Abstract
Epidermal keratinocyte differentiation on the body surface is a carefully choreographed process that leads to assembly of a barrier that is essential for life. Perturbation of keratinocyte differentiation leads to disease. Activator protein 1 (AP1) transcription factors are key controllers of this process. We have shown that inhibiting AP1 transcription factor activity in the suprabasal murine epidermis, by expression of dominant-negative c-jun (TAM67), produces a phenotype type that resembles human keratoderma. However, little is understood regarding the structural and molecular changes that drive this phenotype. In the present study we show that TAM67-positive epidermis displays altered cornified envelope, filaggrin-type keratohyalin granule, keratin filament, desmosome formation and lamellar body secretion leading to reduced barrier integrity. To understand the molecular changes underlying this process, we performed proteomic and RNA array analysis. Proteomic study of the corneocyte cross-linked proteome reveals a reduction in incorporation of cutaneous keratins, filaggrin, filaggrin2, late cornified envelope precursor proteins, hair keratins and hair keratin-associated proteins. This is coupled with increased incorporation of desmosome linker, small proline-rich, S100, transglutaminase and inflammation-associated proteins. Incorporation of most cutaneous keratins (Krt1, Krt5 and Krt10) is reduced, but incorporation of hyperproliferation-associated epidermal keratins (Krt6a, Krt6b and Krt16) is increased. RNA array analysis reveals reduced expression of mRNA encoding differentiation-associated cutaneous keratins, hair keratins and associated proteins, late cornified envelope precursors and filaggrin-related proteins; and increased expression of mRNA encoding small proline-rich proteins, protease inhibitors (serpins), S100 proteins, defensins and hyperproliferation-associated keratins. These findings suggest that AP1 factor inactivation in the suprabasal epidermal layers reduces expression of AP1 factor-responsive genes expressed in late differentiation and is associated with a compensatory increase in expression of early differentiation genes.
Highlights
Previous studies suggest that activator protein 1 (AP1) factors are important regulators
of differentiation in the human epidermis.[1,2,55] The present study expands the number of potential genes that may be regulated by AP1 factors
altered expression of these genes is associated with other skin diseases
Summary
The TAM67-rTA mice used in this study are the offspring of a cross of hINV-rTA +/ − mice and TetO-TAM67-FLAG mice and are maintained in a hairless SKH1 background.[5] The hINV-rTA mice express the reverse-tetracycline activator (rTA) protein targeted to the suprabasal epidermis. Interaction with doxycycline converts the rTA protein to an active conformation that binds to the TetO in the TetO-TAM67-FLAG cassette to drive TAM67 expression. Treatment of TAM67rTA mice with doxycycline in the drinking water results in suprabasal TAM67FLAG expression.[4,5] The transgenes are maintained in the SKH-1 hairless genetic background to facilitate visualization of the epidermal phenotype.[5] Mice were
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