A novel translation re-initiation mechanism for the p63 gene revealed by amino-terminal truncating mutations in Rapp-Hodgkin/Hay-Wells-like syndromes.

Missense mutations in the 3' end of the p63 gene are associated with either RHS (Rapp-Hodgkin syndrome) or AEC (Ankyloblepharon Ectodermal defects Cleft lip/palate) syndrome. These mutations give rise to mutant p63alpha protein isoforms with dominant effects towards their wild-type counterparts. Here we report four RHS/AEC-like patients with mutations (p.Gln9fsX23, p.Gln11X, p.Gln16X), that introduce premature termination codons in the N-terminal part of the p63 protein. These mutations appear to be incompatible with the current paradigms of dominant-negative/gain-of-function outcomes for other p63 mutations. Moreover it is difficult to envisage how the remaining small N-terminal polypeptide contributes to a dominant disease mechanism. Primary keratinocytes from a patient containing the p.Gln11X mutation revealed a normal and aberrant p63-related protein that was just slightly smaller than the wild-type p63. We show that the smaller p63 protein is produced by translation re-initiation at the next downstream methionine, causing truncation of a non-canonical transactivation domain in the DeltaN-specific isoforms. Interestingly, this new DeltaDeltaNp63 isoform is also present in the wild-type keratinocytes albeit in small amounts compared with the p.Gln11X patient. These data establish that the p.Gln11X-mutation does not represent a null-allele leading to haploinsufficiency, but instead gives rise to a truncated DeltaNp63 protein with dominant effects. Given the nature of other RHS/AEC-like syndrome mutations, we conclude that these mutations affect only the DeltaNp63alpha isoform and that this disruption is fundamental to explaining the clinical characteristics of these particular ectodermal dysplasia syndromes.

Loss-of-function FERMT1 mutations in kindler syndrome implicate a role for fermitin family homolog-1 in integrin activation.

Kindler syndrome is an autosomal recessive disorder characterized by skin atrophy and blistering. It results from loss-of-function mutations in the FERMT1 gene encoding the focal adhesion protein, fermitin family homolog-1. How and why deficiency of fermitin family homolog-1 results in skin atrophy and blistering are unclear. In this study, we investigated the epidermal basement membrane and keratinocyte biology abnormalities in Kindler syndrome. We identified altered distribution of several basement membrane proteins, including types IV, VII, and XVII collagens and laminin-332 in Kindler syndrome skin. In addition, reduced immunolabeling intensity of epidermal cell markers such as beta1 and alpha6 integrins and cytokeratin 15 was noted. At the cellular level, there was loss of beta4 integrin immunolocalization and random distribution of laminin-332 in Kindler syndrome keratinocytes. Of note, active beta1 integrin was reduced but overexpression of fermitin family homolog-1 restored integrin activation and partially rescued the Kindler syndrome cellular phenotype. This study provides evidence that fermitin family homolog-1 is implicated in integrin activation and demonstrates that lack of this protein leads to pathological changes beyond focal adhesions, with disruption of several hemidesmosomal components and reduced expression of keratinocyte stem cell markers. These findings collectively provide novel data on the role of fermitin family homolog-1 in skin and further insight into the pathophysiology of Kindler syndrome.

The molecular skin pathology of familial primary localized cutaneous amyloidosis.

Familial primary localized cutaneous amyloidosis (FPLCA) is an autosomal dominant disorder associated with chronic itching and skin lichenification. In lesional skin, there are apoptotic basal keratinocytes and deposits of amyloid material on degenerate keratin filaments in the upper dermis. The genetic basis of FPLCA involves mutations in the OSMR and IL31RA genes but the disease pathophysiology is not fully understood. In this study, we identified new pathogenic heterozygous missense mutations in the OSMR gene (p.Val631Leu and p.Asp647Tyr) in two Dutch FPLCA families. We then compared gene expression profiles between FPLCA lesional skin (n = 4) and site-matched control skin (n = 6). There was twofold or greater upregulation of 34 genes and downregulation of 43 genes. Most changes in gene expression (verified by quantitative RT-PCR) reflected alterations in epidermal differentiation and proliferation consistent with lichenification, but we also noted a reduction in several interfollicular keratinocyte stem cell markers in FPLCA skin. Differences in gene expression were also noted for proteins involved in apoptosis and nerve conduction. Collectively, this study expands the molecular basis of FPLCA and provides new insight into the skin pathology of this condition.

Classifying ectodermal dysplasias: Incorporating the molecular basis and pathways (Workshop II).

Hereditary conditions are traditionally classified based either on physical/physiological attributes or using the names of the individuals credited with identifying the condition. For the 170 plus conditions classified as ectodermal dysplasias (EDs), both of these nosological systems are used, at times interchangeably. Over the past decade our knowledge of the human genome and the molecular basis of the EDs have greatly expanded providing the impetus to consider alternative classification systems. The incorporation of the molecular basis of hereditary conditions adds important information allowing effective transfer of objective genetic information that can be lacking from traditional classification systems. Molecular information can be added to the nosological system for the EDs through a hierarchical- and domain-based approach that encompasses the condition's name, mode of inheritance, molecular pathway affected, and specific molecular change. As new molecular information becomes available it can be effectively incorporated using this classification approach. Integrating molecular information into the ED classification system, while retaining well-recognized traditional syndrome names, facilitates communication at and between different groups of people including patients, families, health care providers, and researchers.

International Research Symposium on Ankyloblepharon-Ectodermal Defects-Cleft Lip/Palate (AEC) syndrome.

Ankyloblepharon-ectodermal defects-cleft lip/palate (AEC) syndrome (Hay-Wells syndrome, MIM #106220) is a rare autosomal dominant ectodermal dysplasia syndrome. It is due to mutations in the TP63 gene, known to be a regulatory gene with many downstream gene targets. TP63 is important in the differentiation and proliferation of the epidermis, as well as many other processes including limb and facial development. It is also known that mutations in TP63 lead to skin erosions. These erosions, especially on the scalp, are defining features of AEC syndrome and cause significant morbidity and mortality in these patients. It was this fact that led to the 2003 AEC Skin Erosion Workshop. That conference laid the groundwork for the International Research Symposium for AEC Syndrome held at Texas Children's Hospital in 2006. The conference brought together the largest cohort of individuals with AEC syndrome, along with a multitude of physicians and scientists. The overarching goals were to define the clinical and pathologic findings for improved diagnostic criteria, to obtain tissue samples for further study and to define future research directions. The symposium was successful in accomplishing these aims as detailed in this conference report. Following our report, we also present 11 manuscripts within this special section that outline the collective clinical, pathologic, and mutational data from 18 individuals enrolled in the concurrent Baylor College of Medicine IRB-approved protocol: Characterization of AEC syndrome. These collaborative findings will hopefully provide a stepping-stone to future translational projects of TP63 and TP63-related syndromes.

Focal dermal hypoplasia resulting from a new nonsense mutation, p.E300X, in the PORCN gene.

BACKGROUND: Focal dermal hypoplasia (FDH) (OMIM 305600) is an X-linked dominant disorder of ecto-mesodermal development. Also known as Goltz syndrome, FDH presents with characteristic linear streaks of hypoplastic dermis and variable abnormalities of bone, nails, hair, limbs, teeth and eyes. The molecular basis of FDH involves mutations in the PORCN gene, which encodes an enzyme that allows membrane targeting and secretion of several Wnt proteins critical for normal tissue development. OBJECTIVES: To investigate the molecular basis of FDH in a 2-year-old Thai girl who presented at birth with depressed, pale linear scars on the trunk and limbs, sparse brittle hair, syndactyly of the right middle and ring fingers, dental caries and radiological features of osteopathia striata. METHODS: Sequencing of genomic DNA from the affected individual and both parents to search for pathogenic mutations in PORCN gene. RESULTS: DNA sequencing disclosed a heterozygous G>T substitution at nucleotide c.898 within exon 10 (NM_203475.1), converting a glutamic acid residue (GAA) to a premature termination codon (TAA). This mutation, designated p.E300X, was not detected in DNA from either parent or in 100 control chromosomes. CONCLUSION: Identification of this new de novo nonsense mutation confirms the diagnosis of FDH in this child and highlights the clinical importance of PORCN and Wnt signalling pathways in embryogenesis.