Epidermal protein kinase C-beta 2 is highly sensitive to downregulation and is exclusively expressed in Langerhans cells: downregulation is associated with attenuated contact hypersensitivity.
Treatment of mice with multiple topical applications of 12-O-tetradecanoylphorbol-13-acetate (TPA) or diacylglycerol resulted in a preferential decrease in epidermal protein kinase C-beta 2 (PKC-beta 2) compared with PKC-alpha as determined by western analysis. When PKC-alpha was decreased by 40%, PKC-beta 2 could no longer be detected, suggesting that PKC-beta 2 is more sensitive to downregulation, and/or specific epidermal cell types that contain PKC-beta 2 are more sensitive to TPA/diacylglycerol. To address this issue, we isolated Langerhans cells (LCs) from epidermal cell suspensions with immunomagnetic beads and an antibody to the class II major histocompatibility complex. Northern blot analysis revealed a PKC-beta 2 signal in isolated LCs that was 40-fold greater than that observed in unfractionated epidermal cells, and no PKC-beta 2 signal was detected in epidermal cells depleted of LCs, indicating that PKC-beta 2 is expressed exclusively in LCs within the epidermis. Western blot analysis confirmed the presence of PKC-beta 2 in LCs. PKC-beta 2 was highly sensitive to downregulation, because a single application of TPA resulted in a 90% loss of PKC-beta 2 within 6 h without a decrease in the number of LCs. To determine whether the decreased level of PKC-beta 2 within LCs was associated with an alteration in contact hypersensitivity, we treated mice with only a single application of TPA, and 6 hours later mice were sensitized with 2,4-dinitrofluorobenzene on the same dorsal area. Subsequent challenge revealed a 60% decrease in contact hypersensitivity in TPA-treated mice. These data indicate that (i) within the epidermis, PKC-beta 2 is highly sensitive to downregulation and is exclusively expressed in LCs, and (ii) the downregulation of PKC-beta 2 is associated with impaired LC function with respect to contact hypersensitivity.
Goodell
A L
AL
Department of Toxicology, North Carolina State University, Raleigh 27695-7633, USA.
Oh
H S
HS
Meyer
S A
SA
Smart
R C
RC
D000818:Animals; D003877:Dermatitis, Contact / Q000378:metabolism* / Q000503:physiopathology; D004075:Diglycerides / Q000494:pharmacology; D015536:Down-Regulation; D004817:Epidermis / Q000201:enzymology*; D005260:Female; D007150:Immunohistochemistry; D007527:Isoenzymes / Q000378:metabolism; D007801:Langerhans Cells / Q000187:drug effects / Q000201:enzymology*; D051379:Mice; D008815:Mice, Inbred Strains; D011493:Protein Kinase C / Q000378:metabolism*; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D004075:Diglycerides; D007527:Isoenzymes; C044297:1,2-didecanoylglycerol; C496630:Prkca protein, mouse; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate