Unexpected inactivation of acceptor consensus splice sequence by a −3 C to T transition in intron 2 of the CFTR gene

Human Genetics - Cystic fibrosis is caused by mutations in the cystic fibrosis transmembrane conductance regulator gene (CFTR). Analysis of DNA from a pancreatic sufficient patient by means of...     

Skewed inactivation of an X chromosome deleted at the dystrophin gene in an asymptomatic mother and her affected daughter

A girl with severe Becker muscular dystrophy and apparently normal chromosomes had a heterozygous deletion for exons 51, 52, and 53 of the dystrophin gene. This deletion was transmitted by her mother, who was unaffected. To differentiate the normal and the deleted X chromosomes, fluorescence in situ hybridization (FISH) was applied to metaphase chromosomes, using probes for both exons 51 and 52, which are only 388 and 113 base pairs long, respectively. FISH signals were observed in one or both chromatids of one chromosome, but never on both chromosomes, suggesting the lack of hybridization on the deleted X chromosome. Using 5-bromodeoxyuridine incorporation to differentiate the late (inactive) and the early replicating (active) X chromosomes, 77% of the signals were observed on the active X chromosomes in the mother. This percentage was only 18% in the daughter, suggesting that skewed inactivation of the X chromosomes was responsible for the phenotypic differences.     

Nucleotide substitutions at the -6 position in the promoter region of the factor IX gene result in different severity of hemophilia B Leyden: consequences for genetic counseling

Mutations in the promoter region of the factor IX gene result in hemophilia B Leyden, which is characterized by considerable improvement in the disease after puberty. We have found that distinct nucleotide substitutions at the -6 position in the Leyden-specific (LS) region are associated with a different severity of hemophilia B. The proband (aged 2) from one family is a severe hemophiliac with factor IX activity (F.IXC) and antigen (F.IXAg) levels less than 1.0U/dl. F.IXC and F.IXAg levels in two affected uncles are approximately 30% of normal levels. The LS region was targeted for analysis because the phenotypes suggested the inheritance of a factor IX Leyden gene. An abnormal TaqI digestion pattern was found in amplified DNA from the proband, and sequencing showed a G (-6) to C transversion that was linked to the disease in the family. In another family, two brothers (aged 8 and 9) suffer from mild hemophilia with F.IXC ranging from 7 to 10 U/dl and F.IXAg from 3 to 4 U/dl. They are the only documented members of the family with a bleeding tendency. Denaturing gradient gel electrophoresis on amplified fragments from one of the patient's genomic DNA corresponding to the 8 exons and flanking sequences of the factor IX gene suggested a defect only in a segment from the 5 region. This segment showed an altered TaqI digestion pattern, and sequencing demonstrated a G(-6) to A transition that was traced to the patients's mother and a grandmother. The different phenotypes associated with the G (-6) to A purine nucleotide transition compared with a G(-6) to C transversion provide evidence that this area is directly involved in the regulation of the human factor IX gene expression in vivo by binding of regulatory factors. The ability to predict that the conditions of a hemophilia B patient will improve with age has important implications for genetic counseling of the family. Therefore, the LS region should always be included when scanning the factor IX gene for mutations.     

A general suppressor of RNA polymerase I,II and III mutations in Saccharomyces cerevisiae

The pem locus, which is responsible for the stable maintenance of the low copy number plasmid R100, contains the pemK gene, whose product has been shown to be a growth inhibitor. Here, we attempted to isolate mutants which became tolerant to transient induction of the PemK protein. We obtained 20 mutants (here called pkt for PemK tolerance), of which 9 were temperature sensitive for growth. We analyzed the nine mutants genetically and found that they could be classified into three complementation groups, pktA, pktB and pktC, which corresponded to three genes, ileS, gltX and asnS, encoding isoleucyl-, glutamyl- and asparaginyl-tRNA synthetases, respectively. Since these aminoacyl-tRNA synthetase mutants did not produce the PemK protein upon induction at the restrictive temperature, these mutants could be isolated because they behaved as if they were tolerant to the PemK protein. The procedure is therefore useful for isolating temperature-sensitive mutants of aminoacyl-tRNA synthetases.     

Chiral symmetry breaking: Experimental results and computer analysis of a liquid-phase autoxidation

The autoxidation of tetralin is treated as a model reaction system to define the applicability of stereospecific autocatalysis. This concept, predicting a spontaneous amplification of enantiomeric excess generated by an autocatalytic chemical reaction, is used in several theoretical models as an explanation for the origin of natural optical activity. The reaction system investigated obeys the basic criteria of these models: a chiral intermediate (tetralin hydroperoxide) is produced from an achiral substrate (tetralin) via an autocatalytic pathway where the feedback mechanism is expected to generate a state of broken chiral symmetry. In order to test the amplification capacity of this reaction a computer analysis of the kinetic scheme is performed. This simulation is derived from the known kinetic scheme of autoxidation and is validated by fitting the experimentally observed data of hydroperoxide evolution. Calculations show that this model allows powerful amplification of enantiomeric excess and a transient amplification of the optical rotation. It is also demonstrated that the model system exhibits pronounced sensitivity toward any loss of absolute configuration of the involved chiral species. Since an amplification effect results exclusively at a high degree of stereoselectivity, it is concluded that stereospecific autocatalysis is possible in systems which show template reactions, crystallization, or colloidal effects. © 1993 Wiley-Liss, Inc.     

The gene coding for the α1 subunit of the skeletal dihydropyridine receptor (Cchl1a3=mdg) maps to mouse Chromosome 1 and human 1q32

Using both chromosomal in situ hybridization and molecular techniques, we report the genetic localization of the gene coding for the alpha 1 subunit of the skeletal slow Ca²⁺ current channel/DHP receptor gene (Cchl1a3) on human Chromosome (Chr) 1 (1q31–1q32 region) and on mouse Chr 1 region (F-G). On the basis of single-strand conformation polymorphism (SSCP-PCR) analysis in an interspecific backcross, we have determined that the Cchl1a3=mdg (muscular dysgenesis) locus is very closely linked to the myogenin (Myog) locus.     

Fine genetic mapping of the proximal part of mouse Chromosome 2 excludes Pax-8 as a candidate gene for Danforth's short tail (Sd)

Danforth's short tail (Sd) is a semidominant mutation of the mouse with effects on the skeleton and the urogenital system. In view of its phenotype and its position in the proximal part of Chromosome (Chr) 2, three genes qualified as possible candidates: Pax-8, a paired box-containing gene; Midkine (Mdk), a retinoic acid-responsive gene; and a new locus (Etl-4) identified by enhancer trapping with a lacZ reporter gene which showed expression in the notochord, the mesonephric mesenchyme, and the apical ectodermal ridge. Three different backcrosses involving all three genes in different combinations were set up and analyzed. From our results we conclude that Sd, Etl-4, Pax-8, and Mdk are independent loci, with Etl-4 being the closest genetic marker (1.1±1.4 cM) to the Danforth's short tail (Sd) gene.