Terminally differentiated epidermal keratinocytes form the body's outer protective barrier that undergoes constant renewal as cornified squames slough off the exterior surface and are replaced by material migrating outwards from the inner living layers of the epidermis. Cornified squames are composite biomaterials with two components: a matrix of keratin filaments encased within the cornified cell envelope (CE). The CE is a multi-component 10nm-thick layer of insoluble protein deposited on the inner surface of the plasma membrane of the cells during terminal differentiation. In the epidermis, a 5nm-thick layer of ceramide lipids (lipid envelope) is attached to the exterior surface. The insolubility of the protein envelope is due in large part to the cross-linking of several structural proteins by transglutaminases. Intermediate filaments (IF) are ubiquitous constituents of the cytoskeletons of eukaryotic cells. They consist of five different types, of which the most numerous and complex are the type I and type II keratins that are widely expressed in epithelia. We are interested also in the related IF of other cell types and in the interactions beetween keratin IF and the CE. The long-term goal of this project, previously pursued in both the Laboratory of Skin Biology (P. M. Steinert, P.I.) and the LSBR is to elucidate the production, assembly, structure and biomechanical properties of these cells and their components in the context of both normal and diseased skin. In FY05, we performed experiments to investigate interactions between the CE and the keratin intermediate filament network. In particular, we published a study that suggested that epiplakin is a cytolinker involved in maintaining the integrity of IF networks in simple epithelial cells and demonstrated the involvement of epiplakin in the process of keratinocyte differentiation.These results were based on immunohistochemistry experiments with knocked out or cotransfected epiplakin.In a second line of investigation, overlay experiments were performed. Involucrin, vimentin, keratins 5,14,8 and 18 and different epiplakin domains were recombinantly expressed, purified and MS-characterized by mass specttrometry. Dot-blot binding assay of purified monomeric or polymerized IF proteins with expressed epiplakin fragments revealed that the repeat unit (linker plus PRD) of epiplakin preferentially associates with assembled keratin filaments During FY05, a U.S. patent was granted to P.M. Steinert, L. N. Marekov, and Z. Nemes for Methods for Ameliorating Icthyosiform Skin Diseases. Progressive mitochondrial dysfunction may play a central role in the pathogenesis of several neurodegenerative diseases including Huntington's disease (HD). We found that TGase activity is increased about two fold in the mitochondrial fraction of HD caudate. A 78 kDa protein found to be progressively polymerized by TGase in isolated rat brain mitochondria was identified as mitochondrial aconitase (Aco 2) by MALDI-TOF analysis. As the enzyme is a part of the energy generating Krebs citric acid cycle its loss by polymerization will result in brain starvation.
| Markova, Nelli G; Karaman-Jurukovska, Nevena; Pinkas-Sarafova, Adriana et al. (2007) Inhibition of histone deacetylation promotes abnormal epidermal differentiation and specifically suppresses the expression of the late differentiation marker profilaggrin. J Invest Dermatol 127:1126-39 |
