RESUMO
The p53 family member p63 has been implicated in both the development and maintenance of stratified epithelial tissues, including the epidermis. Increasing data support p63 function in the regenerative capacity of basal keratinocytes by maintaining cell proliferation. Recent studies further suggest this regulation relies on inhibition of p53 activity. In addition, p63 appears to exert separate control over epidermal differentiation, which may involve control of such key signaling molecules as IKKalpha and Notch. While studies over the past decade have greatly expanded our knowledge of p63 function, much remains to be understood regarding how p63 regulates epidermal homeostasis. Future efforts to identify and validate direct p63 target genes as well as to understand the expression and function of individual p63 isoforms will be important to further define how p63 functions in the control of keratinocyte proliferation and differentiation.
Assuntos
Diferenciação Celular , Proliferação de Células , Proteínas de Ligação a DNA/fisiologia , Queratinócitos/citologia , Transativadores/fisiologia , Proteínas Supressoras de Tumor/fisiologia , Animais , Proteínas de Ligação a DNA/genética , Humanos , Queratinócitos/metabolismo , Modelos Biológicos , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiologia , Transativadores/genética , Fatores de Transcrição , Proteínas Supressoras de Tumor/genéticaRESUMO
p63 is a multi-isoform p53 family member required for epidermal development. Contrasting roles for p63 in either the initial commitment to the stratified epithelial cell fate or in stem cell-based self-renewal have been proposed. To investigate p63 function in a post-developmental context, we used siRNAs directed against p63 to down-regulate p63 expression in regenerating human epidermis. Loss of p63 resulted in severe tissue hypoplasia and inhibited both stratification and differentiation in a cell-autonomous manner. Although p63-deficient cells exhibited hypoproliferation, differentiation defects were not due to tissue hypoplasia. Simultaneous p63 and p53 knockdown rescued the cell proliferation defect of p63 knockdown alone but failed to restore differentiation, suggesting that defects in epidermal proliferation and differentiation are mediated via p53-dependent and -independent mechanisms, respectively. Furthermore, DeltaNp63 isoforms are the main mediators of p63 effects, although TAp63 isoforms may contribute to late differentiation. These data indicate that p63 is required for both the proliferative and differentiation potential of developmentally mature keratinocytes.
Assuntos
Diferenciação Celular , Proteínas de Ligação a DNA/metabolismo , Queratinócitos/citologia , Queratinócitos/fisiologia , Transativadores/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Ciclo Celular , Proliferação de Células , Proteínas de Ligação a DNA/genética , Regulação para Baixo , Células Epidérmicas , Humanos , Isoformas de Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transativadores/genética , Fatores de Transcrição , Proteína Supressora de Tumor p53/deficiência , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor/genéticaRESUMO
Phosphoinositide 3-kinases (PI3Ks) regulate an array of cellular processes and are comprised of three classes. Class I PI3Ks include the well-studied agonist-sensitive p110 isoforms; however, the functions of class II and III PI3Ks are less well characterized. Of the three class II PI3Ks, C2alpha and C2beta are widely expressed in many tissues, including the epidermis, while C2gamma is confined to the liver. In contrast to the class I PI3K p110alpha, which is expressed throughout the epidermis, C2beta was found to be localized in suprabasal cells, suggesting a potential role for C2beta in epidermal differentiation. Overexpressing C2beta in epidermal cells in vitro induced differentiation markers. To study a role for C2beta in tissue, we generated transgenic mice overexpressing C2beta in both suprabasal and basal epidermal layers. These mice lacked epidermal abnormalities. Mice deficient in C2beta were then generated by targeted gene deletion. C2beta knockout mice were viable and fertile and displayed normal epidermal growth, differentiation, barrier function, and wound healing. To exclude compensation by C2alpha, RNA interference was then used to knock down both C2alpha and C2beta in epidermal cells simultaneously. Induction of differentiation markers was unaffected in the absence of C2alpha and C2beta. These findings indicate that class II PI3Ks are not essential for epidermal differentiation.
Assuntos
Células Epidérmicas , Epiderme/enzimologia , Fosfatidilinositol 3-Quinases/metabolismo , Animais , Diferenciação Celular/genética , Células Cultivadas , Classe II de Fosfatidilinositol 3-Quinases , Deleção de Genes , Humanos , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Mutação , Fosfatidilinositol 3-Quinases/genética , Inibidores de Fosfoinositídeo-3 Quinase , Interferência de RNA , Ativação Transcricional , Cicatrização/genéticaRESUMO
14-3-3 proteins are a family of conserved dimeric molecules that interact with a broad range of target proteins, most of which contain phosphoserine/threonine. The amphipathic groove of 14-3-3 is the main structural feature involved in mediating its associations. We have studied another domain of 14-3-3, the C-terminal loop, to determine what role it plays in ligand interaction. A truncated form of 14-3-3zeta lacking this C-terminal loop was generated and found to bind with higher affinity than the wild-type 14-3-3zeta protein to the ligands Raf-1 and Bad. Interestingly, the truncated 14-3-3zeta also showed increased association with the 14-3-3 binding-deficient Bad/S136A mutant. Taken together, these data support a role for the C-terminal loop as a general inhibitor of 14-3-3/ligand interactions. This may provide a mechanism by which inappropriate associations with 14-3-3 are prevented.