Keratin Intermediate Filaments Research Articles

Keratin 8/18a.1 Expression Influences Embryonic Neural Crest Cell Dynamics and Contributes to Postnatal Corneal Regeneration in Zebrafish.

A complete understanding of neural crest cell mechanodynamics during ocular development will provide insight into postnatal neural crest cell contributions to ophthalmic abnormalities in adult tissues and inform regenerative strategies toward injury repair. Herein, single-cell RNA sequencing in zebrafish during early eye development revealed keratin intermediate filament genes krt8 and krt18a.1 as additional factors expressed during anterior segment development. In situ hybridization and immunofluorescence microscopy confirmed krt8 and krt18a.1 expression in the early neural plate border and migrating cranial neural crest cells. Morpholino oligonucleotide (MO)-mediated knockdown of K8 and K18a.1 markedly disrupted the migration of neural crest cell subpopulations and decreased neural crest cell marker gene expression in the craniofacial region and eye at 48 h postfertilization (hpf), resulting in severe phenotypic defects reminiscent of neurocristopathies. Interestingly, the expression of K18a.1, but not K8, is regulated by retinoic acid (RA) during early-stage development. Further, both keratin proteins were detected during postnatal corneal regeneration in adult zebrafish. Altogether, we demonstrated that both K8 and K18a.1 contribute to the early development and postnatal repair of neural crest cell-derived ocular tissues.

Changes of epidermal proteins immunolocalization in the corneous layer from embryonic to definitive avian beak

The process of keratinization and cornification in the developing beak has been studied through immunofluorescence and immunogold electron microscopy in chick and zebrafinch embryos. After the curved beak anlagen appears at the tip of the maxillar bone, 5–8 layers of embryonic epidermis are generated from the basal layer of the epidermis. These cells are weakly immunoabeled for IFKs (Intermediate Filament Keratins) and more intensely for scaffoldin, a protein of the EDC (Epidermal Differentiation Complex) involved in the soft keratinization of the embryonic epidermis. Immunolabeling for CBPs (Corneous Beta Proteins) is visible in the transitional embryonic layers that are temporarily generated between the embryonic and definitive beak epidermis. The electron microscope reveals that intermediate layers contain immunolabeled periderm granules for scaffoldin mixed with bundles of corneous material immunolabeled for CBPs. Intense CBPs labeling occurs in the compacting corneous bundles of beta-keratinocytes in the definitive beak while scaffolding labeling disappears. The embryonic epidermis is sloughed before hatching. Sox (Sulfhydryl Oxidase) immunolabeling reveals that the enzyme is almost absent in embryonic layers but is present in transitional and definitive beta-keratinocytes. This indicates the formation of cross-linked disulfide bonds in the definitive corneous layer of the beak. Some calcium precipitation, suggested from von Kossa staining, occurs in the corneous layers only on the 18th day of development in the chick, in preparation for hatching.

Variation in the Exon 3-4 Region of Ovine KRT85 and Its Effect on Wool Traits.

α-keratins are structural proteins in the cortex of wool fibres and assemble in an organized fashion into keratin intermediate filaments. Variation in these keratin proteins affects the structure and characteristics of wool fibre, making keratin genes ideal candidates for the development of gene markers that describe variations in wool traits. A region of KRT85 spanning exon 3-4 (including the entire exon 3, intron 3, exon 4 and part of intron 4) was investigated. Two banding patterns defining two variant sequences (A and B) were observed in this region, and these were characterised by the presence of two single nucleotide polymorphisms. The effect of this variation in the exon 3-4 region of KRT85 on wool traits was investigated in 463 Merino × Southdown-cross lambs. The frequencies of these two variants in these sheep were 55.6% and 44.4%, respectively. Three different genotypes were observed with frequencies of 32.6%, 46.1% and 21.3% for AA, AB and BB, respectively. The presence of A was associated with an increase in greasy fleece weight and clean fleece weight, while the presence of B was associated with an increased wool prickle factor. These findings should be replicated in a broader range of sheep breeds to determine whether the associations are robust and to clarify whether the observed effects are attributable to breed differences or to gene effects themselves.

Shifts in keratin isoform expression activate motility signals during wound healing

Keratin intermediate filaments confer structural stability to epithelial tissues, but the reason this simple mechanical function requires a protein family with 54 isoforms is not understood. During skin wound healing, a shift in keratin isoform expression alters the composition of keratin filaments. If and how this change modulates cellular functions that support epidermal remodeling remains unclear. We report an unexpected effect of keratin isoform variation on kinase signal transduction. Increased expression of wound-associated keratin 6A, but not of steady-state keratin 5, potentiated keratinocyte migration and wound closure without compromising mechanical stability by activating myosin motors to increase contractile force generation. These results substantially expand the functional repertoire of intermediate filaments from their canonical role as mechanical scaffolds to include roles as isoform-tuned signaling scaffolds that organize signal transduction cascades in space and time to influence epithelial cell state.

Tandem LIM domain-containing proteins, LIMK1 and LMO1, directly bind to force-bearing keratin intermediate filaments

The cytoskeleton of the cell is constantly exposed to physical forces that regulate cellular functions. Selected members of the LIM (Lin-11, Isl-1, and Mec-3) domain-containing protein family accumulate along force-bearing actin fibers, with evidence supporting that the LIM domain is solely responsible for this force-induced interaction. However, LIM domain's force-induced interactions are not limited to actin. LIMK1 and LMO1, both containing only two tandem LIM domains, are recruited to force-bearing keratin fibers in epithelial cells. This unique recruitment is mediated by their LIM domains and regulated by the sequences outside the LIM domains. Based on invitro reconstitution of this interaction, LIMK1 and LMO1 directly interact with stretched keratin 8/18 fibers. These results show that LIM domain's mechano-sensing abilities extend to the keratin cytoskeleton, highlighting the diverse role of LIM proteins in force-regulated signaling.

Colonocyte keratins stabilize mitochondria and contribute to mitochondrial energy metabolism.

Keratin intermediate filaments form dynamic filamentous networks, which provide mechanical stability, scaffolding, and protection against stress to epithelial cells. Keratins and other intermediate filaments have been increasingly linked to the regulation of mitochondrial function and homeostasis in different tissues and cell types. While deletion of keratin 8 (K8-/-) in mouse colon elicits a colitis-like phenotype, epithelial hyperproliferation, and blunted mitochondrial ketogenesis, the role of K8 in colonocyte mitochondrial function and energy metabolism is unknown. We used two K8 knockout mouse models and CRISPR/Cas9 K8-/- colorectal adenocarcinoma Caco-2 cells to answer this question. The results show that K8-/- colonocyte mitochondria in vivo are smaller and rounder and that mitochondrial motility is increased in K8-/- Caco-2 cells. Furthermore, K8-/- Caco-2 cells displayed diminished mitochondrial respiration and decreased mitochondrial membrane potential compared with controls, whereas glycolysis was not affected. The levels of mitochondrial respiratory chain complex proteins and mitochondrial regulatory proteins mitofusin-2 and prohibitin were decreased both in vitro in K8-/- Caco-2 cells and in vivo in K8-/- mouse colonocytes, and reexpression of K8 into K8-/- Caco-2 cells normalizes the mitofusin-2 levels. Mitochondrial Ca2+ is an important regulator of mitochondrial energy metabolism and homeostasis, and Caco-2 cells lacking K8 displayed decreased levels and altered dynamics of mitochondrial matrix and cytoplasmic Ca2+. In summary, these novel findings attribute an important role for colonocyte K8 in stabilizing mitochondrial shape and movement and maintaining mitochondrial respiration and Ca2+ signaling. Further, how these metabolically compromised colonocytes are capable of hyperproliferating presents an intriguing question for future studies.NEW & NOTEWORTHY In this study, we show that colonocyte intermediate filament protein keratin 8 is important for stabilizing mitochondria and maintaining mitochondrial energy metabolism, as keratin 8-deficient colonocytes display smaller, rounder, and more motile mitochondria, diminished mitochondrial respiration, and altered Ca2+ dynamics. Changes in fusion-regulating proteins are rescued with reexpression of keratin 8. These alterations in colonocyte mitochondrial homeostasis contribute to keratin 8-associated colitis pathophysiology.

Identification of a Novel miR-195-5p/PNN Axis in Colorectal Cancer.

Pinin (PNN) is a desmosome-associated protein that reinforces the organization of keratin intermediate filaments and stabilizes the anchoring of the cytoskeleton network to the lateral surface of the plasma membrane. The aberrant expression of PNN affects the strength of cell adhesion as well as modifies the intracellular signal transduction pathways leading to the onset of CRC. In our previous studies, we characterized the role of miR-195-5p in the regulation of desmosome junctions and in CRC progression. Here, with the aim of investigating additional mechanisms related to the desmosome complex, we identified PNN as a miR-195-5p putative target. Using a public data repository, we found that PNN was a negative prognostic factor and was overexpressed in colon cancer tissues from stage 1 of the disease. Then, we assessed PNN expression in CRC tissue specimens, confirming the overexpression of PNN in tumor sections. The increase in intracellular levels of miR-195-5p revealed a significant decrease in PNN at the mRNA and protein levels. As a consequence of PNN regulation by miR-195-5p, the expression of KRT8 and KRT19, closely connected to PNN, was affected. Finally, we investigated the in vivo effect of miR-195-5p on PNN expression in the colon of AOM/DSS-treated mice. In conclusion, we have revealed a new mechanism driven by miR-195-5p in the regulation of desmosome components, suggesting a potential pharmacological target for CRC therapy.

The Tilapia Cyst Tissue Enclosing the Proliferating Myxobolus bejeranoi Parasite Exhibits Cornified Structure and Immune Barrier Function.

Myxozoa, a unique group of obligate endoparasites within the phylum Cnidaria, can cause emerging diseases in wild and cultured fish populations. Recently, the myxozoan Myxobolus bejeranoi has been identified as a prevalent pathogen infecting the gills of cultured hybrid tilapia, leading to systemic immune suppression and considerable mortality. Here, we employed a proteomic approach to examine the impact of M. bejeranoi infection on fish gills, focusing on the structure of the granulomata, or cyst, formed around the proliferating parasite to prevent its spread to surrounding tissue. Enrichment analysis showed increased immune response and oxidative stress in infected gill tissue, most markedly in the cyst's wall. The intense immune reaction included a consortium of endopeptidase inhibitors, potentially combating the myxozoan arsenal of secreted proteases. Analysis of the cyst's proteome and histology staining indicated that keratin intermediate filaments contribute to its structural rigidity. Moreover, we uncovered skin-specific proteins, including a grainyhead-like transcription factor and a teleost-specific S100 calcium-binding protein that may play a role in epithelial morphogenesis and cysts formation. These findings deepen our understanding of the proteomic elements that grant the cyst its distinctive nature at the critical interface between the fish host and myxozoan parasite.

Ectopic Expression of a Truncated Isoform of Hair Keratin 81 in Breast Cancer Alters Biophysical Characteristics to Promote Metastatic Propensity.

Keratins are an integral part of cell structure and function. Here, it is shown that ectopic expression of a truncated isoform of keratin 81 (tKRT81) in breast cancer is upregulated in metastatic lesions compared to primary tumors and patient-derived circulating tumor cells, and is associated with more aggressive subtypes. tKRT81 physically interacts with keratin 18 (KRT18) and leads to changes in the cytosolic keratin intermediate filament network and desmosomal plaque formation. These structural changes are associated with a softer, more elastically deformable cancer cell with enhanced adhesion and clustering ability leading to greater in vivo lung metastatic burden. This work describes a novel biomechanical mechanism by which tKRT81 promotes metastasis, highlighting the importance of the biophysical characteristics of tumor cells.

In vitro effects of wool-derived keratin on human dental pulp-derived stem cells for endodontic applications

In this study we examine the influence of wool-derived keratin intermediate filament proteins (kIFPs) on human dental pulp-derived stem cells (hDPSCs). kIFPs were diluted (10 mg/mL to 0.001 mg/mL) in cell culture media. Effects on hDPSCs proliferation were measured using Alamar blue assay. Keratin concentrations of 1 mg/mL and 0.1 mg/mL were tested for odontogenic differentiation and mineralisation. Alkaline phosphatase (ALP) quantification (7th, 14th, and 21st days), alizarin red S (AR-S) staining and calcium quantification (21st day), reverse transcription polymerase chain reaction (RT-PCR, collagen expression), and immunocytochemical staining for dentin matrix protein (DMP) were performed. hDPSCs showed higher proliferation with kIFPs of 0.1 mg/mL or less (p < 0.0001). The 0.1 mg/mL keratin concentration promoted odontogenic differentiation, confirmed by increased ALP activity, significant calcium deposits (AR-S staining, p < 0.05), up-regulated collagen expression (RT-PCR, p < 0.05), and positive DMP staining. These results suggest that kIFPs could be a potential biomaterial for pulp-dentin regeneration.

Investigation of lncRNA in Bos taurus Mammary Tissue during Dry and Lactation Periods.

This study aims to collect RNA-Seq data from Bos taurus samples representing dry and lactating mammary tissue, identify lncRNA transcripts, and analyze findings for their features and functional annotation. This allows for connections to be drawn between lncRNA and the lactation process. RNA-Seq data from 103 samples of Bos taurus mammary tissue were gathered from publicly available databases (60 dry, 43 lactating). The samples were filtered to reveal 214 dry mammary lncRNA transcripts and 517 lactating mammary lncRNA transcripts. The lncRNAs met common lncRNA characteristics such as shorter length, fewer exons, lower expression levels, and less sequence conservation when compared to the genome. Interestingly, several lncRNAs showed sequence similarity to genes associated with strong hair keratin intermediate filaments. Human breast cancer research has associated strong hair keratin filaments with mammary tissue cellular resilience. The lncRNAs were also associated with several genes/proteins that linked to pregnancy using expression correlation and gene ontology. Such findings indicate that there are crucial relationships between the lncRNAs found in mammary tissue and the development of the tissue, to meet both the animal's needs and our own production needs; these relationships should be further investigated to ensure that we continue to breed the most resilient, efficient dairy cattle.

Alterations promoted by acid straightening and/or bleaching in hair microstructures.

Human hair is a biopolymer constituted mainly of keratin intermediate filaments, lipids, pigments and water. Cosmetic treatments usually interact with the hair at the molecular level, inducing changes in its components and modifying the physicochemical and mechanical properties of the fibers. Here, the effect of acid straightening on the morphology and ultrastructure of Caucasian hair was investigated by a group of complementary experimental methods: wide-, small- and ultra-small-angle X-ray scattering; high-resolution 3D X-ray microscopy; quasi-elastic neutron scattering and inelastic neutron scattering; thermogravimetry-mass spectrometry; and differential scanning calorimetry (DSC). X-ray diffraction patterns showed that acid straightening associated with a flat iron (∼180°C) changed the cortex of the fiber, shown by denaturation of the intermediate filaments (measured by DSC). The increase in the spacing of the lipid layers and the observation of the dehydration behavior of the fiber provided indications that water may be confined between these layers, while neutron spectroscopy showed alterations in the vibration mode of the CH2 groups of the lipids and an increase of the proton (H+) mobility in the hair structure. The latter may be associated with the extremely low pH of the formulation (pH ≃ 1). Additionally, this investigation showed that bleached hair (one-time bleached) is more damaged by the action of acid straightening than virgin hair, which was shown by a threefold increase in the percentage of total porosity of the tresses. The obtained results demonstrate that the investigation approach proposed here can provide very important thermodynamic and structural information on induced changes of hair structure, and certainly can be applied for the evaluation of the action mode and efficiency of cosmetic treatments.

Deacetylation via SIRT2 prevents keratin-mutation-associated injury and keratin aggregation.

Keratin (K) and other intermediate filament (IF) protein mutations at conserved arginines disrupt keratin filaments into aggregates and cause human epidermolysis bullosa simplex (EBS; K14-R125C) or predispose to mouse liver injury (K18-R90C). The challenge for more than 70 IF-associated diseases is the lack of clinically utilized IF-targeted therapies. We used high-throughput drug screening to identify compounds that normalized mutation-triggered keratin filament disruption. Parthenolide, a plant sesquiterpene lactone, dramatically reversed keratin filament disruption and protected cells and mice expressing K18-R90C from apoptosis. K18-R90C became hyperacetylated compared with K18-WT and treatment with parthenolide normalized K18 acetylation. Parthenolide upregulated the NAD-dependent SIRT2, and increased SIRT2-keratin association. SIRT2 knockdown or pharmacologic inhibition blocked the parthenolide effect, while site-specific Lys-to-Arg mutation of keratin acetylation sites normalized K18-R90C filaments. Treatment of K18-R90C-expressing cells and mice with nicotinamide mononucleotide had a parthenolide-like protective effect. In 2 human K18 variants that associate with human fatal drug-induced liver injury, parthenolide protected K18-D89H- but not K8-K393R-induced filament disruption and cell death. Importantly, parthenolide normalized K14-R125C-mediated filament disruption in keratinocytes and inhibited dispase-triggered keratinocyte sheet fragmentation and Fas-mediated apoptosis. Therefore, keratin acetylation may provide a novel therapeutic target for some keratin-associated diseases.

BG06 Beyond keratin: when simplex is not so simple

Abstract Epidermolysis bullosa simplex (EBS) is a heterogeneous, congenital, mucocutaneous fragility syndrome, characterized ultrastructurally by cleavage within the basal keratinocyte. Seventy-five per cent of cases are attributable to mutations in KRT5 and KRT14, the vast majority of which are dominant negative mutations, resulting in impaired keratin intermediate filament dynamics within the basal layer of the epidermis. The presenting phenotype, including extracutaneous manifestations and overall severity, is often closely linked to the underlying genetic mutation; however, genotype–­phenotype correlation is poorly resolved in a minority of cases, and there is an approximately additional 15% of patients in whom genetic abnormalities are not identified. However, recent advances in next-generation sequencing technologies have expanded the mutational database. We describe two cases of EBS with rare underlying mutations and distinct clinical phenotypes. A 46-year-old man presented with widespread blistering primarily affecting his hands, feet, elbows and knees since childhood. He had unaffected, consanguineous parents and one of his three siblings was affected. Other presenting features included dystrophic nails, focal plantar keratoderma, truncal mottled pigmentation and large areas of hypopigmentation on the back. We considered a diagnosis of ‘EBS with mottled pigmentation’, but the autosomal recessive mode of inheritance and dystrophic nails were atypical. DNA whole-exome sequencing (WES) and bidirectional Sanger sequencing revealed a novel homozygous PLEC mutation [c.94dupG p.(Ala32GlyfsTer3)]. Pathogenic PLEC variants are known to cause autosomal dominant EBS, the Ogna type (MIM131950), autosomal recessive EBS with muscular dystrophy (MIM226670) and autosomal recessive EBS with pyloric atresia (MIM612138). To date, our patient has not manifested any signs of progressive muscle weakness, but the presenting age can vary. The second patient was a 30-year-old woman who presented with blistering on her flexures, feet, hands and friction sites since the age of 4 months. She had unaffected, consanguineous parents and eight unaffected siblings. WES and confirmatory bidirectional Sanger sequencing identified a homozygous nonsense mutation in DST [c.3370C&amp;gt;T p.(Gln1124Ter)]. DST encodes the epithelial isoform of bullous pemphigoid antigen-1, present in the skin, muscle and neuronal tissues. Only 11 patients with DST mutation-induced EBS have been reported to date. The cutaneous phenotype is reported to be blisters, postinflammatory dyspigmentation, scarring, nail dystrophy, keratoderma and hyperhidrosis. If the affected exon is present in neuronal isoforms and neuronal tissue, neurological symptoms are expected. Unlike the patient reported elsewhere who harbours exactly the same mutation, our patient has reported no neurological issues to date. These cases highlight the importance of unravelling genotype–phenotype correlation in rare autosomal recessive forms of EBS, with possible implications for future prognosis and management.

Retromer Chaperones: Potential Therapeutics for Treatment of Skin Disease?

Retromer Chaperones: Potential Therapeutics for Treatment of Skin Disease?

An in vitro analysis of the hemostatic efficacy of fibrinogen precipitation with varied keratin fraction compositions

In preclinical studies, human hair has demonstrated effective hemostatic properties, potentially attributed to keratin proteins facilitating rapid conversion of fibrinogen to fibrin during coagulation. However, the rational use of human hair keratin for hemostasis remains unclear, given its complex mixture of proteins with diverse molecular weights and structures, leading to variable hemostatic capacity. To optimize the rational utilization of human hair keratin for hemostasis, we investigated the effects of different keratin fractions on keratin-mediated fibrinogen precipitation using a fibrin generation assay. Our study focused on high molecular weight keratin intermediate filaments (KIFs) and lower molecular weight keratin-associated proteins (KAPs) combined in various ratios during the fibrin generation. Scanning electron microscope analysis of the precipitates revealed a filamentous pattern with a broad distribution of fiber diameters, likely due to the diversity of keratin mixtures involved. An equal proportion of KIFs and KAPs in the mixture yielded the most extensive precipitation of soluble fibrinogen in an in vitro study, potentially due to structure-induced exposure of active sites. However, all hair protein samples exhibited diverse catalytic behaviors compared to thrombin, highlighting the potential of utilizing specific hair fractions to develop hair protein-based hemostatic materials with optimized capacity.

Architecture and dynamics of a desmosome-endoplasmic reticulum complex.

The endoplasmic reticulum (ER) forms a dynamic network that contacts other cellular membranes to regulate stress responses, calcium signalling and lipid transfer. Here, using high-resolution volume electron microscopy, we find that the ER forms a previously unknown association with keratin intermediate filaments and desmosomal cell-cell junctions. Peripheral ER assembles into mirror image-like arrangements at desmosomes and exhibits nanometre proximity to keratin filaments and the desmosome cytoplasmic plaque. ER tubules exhibit stable associations with desmosomes, and perturbation of desmosomes or keratin filaments alters ER organization, mobility and expression of ER stress transcripts. These findings indicate that desmosomes and the keratin cytoskeleton regulate the distribution, function and dynamics of the ER network. Overall, this study reveals a previously unknown subcellular architecture defined by the structural integration of ER tubules with an epithelial intercellular junction.

General aspects on skin development in vertebrates with emphasis on sauropsids epidermis

General cellular aspects of skin development in vertebrates are presented with emphasis on the epidermis of sauropsids. Anamniote skin develops into a multilayered mucogenic and soft keratinized epidermis made of Intermediate Filament Keratins (IFKs) that is reinforced in most fish and few anurans by dermal bony and fibrous scales. In amniotes, the developing epidermis in contact with the amniotic fluid initially transits through a mucogenic phase recalling that of their anamniotes progenitors. A new gene cluster termed EDC (Epidermal Differentiation Complex) evolved in amniotes contributing to the origin of the stratum corneum. The EDC contains numerous genes coding for over 100 types of corneous proteins (CPs). In sauropsids 2–8 layers of embryonic epidermis accumulate soft keratins (IFKs) but do not form a compact corneous layer. The embryonic epidermis of reptiles and birds produces small amount of other, poorly known proteins in addition to IFKs and mucins. In the following development, a resistant corneous layer is formed underneath the embryonic epidermis that is shed before hatching. The definitive corneous epidermis of sauropsids is mainly composed of CBPs (Corneous beta proteins, formerly indicated as beta-keratins) derived from the EDC. CBPs belong to a gene sub-family of CPs unique for sauropsids, contain an inner amino acid region formed by beta-sheets, are rich in cysteine and glycine, and make most of the protein composition of scales, claws, beaks and feathers. In mammalian epidermis CPs missing the beta-sheet region are instead produced, and include loricrin, involucrin, filaggrin and various cornulins. Small amount of CPs accumulate in the 2–3 layers of mammalian embryonic epidermis and their appendages, that is replaced with the definitive corneous layers before birth. Differently from sauropsids, mammals utilize KAPs (keratin associated proteins) rich in cysteine and glycine for making the hard corneous material of hairs, claws, hooves, horns, and occasionally also scales.

Development and structure of the skin in the Australian lungfish (Neoceratodus forsteri) in relation to epidermal adaptation of tetrapods

AbstractAnalysis of skin development in the Australian lungfish, Neoceratodus forsteri, indicates that keratinocytes form 2 to 3 irregular layers in tadpoles of 2–5 cm in length and are fully stratified (7–12 layers) in juveniles of 16.5–26 cm in length. The epidermis produces mucus and mainly contains intermediate filament keratins of acidic nature but also a minute amount of neutral‐basic keratins (pI 7.0–7.3), which is unique among fish. The latter keratins of 60–62 kDa, are generally present in tetrapods but the specific gene structure and amino sequence of acidic, neutral and basic keratins in N. forsteri are presently unknown. No other epidermal proteins are detected although biochemical data suggest the presence of non‐keratin proteins, including a basic type (pI 8.0–8.2) of around 32 kDa of molecular weight. The spinulated bony layer of elasmoid scales contains glycoproteins involved in mineralization, such as osteonectin and osteopontin. Also, alkaline phosphatase, involved in calcium precipitation onto the collagen matrix, has been detected in scales of juveniles up to 26 cm in length. The present review concludes that the knowledge of the data derived from epidermal and dermal protein annotation from the recently sequenced genome of this fish species will contribute to address the study on the evolution of tetrapod skin.

Mechanical anisotropy of hair affected by genetic diseases highlights structural information related to differential crosslinking in keratins.

Previous work with Atomic Force Microscope (AFM) nanoindentation, on longitudinal and cross-sections of the human hair fibre, allowed for the derivation of a model for the mechanical behaviour of human hair, called the Anisotropic Index. Expanding that research further, and by applying this model, the nanomechanical behaviour of hairs from patients with the disease Trichothiodystrophy (TTD) has been examined and structural insights, gained from combining the AFM results with Differential Scanning Calorimetry (DSC) experiments and tensile measurements, suggests that TTD-affected hairs have a relatively increased amount of Keratin Intermediate Filaments, contained in compartments of differing crosslinking extent. The associated calculations of axial and transverse Young's Moduli deliver values in good agreement with the measured fibre mechanics. Furthermore, comparing these findings with the results previously obtained from the study of hairs from patients with the disease Monilethrix, it is shown that the Anisotropic Index correlates well with the known deficiencies in both hair types obtained from such patients and allows for discerning between the Control hair and from those affected by the two diseases. AFM nanoindentation along and across the fibre axis and the Anisotropic Index thus appear to reveal structural details of hair not otherwise acquirable, whilst DSC may offer a quick and simple method for distinguishing between different severities of TTD.