Complete Cytolysis and Neonatal Lethality in Keratin 5 Knockout Mice Reveal Its Fundamental Role in Skin Integrity and in Epidermolysis Bullosa Simplex

Abstract

In human patients, a wide range of mutations in keratin (K) 5 or K14 lead to the blistering skin disorder epidermolysis bullosa simplex. Given that K14 deficiency does not lead to the ablation of a basal cell cytoskeleton because of a compensatory role of K15, we have investigated the requirement for the keratin cytoskeleton in basal cells by inactivating the K5 gene in mice. We report that the K5 Ϫ/Ϫ mice die shortly after birth, lack keratin filaments in the basal epidermis, and are more severely affected than K14 Ϫ/Ϫ mice. In contrast to the K14 Ϫ/Ϫ mice, we detected a strong induction of the wound-healing keratin K6 in the suprabasal epidermis of cytolyzed areas of postnatal K5 Ϫ/Ϫ mice. In addition, K5 and K14 mice differed with respect to tongue lesions. Moreover, we show that in the absence of K5 and other type II keratins, residual K14 and K15 aggregated along hemidesmosomes, demonstrating that individual keratins without a partner are stable in vivo. Our data indicate that K5 may be the natural partner of K15 and K17. We suggest that K5 null mutations may be lethal in human epidermolysis bullosa simplex patients. INTRODUCTIONKeratin (K) intermediate filaments (IFs) belong to a gene family that is organized into two subfamilies, coding for type I (K9 -20) and type II keratins (K1-8). At least one member of each family is necessary to form heterodimeric IFs. In epidermis, keratins display a complex expression pattern that is widely assumed to reflect the structural requirements of distinct epidermal compartments (Fuchs and Cleveland, 1998).The major keratins in the basal layer of stratified epithelia are K5, K14, and K15 (Fuchs and Green, 1978, 1980;Moll et al., 1982;Leube et al., 1988;Lloyd et al., 1995). In wound healing or in hyperproliferative disorders, the keratin pair K6 and K16 is transiently expressed in the suprabasal layers instead of K1, K2e, and K10 (for review, see McGowan and Coulombe, 1998). The functional significance of the highly patterned expression profile of keratins is still poorly understood, but evidence is accumulating that they have distinct functions in epidermis (Paladini and Coulombe, 1999).The structural function of keratins was elucidated by the discovery of point mutations in human epidermal keratin genes (Corden and McLean, 1996), preceded by studies on transgenic mice expressing mutant keratin subunits . These mutations cause a number of inherited human skin disorders such as epidermolysis bullosa simplex (EBS) (Bonifas et al., 1991;Coulombe et al., 1991;Lane et al., 1992;Rothnagel et al., 1995;Corden and McLean, 1996). The characteristic feature of EBS is epidermal blistering resulting from cytolysis of basal keratinocytes. On the basis of these observations, it was suggested that the overall function of epidermal keratins was to provide the reinforcement of the epidermis and the maintenance of cellular integrity under mechanical and thermal stress. Several mice carrying null or dominant keratin mutations that affect epidermal integrity have added further support to t...

Keratins as the main component for the mechanical integrity of keratinocytes

“…The most prominent keratins of basal epidermal cells are the type II K5 and the type I keratins K14 and K15 (14). K5 deletion results in pronounced cytolysis and neonatal lethality (38), whereas K14 depletion leads to epidermolysis bullosa simplex-like disease where mechanical trauma exacerbates blistering (39). Assuming a similar role of K14 and K15, we decided to test for rescue of overall type I KO by reexpression of K14 only.…”

Desmosomes and Intermediate Filaments: Their Consequences for Tissue Mechanics

“…EBS is characterized by cytoplasmic keratin aggregates, cytolysis of basal keratinocytes, and bullous lesions following mild trauma to the skin. Although it is recognized that the pathomechanisms contributing to EBS and additional keratinopathies are more complex than originally considered Roth et al 2012;Bohnekamp et al 2015;Hobbs et al 2016;Kumar et al 2016), it is evident that loss of an intact keratin cytoskeleton renders keratinocytes fragile on mild physical stress, shown by KRT5 and KRT14 KO mice (Lloyd et al 1995;Peters et al 2001). Of note, even mutations causing severe disease do not prevent formation of long keratin intermediate filaments (KIFs) in vitro (Herrmann et al 2002), suggesting that mutations and physical stress act at the level of keratin bundling, network organization, dynamics, or by affecting association with other proteins.…”

Genética Molecular das Epidermólises Bolhosas

“…54 Modelo animal com ratos transgênicos, simulando a doença humana, tem acrescentado informações relevantes na pesquisa das EB. 10,12 Alguns autores consideram que as correlações entre genótipo e fenótipo estejam apenas começando 34 e que a expansão dos bancos de dados sobre as alterações gênicas seja de extrema importância, pois permitirá, cada vez mais, que se melhore essa correlação e talvez até se reclassifique, com base em aspectos moleculares, parte das genodermatoses. …”

“…É interessante observar que defeitos genéti-cos distintos na EBS, um afetando a citoqueratina 5, outro, a 14,6,8,9 levam à mesma alteração histológica, pois todos esses defeitos produzem alterações estruturais de uma ou outra citoqueratina, 10 impedindo a função estrutural das mesmas no citoesqueleto 11 -a formação dos heterodímeros, responsáveis pela configuração tridimensional da célula. Essa alteração é vista facilmente na histologia e culmina com a formação das bolhas, sendo esse o único subgrupo das EB decorrente de citólise e não de defeito de adesão.…”

“…1,17 Outro componente da placa interna do hemidesmossoma é o antígeno do penfigóide bolhoso de 230 KD de peso molecular, não havendo até o momento descrição de mutação no gen que o regula. 14,6,8,9 lead to the same histological alteration, because all these defects produce structural alterations in one or another cytokeratin, 10 impeding their structural function in the cytoskeleton 11 (Figure 2) 13,14 of the basal cytokeratins.…”