|Year : 2011 | Volume
| Issue : 3 | Page : 23-24
Session K: Genotrichoses
|Date of Web Publication||16-Jun-2011|
Source of Support: None, Conflict of Interest: None
|How to cite this article:|
. Session K: Genotrichoses. Int J Trichol 2011;3, Suppl S1:23-4
Functional analysis of splice site mutations in the human hairless gene using a minigene assay
Sandra M. Pasternack 1 *, Melanie Refke 1 , Britta Fiebig 2 , Sabrina Wenzel 3 , Nina Ishorst 1 , Michael Ludwig 4 , Markus M. Nöthen 1,5 , Marieke M. Seyger 6 , Ben C. Hamel 7 , Regina C. Betz 1
1 Institute of Human Genetics, University of Bonn, Bonn, Germany; 2 Center for Human Genetics, Regensburg, Germany; 3 Department of Dermatology, University of Regensburg, Regensburg, Germany; 4 Institute for Clinical Chemistry and Pharmacology, Life and Brain Center, University of Bonn, Bonn, Germany; 5 Department of Genomics, Life and Brain Center, University of Bonn, Bonn, Germany; 6 Department of Dermatology, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands; 7 Department of Human Genetics, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands.
Atrichia universalis congenitalis (AUC) and atrichia with papular lesions (APL) are rare autosomal-recessive forms of isolated alopecia caused by mutations in the human hairless (HR) gene. Patients are born with normal hair that is shed almost completely and irreversibly during the first weeks of life. We investigated the molecular genetic basis of APL and AUC in two patients by direct sequencing of the HR gene. We identified two novel homozygous mutations: an insertion in exon 2 (c.485insT) which leads to a frameshift and premature stop and a splice site mutation (c.2847-1G>A). As biocomputational analyses did not reveal a clear prediction for the consequence of the splice mutation we decided to perform a functional splicing assay which resulted in an unexpected finding. This prompted us to analyse all known HR splice site mutations by this minigene assay. Seven different HR splice site mutations have been published up to date but none of them has been investigated experimentally. Therefore, we cloned the respective sequences into the exontrap cloning vector which allows directed splicing. After transfection of COS7 cells, isolation of RNA and cDNA synthesis, we analysed the products by sequencing. Comparison of the results with two biocomputational programmes revealed that the predictions are concordant with the experiments in five respective two of the eight cases. We also calculated consensus values for the mutations and the wild type sequences according to Shapiro and Senapathy (1987). This is the first study analysing the consequences of HR splice site mutations by a cell-based in vitro assay.
Genetics of congenital generalized hypertrichosis
McKusick-Zhang Center for Genetic Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.
Congenital generalized hypertrichosis (CGH) is a genetically and phenotypically heterogeneous group of rare conditions characterized by universal hair overgrowth. It is the major phenotypic feature of multiple distinct genetic syndromes and can be inherited as an autosomal or X-linked dominant trait. CGH, especially its genetic mechanism, has generated much scientific interest and media attention largely due to the striking hairy phenotype and its apparent atavistic nature. To date, genetic defects have been found in two forms of autosomal dominant CGH. We have shown that copy number variations (CNVs) on chromosome 17q24 are responsible for congenital generalized hypertrichosis terminalis with or without gingival hyperplasia (MIM 135400). Rearrangements of chromosome 8 have been detected in hypertrichosis universalis congenita, Ambras type (MIM 145701). Patel and colleagues mapped the first CGH locus to chromosome Xq24-q27.1 in 1995 in a large Mexican family segregating a unique X-linked hypertrichosis (MIM 307150). We ascertained a five-generation Chinese family with X-linked CGH and mapped the disease locus to a 5.6 Mb region within the interval defined by the previously reported Mexican family. I will present our results of genetic mapping and molecular analysis in X-linked CGH.
Latest news in hypotrichosis research
Regina C. Betz
Institute of Human Genetics, University of Bonn, Bonn, Germany.
Monogenic isolated alopecias comprise a group of clinically and genetically heterogeneous forms of hairlessness or hair loss. Clinical classification of the isolated alopecias is based on the onset of the disorder, the regions affected, and the structure of the hair shaft. Men and women are equally affected, and the mode of inheritance is autosomal dominant or autosomal recessive. Since the identification of the keratin gene KRT86 as a cause of the so-called monilethrix in the late nineties, mutations in several other genes have been identified for various isolated alopecias, e.g. hairless, desmoglein 4, lipase H, P2RY5, APCDD1 and corneodesmosin. Nevertheless, the pathogenetic causes of a large number of families suffering from hypotrichosis or alopecia are still unsolved and these families are in the focus of further research studies. Within the last few years, however, molecular genetic research has progressed enormously, mainly through the application of newly developed methods, e.g. chip technologies and resequencing techniques which might help to identify new genes. The latest data concerning hypotrichosis will be presented. Molecular genetic and pathophysiological studies of these rare disorders of hair development have contributed significantly to our understanding of the pathomechanisms of hair loss as well as the basic physiological mechanisms of hair growth.
New developments in Birt-Hogg-Dubé syndrome
Monique N. H. Luijten, Tijs Claessens, Marigje Vernooij, Barry J Coull, Maurice A. M. van Steensel*
Department of Dermatology, GROW School for Oncology and Developmental Biology, Maastricht University Medical Center, Maastricht, The Netherlands.
Birt-Hogg-Dubé syndrome (BHD, MIM # 135150) is an autosomal dominantly inherited disorder that predisposes to kidney cancer. In addition, patients develop benign hair follicle tumors and are at risk of pneumothorax due to lung cysts. The causative gene is known and codes for a conserved protein called folliculin (FLCN). Although of obvious importance, its function has remained elusive. Conflicting data have suggested an interaction with mTOR signaling, but more recent findings are pointing towards hypoxia inducible factor 1 and transforming growth factor beta signaling as potential culprits. We now present breakthrough findings from our research on FLCN and BHD syndrome that will help to understand FLCN's role in normal and abnormal cellular function. Our data will also shed new light on the pathogenesis of hair follicle tumors in BHD and related disorders.
Pili annulati - Update on the genetic analysis
Kathrin A. Giehl*, Gertrud Eckstein 1 , Anna Benet-Pagès 1 , Sebastian H. Eck 1
Department of Dermatology, Ludwig-Maximilians-University, Munich, Germany; 1 Institute of Human Genetics, Helmholtz-Zentrum, Neuherberg, Germany.
Pili annulati is an autosomal-dominant inherited disorder with alternating dark and light bands in the hair shaft of affected individuals. A linkage analysis in 9 families with 141 family members (65 affected and 76 non-affected) identified a candidate region of 2.9 Mb on chromosome 12q24.33-ter. Although the coding regions of all genes have been sequenced with standard methods, no causative gene could be identified yet. Therefore second generation sequencing was performed with Illumina technology. In total, we generated 125,464,762 36-bp paired-end reads in 7 lanes of an Illumina flow-cell. This resulted in a sequencing yield of 11.25 Gb. Alignment of the reads was performed with the MAQ software (v.0.7.1). We used the human hg19 assembly as reference. Single nucleotide variant calling and filtering was performed using MAQ and custom developed Perl scripts. All discovered variants were annotated using information from the UCSC Genome Browser database. We excluded all intergenic, intronic and known SNPs from dbSNP (v.129) and filtered the variants to the designated target area. This resulted in a list of 10 variants which were selected for further
P-cadherin is a novel key player in the control of human hair growth and pigmentation
Liat Samuelov-Kinori 1 *, Jennifer E. Kloepper 2 , Eli Sprecher 1,3 , Ralf Paus 2,4
1 Department of Dermatology, Tel-aviv Sourasky Medical Center, Tel-aviv, Israel; 2 Department of Dermatology, University of Lübeck, Lübeck, Germany; 3 Department of Human Molecular Genetics and Biochemistry, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, Israel; 4 School of Translational Medicine, University of Manchester, Manchester, UK.
P-cadherin belongs to the classical cadherin family of proteins. It is part of adherens junctions in various epithelia, including the follicular epithelium. Loss-of-function mutations in CDH3, which encodes P-cadherin, result in hypotrichosis with juvenile macular dystrophy (HJMD), an autosomal recessive disorder featuring sparse and short hair throughout life associated with progressive degeneration of the retinal macula leading to blindness during the second decade of life. In an attempt to recapitulate the functional effects of defective P-cadherin in a normal human hair model, we compared HF shaft production, cycling and pigmentation in human anagen VI scalp hair follicles (HFs) transfected with lipofectamine and CDH3-specific or scrambled control siRNAs. Successful know-down of P-cadherin was achieved. P-cadherin silencing inhibited hair shaft growth, caused premature catagen induction, reduced HF pigmentation, and inhibited hair matrix keratinocyte proliferation. When only anagen VI and control HFs were compared, P-cadherin knock-down significantly reduced gp100, MITF and SCF expression at the RNA and protein level as well as tyrosinase activity in situ. c-kit and β-catenin expression were also reduced. To the best of our knowledge, we report the first successful knock-down of a cadherin family member in an intact human organ. Moreover, our data provide direct functional evidence that P-cadherin-mediated signaling is important for the control of human HF growth and cycling and the first evidence that P-cadherin plays an unexpectedly important regulatory role in the control of human HF pigmentation.
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