P4.41 INHIBITION OF HAIR PLACODE FORMATION IN MICE WITH SUPPRESSED NF-kB ACTIVITY: THE MOLECULAR MECHANISMS

Ruth Schmidt-Ullrich¹, Desmond Tobin², Joerg Huelsken³, Gundula Pilnitz-Stolze⁴, Walter Birchmeier¹, Ralf Paus⁴ & Claus Scheidereit¹

¹, Max-Delbrück-Center, Robert-Rössle-Str. 10, 13092 Berlin, Germany ^2, University of Biomedical Sciences, University of Bradford, Bradford, West Yorkshire BD7 1DP, England ³, ISREC, Chemin des Boveresses 155, 1066 Epalinges s/Lausanne, Switzerland ⁴, Hautklinik UKE, Universität Hamburg, Martinistr. 52, 20246 Hamburg, Germany

Objectives: The transcription factor family NF-kB /Rel plays an important role in many cellular events including cell cycle and cell death, as well as in processes of the immune response. NF-kB /Rel activity is regulated by a specific group of inhibitors, called IkB (Inhibitor of kB), which in most cell types of the body sequester NF-kB /Rel molecules in the cytoplasm. Upon specific extracellular stimuli, these inhibitors are phosphorylated by the IkB kinase complex (IKK). Subsequent degradation of IkB leads to the release of NF-kB /Rel molecules, and translocation to the nucleus. We investigated the role of NF-kB during murine hair follicle development, and the cross-talk between the EDA/EDAR/ NF-kB and the Wnt/β-catenin signaling pathway at initial time points of hair follicle formation in mice.

Methodes: We generated mice expressing a transdominant negative mutant of the NF-kB inhibitor IkBa, to block NF-kB activation. IkBaDN cDNA was integrated in frame into the ubiquitously expressed b-catenin gene. Additional mice, and offspring of matings between them, which used in this study were downless and tabby mice, reporter mice expressing an NF-kB responsive β-galactosidase transgene, and mice expressing the β-galactosidase gene in frame in the β-catenin locus.

Results: Knock-in IkBaDN mice revealed defects in the early development of most hair follicle types, exocrine glands and teeth identical to those of mice with mutations in the tabby (ta; eda) and downless (dl; edar) locus, and present, thus, an analogous phenotype in humans with HED (Hypohydrotic Ectodermal Dysplasia). To confirm NF-kB activity in these structures we analyzed mice expressing an NF-kB-responsive b-galactosidase construct ((Igk)₃conalacZ) which showed strong NF-kB activity in these structures as early as E14 for pellage hair and E12 for vibrissae. With the exception of b-catenin, EDA (Ectodysplasin A) and EDAR (Ectodysplasin receptor), expression of important regulators of hair follicle development are absent in guard hairs of IkBaDN mice, but not in the secondary awl hairs.

Conclusions: We demonstrate that NF-kB is needed for ectodermal placode formation of guard hairs at the initial stage, and that it is directly activated downstream by EDA interacting with its receptor EDAR. EDA and EDAR are related to the TNF multigene family of ligands and receptors, respectively, which are known to activate NF-kB. The first mesodermal or ectodermal signal Wnt/β-catenin leads to localization of EDAR to the future placode site, and subsequent binding of EDA leads to activation of NF-kB. Direct or indirect targets of NF-kB activity such as Wnt10b, 10a, Shh, BMP’S Noggin and others lead to the proliferation of the hair placode and later to morphogenesis of the hair follicle. Furthermore, we can show that NF-kB is important for the survival and proliferation of guard hair placode cells.