through computational modeling

When biological cells divide, they divide on a given angle. It has been shown experimentally that the orientation of cell division angle for a single cell can be described by a probability density function. However, the way in which the probability density function underlying cell division orientation influences population or tissue scale morphogenesis is unknown. Here we show that a computational approach, with thousands of stochastic simulations modeling growth and division of a population of cells, can be used to investigate this unknown. In this paper we examine two potential forms of the probability density function: a wrapped normal distribution and a binomial distribution. Our results demonstrate that for the wrapped normal distribution the standard deviation of the division angle, potentially interpreted as biological noise, controls the degree of tissue scale anisotropy. For the binomial distribution, we demonstrate a mechanism by which direction and degree of tissue scale anisotropy can be tuned via the probability of each division angle. We anticipate that the method presented in this paper and the results of these simulations will be a starting point for further investigation of this topic.

Expressed miRNAs target feather related mRNAs involved in cell signaling, cell adhesion and structure during chicken epidermal development

MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression at the post-transcriptional level. Previous studies have shown that miRNA regulation contributes to a diverse set of processes including cellular differentiation and morphogenesis which leads to the creation of different cell types in multicellular organisms and is thus key to animal development. Feathers are one of the most distinctive features of extant birds and are important for multiple functions including flight, thermal regulation, and sexual selection. However, the role of miRNAs in feather development has been woefully understudied despite the identification of cell signaling pathways, cell adhesion molecules and structural genes involved in feather development. In this study, we performed a microarray experiment comparing the expression of miRNAs and mRNAs among three embryonic stages of development and two tissues (scutate scale and feather) of the chicken. We combined this expression data with miRNA target prediction tools and a curated list of feather related genes to produce a set of 19 miRNA-mRNA duplexes. These targeted mRNAs have been previously identified as important cell signaling and cell adhesion genes as well as structural genes involved in feather and scale morphogenesis. Interestingly, the miRNA target site of the cell signaling pathway gene, Aldehyde Dehydrogenase 1 Family, Member A3 (ALDH1A3), is unique to birds indicating a novel role in Aves. The identified miRNA target site of the cell adhesion gene, Tenascin C (TNC), is only found in specific chicken TNC splice variants that are differentially expressed in developing scutate scale and feather tissue indicating an important role of miRNA regulation in epidermal differentiation. Additionally, we found that β-keratins, a major structural component of avian and reptilian epidermal appendages, are targeted by multiple miRNA genes. In conclusion, our work provides quantitative expression data on miRNAs and mRNAs during feather and scale development and has produced a highly diverse, but manageable list of miRNA-mRNA duplexes for future validation experiments.

The anatomical placode in reptile scale morphogenesis indicates shared ancestry among skin appendages in amniotes.

Most mammals, birds, and reptiles are readily recognized by their hairs, feathers, and scales, respectively. However, the lack of fossil intermediate forms between scales and hairs and substantial differences in their morphogenesis and protein composition have fueled the controversy pertaining to their potential common ancestry for decades. Central to this debate is the apparent lack of an "anatomical placode" (that is, a local epidermal thickening characteristic of feathers' and hairs' early morphogenesis) in reptile scale development. Hence, scenarios have been proposed for the independent development of the anatomical placode in birds and mammals and parallel co-option of similar signaling pathways for their morphogenesis. Using histological and molecular techniques on developmental series of crocodiles and snakes, as well as of unique wild-type and EDA (ectodysplasin A)-deficient scaleless mutant lizards, we show for the first time that reptiles, including crocodiles and squamates, develop all the characteristics of an anatomical placode: columnar cells with reduced proliferation rate, as well as canonical spatial expression of placode and underlying dermal molecular markers. These results reveal a new evolutionary scenario where hairs, feathers, and scales of extant species are homologous structures inherited, with modification, from their shared reptilian ancestor's skin appendages already characterized by an anatomical placode and associated signaling molecules.

Tooth and scale morphogenesis in shark: an alternative process to the mammalian enamel knot system.

BackgroundThe gene regulatory network involved in tooth morphogenesis has been extremely well described in mammals and its modeling has allowed predictions of variations in regulatory pathway that may have led to evolution of tooth shapes. However, very little is known outside of mammals to understand how this regulatory framework may also account for tooth shape evolution at the level of gnathostomes. In this work, we describe expression patterns and proliferation/apoptosis assays to uncover homologous regulatory pathways in the catshark Scyliorhinus canicula.ResultsBecause of their similar structural and developmental features, gene expression patterns were described over the four developmental stages of both tooth and scale buds in the catshark. These gene expression patterns differ from mouse tooth development, and discrepancies are also observed between tooth and scale development within the catshark. However, a similar nested expression of Shh and Fgf suggests similar signaling involved in morphogenesis of all structures, although apoptosis assays do not support a strictly equivalent enamel knot system in sharks. Similarities in the topology of gene expression pattern, including Bmp signaling pathway, suggest that mouse molar development is more similar to scale bud development in the catshark.ConclusionsThese results support the fact that no enamel knot, as described in mammalian teeth, can be described in the morphogenesis of shark teeth or scales. However, homologous signaling pathways are involved in growth and morphogenesis with variations in their respective expression patterns. We speculate that variations in this topology of expression are also a substrate for tooth shape evolution, notably in regulating the growth axis and symmetry of the developing structure.Electronic supplementary materialThe online version of this article (doi:10.1186/s12862-015-0557-0) contains supplementary material, which is available to authorized users.

Genomic and expression analysis of a solute carrier protein (CcSLC25a5) gene from Cyprinus carpio Linnaeus

Using the Genefishing method, we identified seven potential regulatory genes involved in the process of scale morphogenesis in fishes. We further characterized a novel solute carrier protein gene (CcSLC), from the common carp which is differentially expressed in mirror carp and Jianli. The ORF encodes a peptide of 298 amino acids with a molecular mass of 31.5 kDa and a theoretical isoelectric point of 7.49. ScanProsite analysis indicated that it is a putative solute carrier protein that contains a substrate binding site. CcSLC was detected in carp embryos by in situ hybridization in the 70%-epiboly, 6-somite, and 14-somite embryonic stages. Gene expression stopped at the long pec stage. However, CcSLC25a5 was re-expressed during the initiation of scale formation in the regions that were scale covered. These findings provide novel insights into the features of early carp embryo and scale development.

Ultrastructure and morphogenesis of the wing scales in Heliconius erato phyllis (Lepidoptera: Nymphalidae): What silvery/brownish surfaces can tell us about the development of color patterning?

Usually the literature on Heliconius show three types of scales, classified based on the correlation between color and ultrastructure: type I – white and yellow, type II – black, and type III – orange and red. The ultrastructure of the scales located at the silvery/brownish surfaces of males/females is for the first time described in this paper. Besides, we describe the ontogeny of pigmentation, the scale morphogenesis and the maturation timing of scales fated to different colors in Heliconius erato phyllis. The silvery/brownish surfaces showed ultrastructurally similar scales to the type I, II and III. The ontogeny of pigmentation follows the sequence red, black, silvery/brownish and yellow. The maturation of yellow-fated scales, however, occurred simultaneously with the red-fated scales, before the pigmentation becomes visible. In spite of the scales at the silvery/brownish surfaces being ultrastructurally similar to the yellow, red and black scales, they mature after them; this suggests that the maturation timing does not show a relationship with the scale ultrastructure, with the deposition timing of the yellow pigment. The analysis of H. erato phyllis scale morphogenesis, as well as the scales ultrastructure and maturation timing, provided new findings into the developmental architecture of color pattern in Heliconius.

Crocodylians evolved scattered multi-sensory micro-organs.

BackgroundDuring their evolution towards a complete life cycle on land, stem reptiles developed both an impermeable multi-layered keratinized epidermis and skin appendages (scales) providing mechanical, thermal, and chemical protection. Previous studies have demonstrated that, despite the presence of a particularly armored skin, crocodylians have exquisite mechanosensory abilities thanks to the presence of small integumentary sensory organs (ISOs) distributed on postcranial and/or cranial scales.ResultsHere, we analyze and compare the structure, innervation, embryonic morphogenesis and sensory functions of postcranial, cranial, and lingual sensory organs of the Nile crocodile (Crocodylus niloticus) and the spectacled caiman (Caiman crocodilus). Our molecular analyses indicate that sensory neurons of crocodylian ISOs express a large repertoire of transduction channels involved in mechano-, thermo-, and chemosensory functions, and our electrophysiological analyses confirm that each ISO exhibits a combined sensitivity to mechanical, thermal and pH stimuli (but not hyper-osmotic salinity), making them remarkable multi-sensorial micro-organs with no equivalent in the sensory systems of other vertebrate lineages. We also show that ISOs all exhibit similar morphologies and modes of development, despite forming at different stages of scale morphogenesis across the body.ConclusionsThe ancestral vertebrate diffused sensory system of the skin was transformed in the crocodylian lineages into an array of discrete multi-sensory micro-organs innervated by multiple pools of sensory neurons. This discretization of skin sensory expression sites is unique among vertebrates and allowed crocodylians to develop a highly-armored, but very sensitive, skin.