Showing papers by "Stylianos E. Antonarakis published in 1990"

A cluster of cystic fibrosis mutations in the first nucleotide-binding fold of the cystic fibrosis conductance regulator protein.

Abstract: THE gene responsible for cystic fibrosis (CF) has recently been identified and is predicted to encode a protein of 1,480 ami no acids called the CF transmembrane conductance regulator (CFTR)1,2. Several functional regions are thought to exist in the CFTR protein, including two areas for ATP-binding, termed nucleotide-binding folds (NBFs), a regulatory (R) region that has many possible sites for phosphor} lation by protein kinases A and C, and two hydro-phobic regions that probably interact with cell membranes2. The most common CF gene mutation leads to omission of phenv lalanine residue 508 in the putative first NBF, indicating that this region is functionally important2–4. To determine whether other mutations occur in the NBFs of CFTR, we determined the nucleotide sequences of exons 9, 10, 11 and 12 (encoding the first NBF) and exons 20, 21 and 22 (encoding most of the second NBF) from 20 Caucasian and 18 American-black CF patients. One cluster of four mutations was discovered in a 30-base-pair region of exon 11. Three of these mutations cause amino-acid substitutions at residues that are highly conserved among the CFTR protein, the multiple-drug-resistance proteins and ATP-binding membrane-associated transport proteins. The fourth mutation creates a premature termination signal. These mutations reveal a functionally important region in the CFTR protein and provide further evidence that CFTR is a member of the family of ATP-dependent transport proteins2,5.

The polydeoxyadenylate tract of Alu repetitive elements is polymorphic in the human genome.

Abstract: To identify DNA polymorphisms that are abundant in the human genome and are detectable by polymerase chain reaction amplification of genomic DNA, we tested the hypothesis that the polydeoxyadenylate tract of the Alu family of repetitive elements is polymorphic among human chromosomes. We analyzed the 3' ends of three specific Alu sequences and found that two (in the adenosine deaminase gene and the beta-globin pseudogene) were polymorphic. This novel class of polymorphisms, termed AluVpA [Alu variable poly(A)] may represent one of the most useful and informative group of DNA markers in the human genome.

Two patients with cystic fibrosis, nonsense mutations in each cystic fibrosis gene, and mild pulmonary disease.

Abstract: CYSTIC fibrosis is the most common lethal inherited disorder in the white population. It is manifested by viscous secretions in the lungs and pancreas and abnormal electrolyte composition of sweat.1 Epithelial cells from patients with cystic fibrosis have abnormal conductance of chloride ions across apical membranes due to defective regulation of a particular chloride channel.2 3 4 Inadequate secretion of chloride is believed to cause the insufficient hydration of mucus in the airways and pancreatic ducts.1 The cystic fibrosis gene has recently been identified, and it is predicted to encode a 1480-amino-acid protein termed the cystic fibrosis transmembrane conductance regulator (CFTR).5 , 6 The . . .

Human erythropoietin gene expression in transgenic mice: multiple transcription initiation sites and cis-acting regulatory elements.

Abstract: Erythropoietin (EPO) is the primary humoral regulator of mammalian erythropoiesis. The single-copy EPO gene is normally expressed in liver and kidney, and increased transcription is induced by anemia or cobalt chloride administration. To identify cis-acting DNA sequences responsible for regulated expression, transgenic mice were generated by microinjection of a 4-kilobase-pair (kb) (tgEPO4) or 10-kb (tgEPO10) cloned DNA fragment containing the human EPO gene, 0.7 kb of 3'-flanking sequence, and either 0.4 or 6 kb of 5'-flanking sequence, respectively. tgEPO4 mice expressed the transgene in liver, where expression was inducible by anemia or cobalt chloride, kidney, where expression was not inducible, and other tissues that do not normally express EPO. Human EPO RNA in tgEPO10 mice was detected only in liver of anemic or cobalt-treated mice. Both tgEPO4 and tgEPO10 mice were polycythemic, demonstrating that the human EPO RNA transcribed in liver is functional. These results suggest that (i) a liver inducibility element maps within 4 kb encompassing the gene, 0.4 kb of 5'-flanking sequence, and 0.7 kb of 3'-flanking sequence; (ii) a negative regulatory element is located between 0.4 and 6 kb 5' to the gene; and (iii) sequences required for inducible kidney expression are located greater than 6 kb 5' or 0.7 kb 3' to the gene. RNase protection analysis revealed that human EPO RNA in anemic transgenic mouse liver and hypoxic human hepatoma cells is initiated from several sites, only a subset of which is utilized in nonanemic transgenic liver and human fetal liver.

Phylogeny of human β‐globin haplotypes and its implications for recent human evolution

Abstract: The evolutionary histories and relationships among African, Eurasian, and Pacific Island populations are investigated by using observations on five polymorphic restriction sites in the beta-globin gene cluster. We present new data on 222 chromosomes from a global sample and combine these with previously published observations on 591 chromosomes. It is shown that the data are rich in rare haplotypes and that rare variants are not helpful for standard methods of population structure analysis. Consequently, a new approach is developed. We first consider the phylogeny of beta-globin haplotypes. The roles of mutation, gene conversion, and recombination in the generation of haplotype diversity are specifically focused upon. The relationships among human populations are then inferred from the phylogenetic relationships among the haplotypes, their presence or absence, and frequencies within populations. Questions regarding whether or not a phyletic process can account for relationships among the major geographical populations and whether or not an extant human population exhibits the qualities that would be expected of an ancestral group are addressed. The results of this analysis support an African origin for modern Homo sapiens and a phyletic structuring of the major geographical regions. However, it is shown that divergence times for the various populations cannot be determined from these data.

Analysis of DNA polymorphisms suggests that most de novo dup(21q) chromosomes in patients with Down syndrome are isochromosomes and not translocations.

Abstract: Down syndrome is rarely due to a de novo duplication of chromosome 21 [dup(21q)]. To investigate the origin of the dup(21q) and the nature of this chromosome, we used DNA polymorphisms in 10 families with Down syndrome due to de novo dup(21q). The origin of the extra chromosome 21q was maternal in six cases and paternal in four cases. Furthermore, the majority (eight of 10) of dup(21q) chromosomes were isochromosomes i(21q) (four were paternal in origin, and four were maternal in origin); however, in two of 10 families the dup(21q) chromosome appeared to be the result of a Robertsonian translocation t(21q;21q) (maternal in origin in both cases).

Gene Defects in β‐Thalassemia and Their Prenatal Diagnosis

Abstract: In contrast to a-thalassemia, P-thalassemia is generally caused by point mutations. These mutations were studied extensively from 1980 to 1986, usually by haplotype analysis of the P-globin gene cluster followed by cloning and sequencing of the mutant P-globin gene. About 40 point mutations producing P-thalassemia were discovered by mid-1987, and many were documented by transient expression studies of the mutant gene.' In 1987 it became possible to amplify regions of the P-globin gene starting with genomic DNA from leukocyte^^^^ and to sequence directly the amplified p r o d u ~ t s . ~ , ~ By use of this approach in selected individuals known to lack the common alleles of the ethnic group represented by the patient under study, about 50 more alleles have been characterized in less than 2 years.6 In July 1990, the total number of point mutations which produce P-thalassemia known to the authors is 91 (TABLE 1 and FIG. 1). Since the total number of P-globin alleles known to produce any clinical phenotype is roughly 200, the P-thalassemia alleles now account for over 45% of the clinically significant alleles at the P-globin locus, and the \"abnormal hemoglobins\" account for the remainder. The spectrum of P-thalassemia alleles has been determined in a wide variety of population groups, including American blacks,',* Asian Indians,','\" Italians and Greeks,\" Spanish,\" Turks,\" Kurdish Jews,14 Chine~e , '~ . '~ Japane~e , '~ and Thai,'\" among others. In most affected population groups, such as Mediterranean peoples, Chinese and Southeast Asians, Asian Indians, and blacks of African origin, a handful of ethnic-group-specific alleles accounts for roughly 90-93% of P-thalassemia genes. For example, among Chinese from Kuontung province, 4 alleles account for about 90% of p-thalassemia genes.\" On the other hand, a large number of rarer alleles have been observed in each ethnic group. For example, to date 11 rare alleles account for the remaining 10% of P-thalassemia genes in Chinese. The total number of mutations obscrvcd in the well-studied groups is, in Chinese, 15; Asian Indians, 10; blacks, 12; Mediterraneans, 31. In the broad group of Mediterranean peoples it is important to note that although many of the 31 alleles have been observed in diverse regions of the Mediterranean basin, allele frequencies vary greatly from one country to another. Because so many alleles are found in each region, most individuals with P-thalassemia major carry two different alleles and are called genetic compounds.

Assignment of the Nance-Horan syndrome to the distal short arm of the X chromosome.

Abstract: There are three types of X-linked cataracts recorded in Mendelian Inheritance in Man (McKusick 1988): congenital total, with posterior sutural opacities in heterozygotes; congenital, with microcornea or slight microphthalmia; and the cataract-dental syndrome or Nance-Horan (NH) syndrome. To identify a DNA marker close to the gene responsible for the NH syndrome, linkage analysis on 36 members in a three-generation pedigree including seven affected males and nine carrier females was performed using 31 DNA markers. A LOD score of 1.662 at theta = 0.16 was obtained with probe 782 from locus DXS85 on Xp22.2-p22.3. Negative LOD scores were found at six loci on the short arm, one distal to DXS85, five proximal, and six probes spanning the long arm were highly negative. These results make the assignment of the locus for NH to the distal end of the short arm of the X chromosome likely.

Apolipoprotein A1 Baltimore (Arg10----Leu), a new ApoA1 variant.

Abstract: A new apolipoprotein A1 (APOA1) gene variant has been identified in a family ascertained through a proband undergoing coronary angiography. The variant, ApoA1 Baltimore, was due to a mutation at codon 34 of the third exon of the APOA1 gene (CGA to CTA) that resulted in an arginine-to-leucine substitution at the tenth amino acid of the mature ApoA1 and a change in charge of -1. The mutation abolishes a TaqI restriction site and it is easily detectable after polymerase chain reaction amplification of genomic DNA. The proband was heterozygous for the mutation. Eight other members of the pedigree had the same ApoA1 variant. Cosegregation of the variant with hypoalphalipoproteinemia could not be demonstrated and the association of this mutation with hypoalphalipoproteinemia was confined to three affected members of the nuclear family. No effect of the mutant on any lipoprotein phenotype could be established.

Evidence for involvement of a Robertsonian translocation 13 chromosome in formation of a ring chromosome 13.

Abstract: We have used molecular and cytogenetic methods to study the derivation of a ring chromosome 13 in the fetus of a woman mosaic for a translocation chromosome 13. DNA analysis showed that the translocation chromosome was a Robertsonian translocation, not an isochromosome. We suggest that the ring is derived from the translocation chromosome by breaks in both long arms and subsequent reunion, r(13) (q12q14).

Microdeletion in the X-Chromosome and Prenatal Diagnosis in a Family With Norrie Disease

Abstract: We have studied a three-generation family in which Norrie disease is segregating and have performed prenatal diagnosis on the fetus of an obligatory carrier. Deletions at loci DXS7 and DXS77 defined by probes L1.28, L1.28-p59, and pX59 were detected in the affected male. DNA studies of chorionic villus biopsy material indicated that the male fetus had inherited the normal allele from the carrier mother. This prediction was confirmed on eye examination at age 5 months.

MspI polymorphic site in intron 22 of the factor VIII gene in the Japanese population.

Abstract: A novel MspI DNA polymorphic site has been found in intron 22 of the human factor VIII gene. This site is informative almost exclusively in the Japanese population (heterozygosity 0.45) and will be of considerable importance in carrier detection and prenatal diagnosis of hemophilia A in this population.

Approaches to the Diagnosis of Renal Genetic Disorders Using DNA Analysis

Abstract: Since the molecular structure of DNA was described by Watson and Crick in 1954, there have been remarkable advances in understanding the structure and function of eukaryotic genomes. Teachniques have been developed to manipulate genes, to reproduce (clone) them in large number, and to analyze them for alterations from the normal state, in individuals with genetic diseases. Assignment of normal and mutant genes to the human chromosomal map has enabled the diagnosis of genetic diseases, even in cases where the basic biochemical defect underlying the disease in question remains unknown.