The epidermis of grhl3-null mice displays altered lipid processing and cellular hyperproliferation

Abstract

The presence of an impermeable surface barrier is an essential homeostatic mechanism in almost all living organisms. We have recently described a novel gene that is critical for the developmental instruction and repair of the integument in mammals. This gene, Grainy head-like 3 (Grhl3) is a member of a large family of transcription factors that are homologs of the Drosophila developmental gene grainy head (grh). Mice lacking Grhl3 fail to form an adequate skin barrier, and die at birth due to dehydration. These animals are also unable to repair the epidermis, exhibiting failed wound healing in both fetal and adult stages of development. These defects are due, in part, to diminished expression of a Grhl3 target gene, Transglutaminase 1 (TGase 1), which encodes a key enzyme involved in cross-linking of epidermal structural proteins and lipids into the cornified envelope (CE). Remarkably, the Drosophila grh gene plays an analogous role, regulating enzymes involved in the generation of quinones, which are essential for cross-linking structural components of the fly epidermis. In an extension of our initial analyses, we focus this report on additional defects observed in the Grhl3-null epidermis, namely defective extra-cellular lipid processing, altered lamellar lipid architecture and cellular hyperproliferation. These abnormalities suggest that Grhl3 plays diverse mechanistic roles in maintaining homeostasis in the skin.

Keywords: Grhl3; grainy head; lipids; proliferation; skin barrier.

Figure 1

Grhl3-null epidermis displays abnormal lamellar body contents, and decreased stratum corneum membrane structures. (A) Ultrastructural analysis by electron microscopy of skin sections from E18.5 embryos. In the wild type skin (left panel, +/+), the stratum corneum (SC) has normal multiple layers of corneocytes (C) with normal EM artefact-induced, well-spaced intercellular layers (asterisk). In comparison, although the number of corneocyte layers is similar in mutant skin (right panel, -/-), the intercellular layers are essentially non-existent (asterisk). Keratohyaline granules (arrows) seem somewhat larger in the mutant skin. Scale bar = 2 µm (B) Morphometric analysis of cornified envelope thickness from multiple E18.5 wild type (+/+) and mutant (-/-) epidermis sections shows a significant decrease (11.5%) in the overall CE thickness in the mutant epidermis. (C) Focusing on the lipid structures shows that the E18.5 wild type (+/+) stratum corneum displays a full complement of extracellular lamellar membrane structures (arrows) and that the lamellar bodies show multiple layers of lipid containing lamellae (insert of panel E, arrows). Whereas, the extracellular spaces of the (D) mutant (-/-) stratum corneum displays a paucity of lamellar bilayers (open arrows), although occasional fragments of replete lamellar arrays are seen (solid arrow). (E) This is as a result of almost all the lamellar organelles lacking internal contents (arrowheads) in the mutant epidermis, even though there are a normal complement of lamellar bodies. Panels (A & E), osmium tetroxide post-fixation; Panels (C & D), ruthenium tetroxide post-fixation. Previously published in Science 2005; 308:411–3.

Figure 2

Abnormalities in differentiation and CE formation in Grhl3-null epidermis. Skin from wild type (+/+) and mutant (-/-) embryos taken at E15.5 and E18.5 as indicated. Epithelial ridges (arrows) in wild type skin at E15.5 (top left panel) are absent in the E15.5 mutant skin (top right panel). Basal disorganisation and suprabasal hyperproliferation and a compacted SC are evident in the E18.5 mutant skin (bottom right) compared to the wild type (bottom left). d, dermis; sb, stratum basale; ss, stratum spinosum; sg, stratum granulosum; sc, stratum corneum. Previously published in Science 2005; 308:411–3.