Mutations in POGLUT1, encoding protein O-glucosyltransferase 1, cause autosomal-dominant Dowling-Degos disease.

Dowling-Degos disease (DDD) is an autosomal-dominant genodermatosis characterized by progressive and disfiguring reticulate hyperpigmentation. We previously identified loss-of-function mutations in KRT5 but were only able to detect pathogenic mutations in fewer than half of our subjects. To identify additional causes of DDD, we performed exome sequencing in five unrelated affected individuals without mutations in KRT5. Data analysis identified three heterozygous mutations from these individuals, all within the same gene. These mutations, namely c.11G>A (p.Trp4*), c.652C>T (p.Arg218*), and c.798-2A>C, are within POGLUT1, which encodes protein O-glucosyltransferase 1. Further screening of unexplained cases for POGLUT1 identified six additional mutations, as well as two of the above described mutations. Immunohistochemistry of skin biopsies of affected individuals with POGLUT1 mutations showed significantly weaker POGLUT1 staining in comparison to healthy controls with strong localization of POGLUT1 in the upper parts of the epidermis. Immunoblot analysis revealed that translation of either wild-type (WT) POGLUT1 or of the protein carrying the p.Arg279Trp substitution led to the expected size of about 50 kDa, whereas the c.652C>T (p.Arg218*) mutation led to translation of a truncated protein of about 30 kDa. Immunofluorescence analysis identified a colocalization of the WT protein with the endoplasmic reticulum and a notable aggregating pattern for the truncated protein. Recently, mutations in POFUT1, which encodes protein O-fucosyltransferase 1, were also reported to be responsible for DDD. Interestingly, both POGLUT1 and POFUT1 are essential regulators of Notch activity. Our results furthermore emphasize the important role of the Notch pathway in pigmentation and keratinocyte morphology.

Genomewide analysis of copy number variants in alopecia areata in a Central European cohort reveals association with MCHR2.

Alopecia areata (AA) is a common hair loss disorder of autoimmune aetiology, which often results in pronounced psychological distress. Understanding of the pathophysiology of AA is increasing, due in part to recent genetic findings implicating common variants at several genetic loci. To date, no study has investigated the contribution of copy number variants (CNVs) to AA, a prominent class of genomic variants involved in other autoimmune disorders. Here, we report a genomewide- and a candidate gene-focused CNV analysis performed in a cohort of 585 patients with AA and 1340 controls of Central European origin. A nominally significant association with AA was found for CNVs in the following five chromosomal regions: 4q35.2, 6q16.3, 9p23, 16p12.1 and 20p12.1. The most promising finding was a 342.5-kb associated region in 6q16.3 (duplications in 4/585 patients; 0/1340 controls). The duplications spanned the genes MCHR2 and MCHR2-AS1, implicated in melanin-concentrating hormone (MCH) signalling. These genes have not been implicated in previous studies of AA pathogenesis. However, previous research has shown that MCHR2 affects the scale colour of barfin flounder fish via the induction of melanin aggregation. AA preferentially affects pigmented hairs, and the hair of patients with AA frequently shows a change in colour when it regrows following an acute episode of AA. This might indicate a relationship between AA, pigmentation and MCH signalling. In conclusion, the present results provide suggestive evidence for the involvement of duplications in MCHR2 in AA pathogenesis.

Mutations in SNRPE, which encodes a core protein of the spliceosome, cause autosomal-dominant hypotrichosis simplex.

Hypotrichosis simplex (HS) comprises a group of hereditary isolated alopecias that are characterized by a diffuse and progressive loss of hair starting in childhood and shows a wide phenotypic variability. We mapped an autosomal-dominant form of HS to chromosome 1q31.3-1q41 in a Spanish family. By direct sequencing, we identified the heterozygous mutation c.1A>G (p.Met1?) in SNRPE that results in loss of the start codon of the transcript. We identified the same mutation in a simplex HS case from the UK and an additional mutation (c.133G>A [p.Gly45Ser]) in a simplex HS case originating from Tunisia. SNRPE encodes a core protein of U snRNPs, the key factors of the pre-mRNA processing spliceosome. The missense mutation c.133G>A leads to a glycine to serine substitution and is predicted to disrupt the structure of SNRPE. Western blot analyses of HEK293T cells expressing SNRPE c.1A>G revealed an N-terminally truncated protein, and therefore the mutation might result in use of an alternative in-frame downstream start codon. Subcellular localization of mutant SNRPE by immunofluorescence analyses as well as incorporation of mutant SNRPE proteins into U snRNPs was found to be normal, suggesting that the function of U snRNPs in splicing, rather than their biogenesis, is affected. In this report we link a core component of the spliceosome to hair loss, thus adding another specific factor in the complexity of hair growth. Furthermore, our findings extend the range of human phenotypes that are linked to the splicing machinery.

Hypotrichosis simplex of the scalp is associated with nonsense mutations in CDSN encoding corneodesmosin.

We have identified nonsense mutations in the gene CDSN (encoding corneodesmosin) in three families suffering from hypotrichosis simplex of the scalp (HSS; OMIM 146520). CDSN, a glycoprotein expressed in the epidermis and inner root sheath (IRS) of hair follicles, is a keratinocyte adhesion molecule. Truncated CDSN aggregates were detected in the superficial dermis and at the periphery of hair follicles. Our findings suggest that CDSN is important in normal scalp hair physiology.

Selected variants of the steroid-5-alpha-reductase isoforms SRD5A1 and SRD5A2 and the sex steroid hormone receptors ESR1, ESR2 and PGR: no association with female pattern hair loss identified.

Female pattern hair loss (FPHL) is a common disorder with a complex mode of inheritance. Although understanding of its etiopathogenesis is incomplete, an interaction between genetic and hormonal factors is assumed to be important. The involvement of an androgen-dependent pathway and sex steroid hormones is the most likely hypothesis. We therefore selected a total of 21 variants from the steroid-5-alpha-reductase isoforms SRD5A1 and SRD5A2, the sex steroid hormone receptors ESR1, ESR2 (oestrogen receptor) and PGR (progesterone receptor) and genotyped these in a case-control sample of 198 patients (145 UK; 53 German patients) and 329 controls (179 UK; 150 German). None of these variants showed any significant association, either in the overall UK and German samples or in the subgroup analyses. In summary, the present results, while based on a limited selection of gene variants, do not point to the involvement of SRD5A1, SRD5A2, ESR1, ESR2 or PGR in FPHL.

Genome-wide scan and fine-mapping linkage study of androgenetic alopecia reveals a locus on chromosome 3q26.

Androgenetic alopecia (AGA, male pattern baldness) is the most common form of hair loss. The origin of AGA is genetic, with the X chromosome located androgen receptor gene (AR) being the only risk gene identified to date. We present the results of a genome-wide linkage study of 95 families and linkage fine mapping of the 3q21-q29, 11q14-q25, 18p11-q23, and 19p13-q13 regions in an extended sample of 125 families of German descent. The locus with strongest evidence for linkage was mapped to 3q26 with a nonparametric linkage (NPL) score of 3.97 (empirical p value = 0.00055). This is the first step toward the identification of new susceptibility genes in AGA, a process which will provide important insights into the molecular and cellular basis of scalp hair loss.

Marie Unna hereditary hypotrichosis: identification of a U2HR mutation in the family from the original 1925 report.

BACKGROUND: In 1925, Dr Marie Unna described a rare form of hereditary hypotrichosis in a German multigenerational family. This was later termed "Marie Unna hereditary hypotrichosis" (MUHH). MUHH is an autosomal dominant disorder that is characterized by the absence or scarcity of scalp hair, eyebrows, and eyelashes at birth; coarse and wiry hair during childhood; and progressive hair loss beginning around puberty. Causal mutations in U2HR, an inhibitory upstream open reading frame in the 5'-untranslated region of the human hairless (HR) gene, were recently identified in several unrelated MUHH families from various ethnic backgrounds. OBJECTIVE: Although there have been several clinical reports of descendants of the originally described family, the molecular cause of disease in this particular family has not been established. The aim of this study was to investigate descendants of this family and to analyze their DNA for the presence of U2HR mutations. METHODS: Descendants of the family (including one affected individual) were examined clinically. Direct sequencing of U2HR was performed. Enzymatic digestion using the restriction enzyme NcoI was performed to confirm the sequencing results. RESULTS: The index patient displayed the typical MUHH pattern of hair loss and was found to carry the disease-causing c.3G>A (p.M1I) U2HR mutation. This mutation was not detected in unaffected family members. LIMITATIONS: Only one affected family member was investigated. CONCLUSIONS: Eighty-five years after the first description of this rare form of alopecia, the disease-causing mutation in the originally reported family has been identified.

Waardenburg syndrome type I with heterochromia iridis and circumscribed hypopigmentation of the skin.

We report a 3-year-old girl with autosomal dominant inherited Waardenburg syndrome type I showing circumscribed hypopigmentation of the skin, heterochromia iridis, sensorineural deafness, and dental aberrations. Clinical diagnosis was confirmed by the identification of an underlying missense mutation (C811T) in the PAX3 gene. Early diagnosis of Waardenburg syndrome among children with pigment anomalies enables a successful interdisciplinary medical care.

Richner-Hanhart syndrome detected by expanded newborn screening.

Richner-Hanhart syndrome (tyrosinemia type 2) is an inborn error of tyrosine metabolism which is clinically characterized mainly by oculocutaneous symptoms including corneal opacities and keratosis palmoplantaris. Skin symptoms usually develop after the first year of life. We report a neonate in whom already on the third day of life diagnosis of Richner-Hanhart syndrome could be suspected because of elevated tyrosine levels in newborn screening by tandem mass spectrometry. Analysis of the tyrosine aminotransferase gene revealed a homozygous missense mutation p.R433W (c.1297C>T). An 8-year-old brother with persistent plantar hyperkeratotic plaques of the soles of yet unknown origin was subsequently identified to be also affected with Richner-Hanhart syndrome. This demonstrates that early diagnosis of Richner-Hanhart syndrome is possible in neonates by extended newborn screening. Early introduction of dietary treatment is a prerequisite to reduce the risk of clinical symptoms.

Genome-Wide MicroRNA Analysis Implicates miR-30b/d in the Etiology of Alopecia Areata.

Alopecia areata (AA) is one of the most common forms of human hair loss. Although genetic studies have implicated autoimmune processes in AA etiology, understanding of the etiopathogenesis is incomplete. Recent research has implicated microRNAs, a class of small noncoding RNAs, in diverse autoimmune diseases. To our knowledge, no study has investigated the role of microRNAs in AA. In this study, gene-based analyses were performed for microRNAs using data of the largest genome-wide association meta-analysis of AA to date. Nominally, significant P-values were obtained for 78 of the 617 investigated microRNAs. After correction for multiple testing, three of the 78 microRNAs remained significant. Of these, miR-30b/d was the most significant microRNA for the follow-up analyses, which also showed lower expression in the hair follicle of AA patients. Target gene analyses for the three microRNAs showed 42 significantly associated target genes. These included IL2RA, TNXB, and ERBB3, which had been identified as susceptibility loci in previous genome-wide association studies. Using luciferase assay, site-specific miR-30b regulation of the AA risk genes IL2RA, STX17, and TNXB was validated. This study implicates microRNAs in the pathogenesis of AA. This finding may facilitate the development of future treatment strategies.

Identification of a keratin-associated protein with a putative role in vesicle transport.

Protection of skin against UV light requires a coordinated interaction between melanocytes and keratinocytes. Melanosomes are lysosome-related organelles that originate in melanocytes and are transferred into keratinocytes where they form a supranuclear cap. The mechanism responsible for melanosome transfer into keratinocytes and their intracellular distribution is poorly understood. Recently, we reported for the first time that loss-of-function mutations in the keratin K5 gene affect melanosome distribution in keratinocytes and results in a reticulate hyperpigmentation disorder, called Dowling-Degos disease. Here, we characterise the distribution and behaviour of individual K5 and K14 domains following transient and stable transfection into cells. We report that the K5 head domain is considerably more stable than the K14 head. Moreover, the distribution of the K5 head domain is altered following depolymerisation of microtubules. Following co-immunoprecipitation, we verified a specific interaction between the head domain of K5 with Hsc70, a chaperone also involved in vesicle uncoating. We hypothesise that this interaction is involved in melanosome formation or transport in keratinocytes. Alternatively, it may have a general function in the regulation of keratin assembly.

Familial aggregation of alopecia areata.

BACKGROUND: Familial aggregation of alopecia areata (AA) has been previously described, but systematic studies with information obtained directly from family members have yet to be undertaken. OBJECTIVE: We sought to study the pattern of familial aggregation of AA by assessing the affection status of patients' relatives. The study included 206 index patients with a total of 1029 first-degree and 2625 second-degree relatives. METHODS: First-degree relatives were directly interviewed, whereas information on second-degree relatives was obtained by interviewing the index patients and their first-degree relatives. RESULTS: Estimated lifetime risks were 7.1% in siblings, 7.8% in parents, and 5.7% in offspring. The risk in second-degree relatives was slightly higher than the reported population risk. Age at onset in index patients and first-degree relatives was significantly correlated. LIMITATIONS: Using patients drawn from specialized hair clinics may have produced results showing a higher proportion of early onset and severe cases. CONCLUSION: The familial aggregation of AA supports the role of genetic factors in the development of the disease. In addition, our data indicate genetic factors might contribute to the age at onset of AA.

Investigation of the HLA-DRB1 locus in alopecia areata.

To further evaluate the nature of the HLA association with alopecia areata (AA), we investigated the HLA-DRB1 locus in 161 AA patients and 165 matched controls from Belgium and Germany. HLA-DRB1 typing was performed using a recently established method that employs a combination of PCR-SSP (sequence specific priming) and Pyrosequencing(TM) technology. No significant differences were observed for HLA allele groups DRB1 *01, *07, *08, *09, *10, *11, *13, *14, *15, and *16. HLA-DRB1*03 was found to confer a protective effect (7.5% versus 13.6%, p = 0.011). Additional genotyping at the allelic level revealed a significant difference in HLA-DRB1*0301 between patients and controls (6.8% versus 11.2%, p = 0.048). The DRB1*04 allele group was confirmed as a risk factor for the development of AA (20.8% versus 13.3%, p = 0.012), with the allele DRB1*0401 accounting for the greatest proportion of the effect (13.4% versus 7.3%, p = 0.014). Results obtained after subgrouping of the patients according to age at onset, severity and family history of the disease suggests that the genetic effects of the HLA system are strongest in familial cases of the disease.

DNA mismatch repair and the significance of a sebaceous skin tumor for visceral cancer prevention.

DNA mismatch repair is a postreplicative DNA repair cascade ensuring genomic integrity. Inactivating germline mutations in DNA mismatch repair genes are responsible for hereditary non-polyposis colorectal carcinoma syndrome (HNPCC), which predisposes to various types of visceral cancer. Most associated tumors exhibit high-grade microsatellite instability. Some patients develop skin tumors of the sebaceous glands. This combined occurrence is known as Muir-Torre syndrome, which has a high probability of an underlying DNA mismatch repair defect. This is also true for individuals selected solely on the basis of sebaceous neoplasias, tumors with the highest frequency of high-grade microsatellite instability. This article focuses on the recent advances in molecular diagnostics for the detection of DNA mismatch repair defects in patients with sebaceous neoplasias, and the potential significance for the secondary prevention of visceral cancer in these patients.

Genome-wide meta-analysis in alopecia areata resolves HLA associations and reveals two new susceptibility loci.

Alopecia areata (AA) is a prevalent autoimmune disease with 10 known susceptibility loci. Here we perform the first meta-analysis of research on AA by combining data from two genome-wide association studies (GWAS), and replication with supplemented ImmunoChip data for a total of 3,253 cases and 7,543 controls. The strongest region of association is the major histocompatibility complex, where we fine-map four independent effects, all implicating human leukocyte antigen-DR as a key aetiologic driver. Outside the major histocompatibility complex, we identify two novel loci that exceed the threshold of statistical significance, containing ACOXL/BCL2L11(BIM) (2q13); GARP (LRRC32) (11q13.5), as well as a third nominally significant region SH2B3(LNK)/ATXN2 (12q24.12). Candidate susceptibility gene expression analysis in these regions demonstrates expression in relevant immune cells and the hair follicle. We integrate our results with data from seven other autoimmune diseases and provide insight into the alignment of AA within these disorders. Our findings uncover new molecular pathways disrupted in AA, including autophagy/apoptosis, transforming growth factor beta/Tregs and JAK kinase signalling, and support the causal role of aberrant immune processes in AA.

Frequency of microsatellite instability in unselected sebaceous gland neoplasias and hyperplasias.

Sebaceous gland neoplasias are the cutaneous manifestation of the Muir-Torre syndrome, which is known to be a phenotypical variant of hereditary nonpolyposis colorectal cancer. Both hereditary nonpolyposis colorectal cancer and Muir-Torre syndrome are caused by inherited DNA mismatch repair defects. As a prominent molecular genetic feature, all tumors associated with a DNA mismatch repair defect exhibit high microsatellite instability. So far, the frequency of DNA mismatch repair defects in patients selected solely on the basis of a sebaceous gland tumor has never been determined. In order to estimate this frequency, we assessed microsatellite instability with up to 10 microsatellite markers in a newly collected unselected series of 25 sebaceous gland neoplasias (six sebaceous adenomas, 16 sebaceous epitheliomas, three sebaceous carcinomas) in comparison to 32 sebaceous gland hyperplasias from unrelated patients. As many as 15 of the 25 sebaceous gland neoplasias (60%), but only one of the 32 sebaceous gland hyperplasias (3%), exhibited high microsatellite instability. Thus, in our study, the majority of patients with a sebaceous gland neoplasia in contrast to patients with a sebaceous gland hyperplasia are highly suspicious for an inherited DNA mismatch repair defect. On the basis of the subsequently collected tumor histories, nine of the 15 patients with a high microsatellite unstable sebaceous gland neoplasia were identified to have Muir-Torre syndrome. In none of these cases, however, were the clinical Amsterdam criteria for diagnosing hereditary nonpolyposis colorectal cancer fulfilled. In the sebaceous tumors of the remaining six patients, high microsatellite instability was an incidental finding. In two of these six patients, single relatives were known to be affected with internal cancer; however, their family histories were not suggestive of Muir-Torre syndrome or hereditary nonpolyposis colorectal cancer. In comparison with microsatellite instability screening studies in a variety of other randomly selected tumors, our study identifies sebaceous gland neoplasias as tumors with the highest frequency of high microsatellite instability reported so far, whereas sebaceous gland hyperplasia rarely exhibits high microsatellite instability. Therefore, screening for microsatellite instability in sebaceous gland neoplasias will be of great value in the detection of an inherited DNA mismatch repair defect, which predisposes to various types of internal cancers.

Arylamine N-acetyltransferase type 2 and glutathione S-transferases M1 and T1 polymorphisms in familial adenomatous polyposis.

Familial adenomatous polyposis (FAP) exhibits a variable phenotype even in carriers of the same adenomatous polyposis coli germline mutation. Xenobiotic-metabolizing enzymes such as N-acetyltransferases (NATs) and glutathione S-transferases (GSTs) were reported to modify the individual risk for colorectal cancer. We examined whether the polymorphisms of the NAT2, GSTM1, and GSTT1 enzymes affect age at diagnosis of first colorectal adenomas or extracolonic manifestations in 411 FAP patients. Neither age at diagnosis of colorectal adenomas nor occurrence of extra-intestinal tumors differed significantly between genotypes at the NAT2 and GSTM1 loci, whereas GSTT1 polymorphism showed an uncertain association with extra-intestinal manifestations. Combinations of supposed at risk genotypes of the three enzymes showed no significant differences either. Thus, NAT2, GSTM1, or GSTT1 are unlikely to modify the disease phenotype in FAP patients.

Loss of DNA mismatch repair proteins in skin tumors from patients with Muir-Torre syndrome and MSH2 or MLH1 germline mutations: establishment of immunohistochemical analysis as a screening test.

Muir-Torre syndrome (MTS) is a rare autosomal-dominant disorder characterized by the predisposition to both sebaceous skin tumors (or multiple keratoacanthomas) and internal malignancies. A subtype of MTS is allelic to hereditary nonpolyposis colorectal cancer and is caused by germline mutations in the DNA mismatch repair genes MSH2 or MLH1. In these cases both internal and skin tumors show characteristic microsatellite instability (MSI). The aim of the present study was to determine whether immunohistochemical examination of MSH2 or MLH1 protein expression in MTS-associated skin tumors can be used as a diagnostic screening tool to identify patients with germline mutations in MSH2 or MLH1. In the present study 28 skin lesions from 17 patients (20 sebaceous gland tumors, 4 sebaceous hyperplasias, 3 keratoacanthomas, and 1 squamous cell carcinoma) were tested immunohistochemically with antibodies against MSH2 and MLH1. Eighteen of these tumors were from eight patients with known MSH2 germline mutations, two tumors were from a patient with a germline mutation in MLH1, and eight microsatellite stable sporadic skin tumors served as controls. One sample had to be excluded because of a lack of immunoreactivity. All eight microsatellite stable tumors expressed both DNA repair proteins. In 15 of the tumors from MSH2 germline mutation carriers, loss of MSH2 expression was observed, one tumor showed reduced MSH2 expression, and one tumor displayed positive immunoreactivity to MSH2. Both tumors of the MLH1 germline mutation carrier showed loss of the MLH1 protein. In conclusion, our findings demonstrate that immunohistochemical testing of MTS-related skin tumors is a reliable screening method with high predictive value for the diagnosis of the DNA mismatch repair-deficient MTS.

On the anisotropy of skeletal muscle tissue under compression.

This paper deals with the role of the muscle fibres and extracellular matrix (ECM) components when muscle tissue is subjected to compressive loads. To this end, dissected tissue samples were tested in compression modes which induced states of fibres in compression (I), in tension (II) or at constant length (III), respectively. A comparison of the stress responses indicated that the tissue behaviour is significantly different for these modes, including differences between the modes (I) and (III). This contradicts the paradigm of many constitutive models that the stress response can be decomposed into an isotropic part relating to the ECM and an anisotropic fibre part the contribution of which can be neglected under compression. Conversely, the results provide experimental evidence that there is an anisotropic contribution of the fibre direction to the compressive stress. Interpreting these results in terms of recent microscopical studies, potential connections between the observed behaviour and the structure of muscle ECM are established.