Interactive Fly, Drosophila

Mammalian Forkhead homologs: Wing-helix nude (Hfh 11) is a product of the nude locus

Mutations at the nude locus of mice and rats disrupt normal hair growth and thymus development, causing nude mice and rats to be immune-deficient. The mouse nude locus has been localized on chromosome 11 within a region of < 1 megabase. One of the genes from this critical region, designated whn, encodes a new member of the winged-helix domain family of transcription factors, and it is disrupted on mouse nu and rat rnuN alleles. Mutant transcripts do not encode the characteristic DNA-binding domain, strongly suggesting that the whn gene is the nude gene. Mutations in winged-helix domain genes cause homeotic transformations in Drosophila and distort cell-fate decisions during vulval development in Caenorhabditis elegans. The whn gene is thus the first member of this class of genes to be implicated in a specific developmental defect in vertebrates (Nehls, 1994).

The development of the thymus depends initially on epithelial-mesenchymal and subsequently on reciprocal lympho-stromal interactions. The genetic steps governing development and differentiation of the thymic microenvironment are unknown. By a targeted disruption of the whn gene, which recapitulates the phenotype of the athymic nude mouse, the Whn transcription factor has been shown to be the product of the nude locus. Formation of the thymic epithelial primordium before the entry of lymphocyte progenitors does not require the activity of Whn. However, subsequent differentiation of primitive precursor cells into subcapsular, cortical, and medullary epithelial cells of the postnatal thymus depends on activity of the whn gene. These results define the first genetically separable steps during thymic epithelial differentiation (Nehls, 1996).

Mutations in the winged-helix nude (whn) gene are associated with the phenotype of congenital athymia and hairlessness in mouse and rat. The whn gene encodes a presumptive transcription factor with a DNA binding domain of the forkhead/winged-helix class. Two previously described null alleles encode truncated whn proteins lacking the characteristic DNA binding domain. In the rat rnu allele described here, a nonsense mutation in exon 8 of the whn gene was identified. The truncated whnrnu protein contains the DNA binding domain but lacks the 175 C-terminal amino acids of the wild-type protein. To facilitate the identification of functionally important regions in this region, a whn homolog from the pufferfish Fugu rubripes was isolated. Comparison of derived protein sequences with the mouse whn gene reveals the presence of a conserved acidic protein domain in the C terminus, in addition to the highly conserved DNA binding domain. Using fusions with a heterologous DNA binding domain, a strong transcriptional activation domain was localized to the C-terminal cluster of acidic amino acids. Since the whnrnu mutant protein lacks this domain, these results indicate that a transactivation function is essential for the activity of the whn transcription factor (Schuddekopf, 1996).

Mutations in the winged-helix nude (whn) gene result in the nude mouse and rat phenotypes. The pleiotropic nude phenotype, which affects the hair, skin, and thymus, suggests that whn plays a pivotal role in the development and/or maintenance of these organs. However, little is known about whn function in these organs. In skin whn is specifically expressed in epithelial cells and not the mesenchymal cells: using a hair reconstitution assay, it has been demonstrated that the abnormal nude mouse hair development is attributable to a functional defect of the epithelial cells. Examination of nude mouse primary keratinocytes in culture reveals that these cells have an increased propensity to differentiate in an abnormal fashion, even under conditions that promote proliferation. Furthermore, nude mouse keratinocytes display a 100-fold increased sensitivity to the growth-inhibitory/differentiation effects of the phorbol ester TPA. In parallel with these findings, it has been directly shown that Whn functions as a transcription factor that can specifically suppress expression of differentiation/TPA-responsive genes. The region of Whn responsible for these effects was mapped to the carboxy-terminal transactivating domain. These results establish whn as a key regulatory factor involved in maintaining the balance between keratinocyte growth and differentiation. The general implications of these findings for an epithelial self-renewal model are discussed (Brissette, 1996).

In the mouse, the product of the nude locus, Whn, is required for the keratinization of the hair shaft and the differentiation of epithelial progenitor cells in the thymus. A bacterially expressed peptide representing the presumptive DNA binding domain of the mouse whn gene in vitro specifically binds to an 11-bp consensus sequence containing the invariant tetranucleotide 5'-ACGC. In transient transfection assays, such binding sites stimulate reporter gene expression about 30- to 40-fold, when positioned upstream of a minimal promotor. Whn homologs from humans, bony fish (Danio rerio), cartilaginous fish (Scyliorhinus caniculus), agnathans (Lampetra planeri), and cephalochordates (Branchiostoma lanceolatum) share at least 80% of amino acids in the DNA binding domain. In agreement with this remarkable structural conservation, the DNA binding domains from zebrafish, which possesses a thymus but no hair, and amphioxus, which possesses neither thymus nor hair, recognize the same target sequence as the mouse DNA binding domain in vitro and in vivo. The genomes of vertebrates and cephalochordates contain only a single whn-like gene, suggesting that the primordial whn gene was not subject to gene-duplication events. Although the role of whn in cephalochordates and agnathans is unknown, its requirement in the development of the thymus gland and the differentiation of skin appendages in the mouse suggests that changes in the transcriptional control regions of whn genes accompanied their functional reassignments during evolution (Schlake, 1997).

Loss-of-function mutations in Whn (wing-helix nude; Hfh 11), a winged-helix/forkhead transcription factor, result in the nude mouse phenotype. To determine the whn expression pattern during development, mice were used in which a beta-galactosidase reporter gene was placed under the control of the wild-type whn promoter by homologous recombination. Sites of reporter expression were confirmed by immunohistochemical staining for Whn protein or by in situ hybridization for whn mRNA. At all developmental stages, whn expression is restricted to epithelial cells. In addition to the skin and thymus, whn is expressed in the developing nails, nasal passages, tongue, palate, and teeth. In embryonic epidermis, suprabasal cells induce whn expression at the same time that terminal differentiation markers first appear. As the epidermis matures, whn promoter activity is found primarily in the first suprabasal layer, which contains keratinocytes in the early stages of terminal differentiation. In developing and mature anagen hair follicles, whn is expressed at high levels in the postmitotic precursor cells of the hair shaft and inner root sheath. Though principally associated with terminal differentiation, whn expression is also detected in progenitor cell compartments; in the hair bulb matrix and basal epidermal layer, a small subclass of cells expresses whn, while in the outer root sheath, whn promoter activity is induced as the follicle completes its elongation. Within these compartments, rare cells exhibit both whn expression and the nuclear proliferation marker Ki-67. The results suggest that whn expression encompasses the transition from a proliferative to a postmitotic state and that whn regulates the initiation of terminal differentiation. During thymus development, whn expression first appears in epithelial cells of the thymic primordium, and in the mature thymus, whn expressing epithelial cells are present throughout the medulla, cortex, and subcapsular region. Given the sites of whn expression in the mouse, as well as the presence of homologs in lower vertebrates and cephalochordates, the whn gene may influence a fundamental or common features of epithelial cell differentiation (Lee, 1999).

Table of contents

The Interactive Fly resides on the
Society for Developmental Biology's Web server.