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Characterization of keratins and associated proteins involved in the corneification of crocodilian epidermis
No full textCrocodilian keratinocytes accumulate keratin and form a corneous cell envelope of which the composition is poorly known. The present immunological study characterizes the molecular weight, isoelectric point (pI) and the protein pattern of alpha- and beta-keratins in the epidermis of crocodilians. Some acidic alpha-keratins of 47-68 kDa are present. Cross-reactive bands for loricrin (70, 66, 55 kDa), sciellin (66, 55-57 kDa), and filaggrin-AE2-positive keratins (67, 55 kDa) are detected while caveolin is absent. These proteins may participate in the formation of the cornified cell membranes, especially in hinge regions among scales. Beta-keratins of 17-20 kDa and of prevalent basic pI (7.0-8.4) are also present. Acidic beta-keratins of 10-16 kDa are scarce and may represent altered forms of the original basic proteins. Crocodilian beta-keratins are not recognized by a lizard beta-keratin antibody (A68B), and by a turtle beta-keratin antibody (A685). This result indicates that these antibodies recognize specific epitopes in different reptiles. Conversely, crocodilian beta-keratins cross-react with the Beta-universal antibody indicating they share a specific 20 amino acid epitope with avian beta-keratins. Although crocodilian beta-keratins are larger proteins than those present in birds our results indicate presence of shared epitopes between avian and crocodilian beta-keratins which give good indication for the future determination of the sequence of these proteins. (C) 2007 Elsevier Ltd. All rights reserved
Distribution and characterization of proteins associated with cornification in the epidermis of gecko lizard
No full textThe distribution and molecular weight of epidermal proteins of gecko lizards have been studied by ultrastructural, autoradiographic, and immunological methods. Setae of the climbing digital pads are cross-reactive to antibodies directed against a chick scutate scale beta-keratin but not against feather beta-keratin. Cross-reactivity for mammalian loricrin, sciellin, filaggrin, and transglutaminase are present in alpha-keratogenic layers of gecko epidermis. Alpha-keratins have a molecular weight in the range 40-58 kDa. Loricrin cross-reactive bands have molecular weights of 42, 50, and 58 kDa. Bands for filaggrin-like protein are found at 35 and 42 kDa, bands for sciellin are found at 40-45 and 50-55 kDa, and bands for transglutaminase are seen at 48-50 and 60 kDa. The specific role of these proteins remains to be elucidated. After injection of tritiated histidine, the tracer is incorporated into keratin and in setae. Tritiated proline labels the developing setae of the oberhautchen and beta layers, and proline-labeled proteins (beta-keratins) of 10-14, 16-18, 22-24 and 32-35 kDa are extracted from the epidermis. In whole epidermal extract (that includes the epidermis with corneous layer and the setae of digital pads), beta-keratins of low-molecular weight (10, 14-16, and 18-19 kDa) are prevalent over those at higher molecular weight (34 and 38 kDa). In contrast, in shed epidermis of body scales (made of corneous layer only while setae were not collected), higher molecular weight beta-keratins are present (25-27 and 30-34 kDa). This suggests that a proportion of the small beta-keratins present in the epidermis of geckos derive from the differentiating beta layer of scales and from the setae of digital pads. Neither small nor large beta-keratins of gecko epidermis cross-react with an antibody specifically directed against the feather beta-keratin of 10-12 kDa. This result shows that the 10 and 14-16 kDa beta-keratins of gecko (lepidosaurian) have a different composition than the 10-12 kDa beta-keratin of feather (archosaurian). It is suggested that the smaller beta-keratins in both lineages of sauropsids were selected during evolution in order to build elongated bundles of keratin filaments to make elongated cells. Larger beta-keratins in reptilian scales produce keratin aggregations with no orientation, used for mechanical protection. © 2005 Elsevier Ltd. All rights reserved
Characterization of beta-keratins in lizard epidermis: Electrophoresis, immunocytochemical and in situ-hybridization study
No full textLizard scales are composed of alpha-(cyto-) keratins and beta-keratins. The characterization of the molecular weight and isoelectric point (pI) of alpha- and beta-keratins of lizard epidermis (Podarcis sicula) has been done by using two-dimensional electrophoresis, immunoblotting, and immunocytochemistry. Antibodies against cytokeratins, against a chicken scale beta-keratin or against lizard beta-keratin bands of 15-16 kDa, have been used to recognize alpha- and beta-keratins. Acid and basic cytokeratins of 42-67 kDa show a pI from 5.0 to 8.9. This indicates the presence of specific keratins for the formation of the stratum corneum. Main protein spots of beta-keratin at 15-17 kDa, and pI at 8.5, 8.2, and 6.7, and one spot at 10 kDa and pI at 7.3 were recognized. Therefore, beta-keratins are mainly basic proteins, and are used for the formation of the hard corneous layer of the epidermis. Ultrastructural immunocytochemistry confirms that beta-keratin is packed into large and dense bundles of beta-keratin cells of lizard epidermis. The use of a probe against a lizard beta-keratin in situ-hybridization studies confirms that the mRNA for beta-keratins is present in beta-cells and is localized around or even associated with beta-keratin filaments. (c) 2006 Elsevier Ltd. All rights reserved
Immuno-cross reactivity of transglutaminase and cornification marker proteins in the epidermis of vertebrates suggests common processes of soft cornification across species
No full textIn differentiating mammalian keratinocytes proteins are linked to the plasma membrane by epidermal transglutaminases through N-ε-(γ- glutamyl)-lysine isopeptide bonds to form the cornified cell envelope. The presence of transglutaminases and their protein substrates in the epidermis of nonmammalian vertebrates is not known. The present study analyses the presence and localization of the above proteins in the epidermis using immuno-cross reactivity across different classes of amniotes. After immunoblotting, some protein bands appear labelled for loricrin, sciellin, and transglutaminase in most species. These proteins are scarce to absent in the epidermis of aquatic species (goldfish and newt) where a stratum corneum is absent or very thin. The molecular weight of transglutaminase immunoreactive bands generally varies between 40 to 62 kDa, with the most represented bands at 52-57 kDa in most species. The more intense loricrin- and sciellin-immunoreactive bands are seen at 50-55-62 kDa, but are weak or absent in aquatic vertebrates. Loricrine-like immunoreactivity is present in the epidermis where α-(soft)-keratinization occurs. Isopeptide bonds are mainly associated to bands in the range of 50-62 kDa. In vertebrates where hard-keratin is expressed (the beta-keratin corneous layer of sauropsids and in feathers) or in hair cortex of mammals, no loricrin-like, transglutaminase-, and isopeptide-bond-immunoreactivities are seen. Immunoblotting however shows loricrin-, sciellin-, and trasnsglutaminase-positive bands in the corneous layers containing β-keratin. Histologically, the epidermis of most amniotes shows variable transglutaminase immunoreactivity, but isopeptide-bond and sciellin immunoreactivities are weak or undetactable in most species. The limitations of immunohistochemical methods are discussed and compared with results from immunoblotting. In reptilian epidermis transglutaminase is mainly localized in 0.15-0.3 μm dense granules or diffuse in transitional α-keratogenic cells. In β-keratogenic cells few small dense granules show a weak immunolabeling. Transglutaminase is present in nuclei of terminal differentiating α- and β-keratinocytes, as in those of mature inner and outer root sheath. The present study suggests that keratinization based on loricrin, sciellin and transglutaminase was probably present in the stratum corneoum of basic amniotes in the Carboniferous. These proteins were mainly maintained in α-keratogenic layers of amniotes but decreased in β-keratogenic layers of sauropsids (reptiles and birds). The study suggests that similar proteins for the formation of the cornified cell envelope are present in α-keratinocytes across vertebrates but not in β-keratinocytes. © 2004 Wiley-Liss, Inc
Ultrastructural Localization of Hair Keratin Homologs in the Claw of the Lizard Anolis carolinensis
No full texthe claw of lizards is largely composed of beta-keratins, also referred to as keratin-associated beta-proteins. Recently, we have reported that the genome of the lizard Anolis carolinensis contains alpha keratin genes homologous to hair keratins typical of hairs and claws of mammals. Molecular and immunohistochemical studies demonstrated that two hair keratin homologs named hard acid keratin 1 (HA1) and hard basic keratin 1 (HB1) are expressed in keratinocytes forming the claws of A. carolinensis. Here, we extended the immunocytochemical localization of the novel reptilian keratins to the ultrastructural level. After sectioning, claws were subjected to immunogold labeling using antibodies against HA1, HB1, and, for comparison, beta-keratins. Electron microscopy showed that the randomly organized network of tonofilaments in basal and suprabasal keratinocytes becomes organized in long and parallel bundles of keratin in precorneous layers, resembling cortical cells of hairs. Entering the cornified part of the claw, the elongated corneous cells fuse and accumulate corneous material. HA1 and HB1 are absent in the basal layer and lower spinosus layers of the claw and are expressed in the upper and precorneous layers, including the elongating corneocytes. The labeling for alpha-keratin was loosely associated with filament structures forming the fibrous framework of the claws. The ultrastructural distribution pattern of hard alpha-keratins resembled that of beta-keratins, which is compatible with the hypothesis of an interaction during claw morphogenesis. The data on the ultrastructural localization of hair keratin homologs facilitate a comparison of lizard claws and mammalian hard epidermal appendages containing hair keratins
Anaerobic Digestion of Microalgal Residues to Enhance the Energetic Profit of Biocrude Production
No full textBioinformatic and molecular characterization of cathelicidin-like peptides isolated from the green lizard Anolis carolinensis (Reptilia: Lepidosauria: Iguanidae)
No full textDistribution of Specific Keratin-Associated Beta-Proteins (Beta-Keratins) in the Epidermis of the Lizard Anolis carolinensis Helps to Clarify the Process of Cornification in Lepidosaurians
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