The molecular basis of X-linked recessive anhidrotic ectodermal dysplasia with immunodeficiency (EDA-ID) has remained elusive. Here we report hypomorphic mutations in the gene IKBKG in 12 males with EDA-ID from 8 kindreds, and 2 patients with a related and hitherto unrecognized syndrome of EDA-ID with osteopetrosis and lymphoedema (OL-EDA-ID). Mutations in the coding region of IKBKG are associated with EDA-ID, and stop codon mutations, with OL-EDA-ID. IKBKG encodes NEMO, the regulatory subunit of the IKK (IkappaB kinase) complex, which is essential for NF-kappaB signaling. Germline loss-of-function mutations in IKBKG are lethal in male fetuses. We show that IKBKG mutations causing OL-EDA-ID and EDA-ID impair but do not abolish NF-kappaB signaling. We also show that the ectodysplasin receptor, DL, triggers NF-kappaB through the NEMO protein, indicating that EDA results from impaired NF-kappaB signaling. Finally, we show that abnormal immunity in OL-EDA-ID patients results from impaired cell responses to lipopolysaccharide, interleukin (IL)-1beta, IL-18, TNFalpha and CD154. We thus report for the first time that impaired but not abolished NF-kappaB signaling in humans results in two related syndromes that associate specific developmental and immunological defects.

Mouse models of human disease can be generated by homologous recombination for germline loss-of-function mutations. However, embryonic-lethal phenotypes and systemic, indirect dysfunction can confound the use of knock-outs to elucidate adult pathophysiology. Site-specific recombination using Cre recombinase can circumvent these pitfalls, in principle, enabling temporal and spatial control of gene recombination. However, direct evidence is lacking for the feasibility of Cre-mediated recombination in postmitotic cells. Here, we exploited transgenic mouse technology plus adenoviral gene transfer to achieve Cre-mediated recombination in cardiac muscle. In vitro, Cre driven by cardiac-specific ␣ -myosin heavy chain ( ␣ MyHC) sequences elicited recombination selectively at loxP sites in purified cardiac myocytes, but not cardiac fibroblasts.

SUMMARY Wnt/β-catenin and NF-κB signaling mechanisms provide central controls in development and disease, but how these pathways intersect is unclear. Using hair follicle induction as a model system, we show that patterning of dermal Wnt/β-catenin signaling requires epithelial β-catenin activity. We find that Wnt/β-catenin signaling is absolutely required for NF-κB activation, and that Edar is a direct Wnt target gene. Wnt/β-catenin signaling is initially activated independently of Eda/Edar/NF-κB activity in primary hair follicle primordia. However, Eda/Edar/NF-κB signaling is required to refine the pattern of Wnt/β-catenin activity, and to maintain this activity at later stages of placode development. We show that maintenance of localized expression of Wnt10b and Wnt10a requires NF-κB signaling, providing a molecular explanation for the latter observation, and identify Wnt10b as a direct NF-κB target. These data reveal a complex interplay and inter-dependence of Wnt/β-catenin and Eda/Edar/NF-κB signaling pathways in initiation and maintenance of primary hair follicle placodes.

SUMMARY Several lines of evidence suggest that mitochondrial dysfunction plays a critical role in the pathogenesis of microvascular complications of diabetes, including diabetic nephropathy. However, the signaling pathways by which hyperglycemia leads to mitochondrial dysfunction are not fully understood. Here we examined the role of Rho-associated coiled-coil containing protein kinase 1 (ROCK1) on mitochondrial dynamics by generating two diabetic mouse models with targeted deletions of ROCK1, and an inducible podocyte-specific knock-in mouse expressing a constitutively active (cA) mutant of ROCK1. Our findings suggest that ROCK1 mediates hyperglycemia-induced mitochondrial fission by promoting dynamin-related protein-1 (Drp1) recruitment to the mitochondria. Deletion of ROCK1 in diabetic mice prevented mitochondrial fission, whereas podocyte-specific cA-ROCK1 mice exhibited increased mitochondrial fission. Importantly, we found that ROCK1 triggers mitochondrial fission by phosphorylating Drp1 at Serine 600 residue. These findings provide insights into the unexpected role of ROCK1 in a signaling cascade that regulates mitochondrial dynamics.

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and collect data. BHC helped generate Tug1-transgenic mice and edit the manuscript. PAO helped edit the manuscript, and FRD oversaw experiments, prepared the manuscript, and provided guidance on overall project design.

Vertebrates have characteristic and conserved left-right (L-R) visceral asymmetries, for example the left-sided heart. In humans, alterations of L-R development can have serious clinical implications, including cardiac defects. Although little is known about how the embryonic L-R axis is established, a recent study in the chick embryo revealed L-R asymmetric expression of several previously cloned genes, including Cnr-1 (for chicken nodal-related-1), and indicated how this L-R molecular asymmetry might be important for subsequent visceral morphogenesis. Here we show that nodal is asymmetrically expressed in mice at similar stages, as is Xnr-1 (for Xenopus nodal related-1) in frogs. We also examine nodal expression in two mouse mutations that perturb L-R development, namely situs inversus viscerum (iv), in which assignment of L-R asymmetry is apparently random and individuals develop either normally or are mirror-image-reversed (situs inversus), and inversion of embryonic turning (inv), in which all individuals develop with situs inversus. In both, nodal expression is strikingly affected, being reversed or converted to symmetry. These results further support a key role for nodal and nodal-related genes in interpreting and relaying L-R patterning information in vertebrates. To our knowledge, our results provide the first direct evidence that iv and inv normally function well before the appearance of morphological L-R asymmetry.

Although inductive interactions are known to be essential for specification of cell fate in many vertebrate tissues, the signals and receptors responsible for transmitting this information remain largely unidentified. Mice with mutations in the downless (dl) gene have defects in hair follicle induction, lack sweat glands and have malformed teeth. These structures originate as ectodermal placodes, which invaginate into the underlying mesenchyme and differentiate to form specific organs. Positional cloning of the dl gene began with identification of the transgenic family OVE1. One branch of the family, dl(OVE1B), carries an approximately 600-kb deletion at the dl locus caused by transgene integration. The mutated locus has been physically mapped in this family, and a 200-kb mouse YAC clone, YAC D9, has been identified and shown to rescue the dl phenotype in the spontaneous dl(Jackson) (dl(J), recessive) and Dl(sleek) (Dl(slk), dominant negative) mutants. Here we report the positional cloning of the dl gene, which encodes a novel member of the tumour necrosis factor (Tnf) receptor (Tnfr) family. The mutant phenotype and dl expression pattern suggests that this gene encodes a receptor that specifies hair follicle fate. Its ligand is likely to be the product of the tabby (Ta) gene, as Ta mutants have a phenotype identical to that of dl mutants and Ta encodes a Tnf-like protein.