Regulation of skin keratin intermediate filament proteins by phosphorylation.

摘要

Keratins are a group of cytoskeletal polymers belonging to the superfamily of intermediate filament (IF) proteins. Genetic and biochemical evidences support two major roles for keratins and other IF proteins. One is family-wide, context-independent, and consists of mechanical support, the abrogation of which leads to cell fragility and clinical diseases. The other occurs in an IF protein-specific and context-dependent fashion, and consists of regulation of several basic metabolic processes (e.g., cell survival, growth, and death. Consistent with this, all IF proteins are subject to regulation via post-translational modifications, including phosphorylation. Previous studies have provided detailed insight into how phosphorylation can regulate the properties and functions of IF proteins, though there is a paucity of knowledge about such keratin regulatory mechanisms in skin epithelium.;Here, we set out to study phosphorylation of mouse keratin 17 (K17) on serine 44 (Ser44), and report that it is phosphorylated in response to extracellular stimuli promoting keratinocyte growth and to various cellular stresses. K17 Ser44 phosphorylation also occurs in basaloid tumors in mouse skin in situ. Upon its stimulation, phosphorylation of K17-Ser44 is induced rapidly but stays on transiently. The majority of the phosphorylated K17-Ser44 pool is polymer-bound and is not obviously related to a change in filament organization. We show that p90 RSK1, an AGC kinase involved in the regulation of cell survival and proliferation, phosphorylates K17-Ser44 in skin keratinocytes. These findings confirm and expand the tight link that has emerged between K17 up-regulation and growth and stress responses in the skin epithelium.;We also mapped global phosphorylation sites on several keratins expressed in the skin in collaboration with Dr. Jerry Hart and colleagues at the Johns Hopkins School of Medicine. Utilizing a shot-gun mass spectrometry approach, we uncovered multiple phosphorylation events occurring on type I keratins K17 and K14 as well as ones on type II keratins K5 and K6. Although the biological significance of these sites is currently unknown, this effort provides a foundation for future investigation regarding the role of site-specific phosphorylation on the regulation of keratin properties and function in skin keratinocytes.