BINDING OF KERATIN INTERMEDIATE FILAMENTS (K10) TO THE CORNIFIED ENVELOPE IN MOUSE EPIDERMIS - IMPLICATIONS FOR BARRIER FUNCTION

The cornified envelope, a structure unique to keratinocytes, is a hall mark of their terminal differentiation and plays an important role in epidermal barrier function. Cornified envelope is formed through the a ction of a membrane-associated transglutaminase, which covalently cros s-links protein precursors into a highly insoluble network at the inne r leaflet of the plasma membrane in granular keratinocytes and stratum corneum. Initial studies, using dansylcadaverine for enzyme-directed labeling of acyl-acceptor transglutaminase substrates in mouse epiderm al homogenates identified a prominent 60-kDa substrate. Specific antib odies raised to this protein stained the cytoplasm of suprabasal cells of stratified squamous epithelia, whereas simple epithelia and nonepi thelial tissues showed no staining. Immunoscreening of a cDNA expressi on library from adult mouse skin identified 18 positive clones. DNA se quencing of the largest clone (which hybridized to a keratinocyte-spec ific transcript of 2.0 kb) showed greater than 99.5% homology with mou se keratin 10. Immunoelectron microscopy using anti-S60 and another an tibody to keratin 10 showed specific binding to cornified envelope ass ociated filamentous structures. Proteolytic fragments of purified corn ified envelope from mouse epidermis showed reactivity to anti-S60. The se data show that mouse keratin 10 is tightly bound to cornified envel ope and may be a cross-linked substrate. The tight binding of keratin filaments and CE suggests a mechanism by which they might interact to enhance the structural integrity of the stratum corneum.