C/EBPβ Modulates the Early Events of Keratinocyte Differentiation Involving Growth Arrest and Keratin 1 and Keratin 10 Expression

Abstract

The epidermis is a stratified squamous epithelium composed primarily of keratinocytes that become postmitotic and undergo sequential changes in gene expression during terminal differentiation. The expression of the transcription factor CCAAT/enhancer binding protein ␤ (C/EBP␤) within mouse epidermis and primary keratinocytes has recently been described; however, the function of C/EBP␤ within the epidermal keratinocyte is unknown. We report here that transient transfection of mouse primary keratinocytes with a C/EBPresponsive promoter-reporter construct resulted in a sevenfold increase in luciferase activity when keratinocytes were switched to culture conditions that induce growth arrest and differentiation. Forced expression of C/EBP␤ in BALB/MK2 keratinocytes inhibited growth, induced morphological changes consistent with a more differentiated phenotype, and upregulated two early markers of differentiation, keratin 1 (K1) and keratin 10 (K10) but had a minimal effect on the expression of late-stage markers, loricrin and involucrin. Analysis of the epidermis of C/EBP␤-deficient mice revealed a mild epidermal hyperplasia and decreased expression of K1 and K10 but not of involucrin and loricrin. C/EBP␤-deficient primary keratinocytes were partially resistant to calcium-induced growth arrest. Analysis of terminally differentiated spontaneously detached keratinocytes or those induced to differentiate by suspension culture revealed that C/EBP␤-deficient keratinocytes displayed striking decreases in K1 and K10, while expression of later-stage markers was only minimally altered. Our results demonstrate that C/EBP␤ plays an important role in the early events of stratified squamous differentiation in keratinocytes involving growth arrest and K1 and K10 expression.

Novel Protein Kinase C Isoforms Regulate Human Keratinocyte Differentiation by Activating a p38δ Mitogen-activated Protein Kinase Cascade That Targets CCAAT/Enhancer-binding Protein α

“…The present results are consistent with a model wherein C/EBP␣ enhances hINV gene expression. In contrast, C/EBP␤ and ␦ inhibits hINV expression in cultured cells (15) or does not influence hINV expression in C/EBP␤ knockout mice (38). This may, at least in part, explain why hINV is expressed in the suprabasal epidermal layers, where C/EBP␣ is expressed, and not in the basal layers, where C/EBP␤ predominates.…”

“…They begin as proliferative cells and are ultimately converted to nonproliferating cells called corneocytes that assemble the epidermal surface (12). This change involves a remarkable number of gene activation events that result in the expression of a group of proteins, including involucrin, that are designed to construct this barrier (12,37,38). Specific PKC isoforms are implicated in this process.…”

Conditional ablation of C/EBPβ demonstrates its keratinocyte-specific requirement for cell survival and mouse skin tumorigenesis

“…Epidermis was removed from the dermis as described (Oh and Smart, 1998) and all organs/tissues were homogenized in RIPA buffer containing 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, pH 7.4, 1 mM PMSF, 1 mM SOV, 1 Â GIBCO complete protease inhibitor cocktail. Samples (20 mg) were separated by SDS-PAGE and western analysis was conducted as described (Zhu et al, 1999). (b) Immunohistochemical staining for C/EBPb in skin of wild type and (c) K5-Cre;C/EBPb fl/fl mice.…”

Decreased survival of C/EBPβ-deficient keratinocytes is due to aberrant regulation of p53 levels and function

“…C/EBPb is expressed in many tissues, including epidermis (Maytin and Habener, 1998;Oh and Smart, 1998) where it plays a role in the early stages of stratified squamous differentiation (Zhu et al, 1999). Recent studies have identified a role for C/EBPb in tumor development (Zhu et al, 2002), senescence (Sebastian et al, 2005), cell survival (Buck et al, 2001;Zhu et al, 2002;Wessells et al, 2004;Li et al, 2005) and cellular transformation (Zhu et al, 2002).…”

“…Recent studies have identified a role for C/EBPb in tumor development (Zhu et al, 2002), senescence (Sebastian et al, 2005), cell survival (Buck et al, 2001;Zhu et al, 2002;Wessells et al, 2004;Li et al, 2005) and cellular transformation (Zhu et al, 2002). In terms of cell transformation, C/EBPb can cooperate with Ras12V to transform NIH3T3 cells (Zhu et al, 1999) and more recently, it has been shown that cell-cycledependent phosphorylation of C/EBPb on Ser64 and Thr189 is required to promote Ras-induced transformation of NIH3T3 cells (Shuman et al, 2004).…”