Cholesterol Depletion Upregulates Involucrin Expression in Epidermal Keratinocytes Through Activation of p38

Abstract

Cholesterol has been recently suggested to regulate the early steps of keratinocyte differentiation through lipid rafts. In many cell types, depletion of cholesterol activates signaling proteins like epidermal growth factor receptor (EGFR), human epidermal growth factor receptor 2 (HER2), or extracellular signal-regulated kinase (ERK) known to affect cell differentiation. In this study, we explored the effects of cholesterol depletion on the phenotype of cultured keratinocytes, using a treatment with methyl-beta-cyclodextrin (MbetaCD) to extract cholesterol and a treatment with lovastatin to inhibit cholesterol neosynthesis. Analysis of the expression of differentiation marker genes in early differentiating confluent cultures reveals that cholesterol depletion induces downregulation of keratin 14 (K14) and keratin 10 (K10) and upregulation of involucrin. MbetaCD treatment induces phosphorylation of EGFR, HER2, and ERK, but not HER3. Inhibition of EGFR with PD153035 impairs the MbetaCD-induced phosphorylation of EGFR, HER2, and ERK, but does not impair the alteration of K14, K10, or involucrin gene expression, indicating that other signaling proteins regulate this phenomenon. p38 has been suggested to regulate the expression of involucrin during keratinocyte differentiation. We found that MbetaCD treatment induces a prolonged phosphorylation of p38 in general and p38alpha in particular. An inhibition of p38 with PD169316 impairs the upregulation of involucrin mRNAs by a treatment with MbetaCD, but not by a p38delta-activating TPA treatment, which might suggest that cholesterol depletion alters involucrin gene expression through activation of p38alpha/beta.

7-Dehydrocholesterol enhances ultraviolet A-induced oxidative stress in keratinocytes: Roles of NADPH oxidase, mitochondria, and lipid rafts

“…5A, row 4), The co-localization of caveolin-1 and CT-B was much less conspicuous than in HK and this image was very similar to the image of SLO-HK. The presence of lipid rafts was reflected in the cell morphology; CD treatment caused shrinkage and rounding of the cells, as previously reported [31]. This shape change, which was reversed when Chol was replaced, may be related to the known disruption of the actin cytoskeleton by CD [32].…”

Methyl-β-cyclodextrin Suppresses Hyaluronan Synthesis by Down-regulation of Hyaluronan Synthase 2 through Inhibition of Akt

“…Epidermal growth factor receptor (11), human ErbB2 (11), ERK (11-13), p38 (11,14), Src (15), and ␤-catenin (16) are activated by M␤CD treatment in various cell types. Like other cell lines, MCF-7 cells showed increased ERK phosphorylation following M␤CD treatment.…”

“…Effect of M␤CD on Akt and p70S6K Phosphorylation-Cholesterol depletion by M␤CD has been suggested to cause changes in the phosphorylation of signaling molecules like ERK and Src (11,15). Using a phosphokinase array we screened the phosphorylation profiles of 46 kinases and their substrates to find out if M␤CD affected the signaling pathways important in HAS2 regulation.…”

“…M␤CD disrupts lipid rafts (10) and activates many signaling proteins, like epidermal growth factor receptor (11), extracellular signal-regulated kinase (ERK) (12,13), p38 (11,14), Src (15), and ␤-catenin (16). On the other hand, M␤CD decreases the growth of MCF-7 breast cancer and A2780 ovarian cancer cells implanted subcutaneously in nude mice (17).…”

p38δ mitogen-activated protein kinase regulates skin homeostasis and tumorigenesis

“…45 Altogether, these data suggest a role for p38δ isoform in activation of differentiation-dependent involucrin gene expression. However, the induction of involucrin expression by treatment with cholesterol-depleting agents involves p38α, 46 and constitutively active MEK6 or MEK7, when overexpressed in keratinocytes, increase involucrin via a mechanism that involves p38α, 42,47 suggesting that the engagement of a specific p38 isoform may be stimulus-dependent. Interestingly, a genomic analysis has shown that MAPK13 (a gene that encodes p38δ protein) and involucrin are among genes commonly upregulated both in psoriasis, a benign inflammatory condition characterized by increased proliferation and differentiation, and in well-differentiated locally invasive cutaneous squamous cell carcinomas (SCC).…”