Relevance of the lariat model to the splicing of the H-2 gene family

We examined the splicing of the H-2 gene family, taking the H-2Kd as a prototype, in the framework of the lariat model. We mainly investigated the mechanism described by Konarska et al. [Nature (Lond.) 313, 552-557 (1985)] who propose a direct interaction between the 5' splicing site and the branching region. We also checked each of the H-2 introns for the presence of patterns resembling the published consensus for the branching region. The known splicing events in the H-2 gene family are not always consistent with the current models, and our results indicate that slightly different mechanisms govern the splicing of different introns. A tentative explanation of the alternative splicing of the first and last intron, previously described, is given. The removal of the large third intron is not easily rationalized unless new rules for an additional multistep processing are postulated.     

Electrophysiological evidence for the autoregulation of β-cell secretion by insulin

Electrophysiological studies of cultured rat pancreatic β-cells using intracellular microelectrodes show that exogenous insulin over the range of 0.1–10.0 μg/ml inhibits the electrical activity due to 27.8 mM glucose in a dose-related manner. This inhibitory effect is manifested by a mean increase of the membrane potential from about ?20 to ?30 mV and inhibition of the manner of cells impaled showing spike activity from 60 to less than 10%. The inhibitory influence of insulin is rapid occuring within 5 min for the highest level used. The results provide evidence for a negative feedback role of insulin in regulating its own release.     

Mutations in three of the putative transmembrane helices of subunit a of the Escherichia coli F1F0-ATPase disrupt ATP-driven proton translocation

Three missense mutants in subunit a of the Escherichia coli F1F0-ATPase were isolated and characterized after hydroxylamine mutagenesis of a plasmid carrying the uncB (subunit a) gene. The mutations resulted in Asp119----His, Ser152----Phe, or Gly197----Arg substitutions in subunit a. Function was not completely abolished by any of the mutations. The F0 membrane sector was assembled in all three cases as judged by restoration of dicyclohexylcarbodiimide sensitivity to the F1F0-ATPase. The H+ translocation capacity of F0 was reduced in all three mutants. ATP-driven H+-translocation was also reduced, with the response in the Gly197----Arg mutant being almost nil and that in the Asp119----His and Ser152----Phe mutants less severely affected. The substituted residues are predicted to lie in the second, third, and fourth transmembrane helices suggested in most models for subunit a. The Gly197----Arg mutation lies in a very conserved region of the protein and the substitution may disrupt a structure that is critical to function. The Asp119----His and Ser152----Phe mutations also lie in areas with sequence conservation. A further analysis of randomly generated mutants may provide more information on regions of the protein that are crucial to function. Heterodiploid transformants, carrying plasmids with either the wild-type uncB gene or mutant uncB genes in an uncB (Trp231----stop) background, were characterized biochemically. The truncated subunit a was not detected in membranes of the background strain by Western blotting, and the uncB+ plasmid complemented strain showed normal biochemistry. The uncB mutant genes were shown to cause equivalent defects in either the heterodiploid background configuration, or after incorporation into an otherwise wild-type unc operon. The subunit a (Trp231----stop) background strain was shown to bind F1-ATPase nearly normally despite lacking subunit a in its membrane.     

Cystic fibrosis typing with DNA probes and screening for ΔF508 deletion in families from Southern France

Summary A sample of 235 individuals from 49 French cystic fibrosis (CF) families with at least one living affected child was typed with probes for restriction fragment length polymorphisms (RFLPs) known to be linked to the CF gene, and was screened for the ΔF508 mutation. Using a combination of six probes, 44 out of the 49 families were sufficiently informative to enable prenatal diagnosis or carrier determination. As in many other populations, linkage disequilibrium was found between the CF locus and the haplotype B (XV2c: allele 1; KM19: allele 2), which accounts for about 78% of CF chromosomes in our families. The ΔF508 deletion was present in 64.3% of CF chromosomes.     

Objective comparison of exon and intron sequences by means of 2-dimensional data analysis methods

Here we advocate the use of 2-dimensional data representation in the context of the informational approach of sequence analysis (Claverie & Bougueleret (1986) Nucleic Acids Research 14, 179-196) by applying these methods to the problem of intron/exon discrimination. Two main findings are reported: i) oligonucleotide patterns complementary to the Ul small nuclear RNA are specifically avoided in exon sequences, ii) vertebrate intron sequences, to the exclusion of other eukaryotic phyla, are characterized by a peculiar distribution of CpG containing patterns.     

The candidate gene for the X-linked Kallmann syndrome encodes a protein related to adhesion molecules

Kallmann syndrome associates hypogonadotropic hypogonadism and anosmia and is probably due to a defect in the embryonic migration of olfactory and GnRH-synthesizing neurons. The Kallmann gene had been localized to Xp22.3. In this study 67 kb of genomic DNA, corresponding to a deletion interval containing at least part of the Kallmann gene, were sequenced. Two candidate exons, identified by multiparameter computer programs, were found in a cDNA encoding a protein of 679 amino acids. This candidate gene (ADMLX) is interrupted in its 3' coding region in the Kallmann patient, in which the proximal end of the KAL deletion interval was previously defined. A 5' end deletion was detected in another Kallmann patient. The predicted protein sequence shows homologies with the fibronectin type III repeat. ADMLX thus encodes a putative adhesion molecule, consistent with the defect of embryonic neuronal migration.     

The Presence and Role of Transmembrane Transforming Growth Factor-α in Cultures of Rat Liver Epithelial Cells

We assessed the presence and the role of membrane TGF-α in two rat liver epithelial cell lines, either parental or transfected with c-fos proto-oncogene. c-fos overexpressing cells had more TGF-α-like activity in their membranes. When TGF-α was removed by elastase or neutralized, the growth rates of both cell lines were markedly reduced, but to a higher extent for parental cells. If membrane TGF-α seemed to play a key contribution in normal cell growth, both cell lines were unable to react to the addition of soluble TGF-α, showing that these two forms of growth factors are not equivalent.