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Biomedical subjects

B R Migeon

Publications and source records attributed to B R Migeon.

At least 73 records · Page 4Linked to original sources
PubMed1983

Glucose-6-phosphate dehydrogenase as a probe for the study of X-chromosome inactivation in hunan females.

The electrophoretic variants of G6PD have provided evidence for a single active X-chromosome in human somatic cells, but two active X-chromosomes in germ cells and in triploid cells with a 69,XXY karyotype. Studies of clonal populations of cells heterozygous for G6PD-A but expressing a single isozyme have provided evidence that the maintenance of the inactivation process is extremely stable, but that an occasional localized derepression event may occur. Such populations have also been used to show that methylation of X-chromosome DNA is not obviously different for XA and XI; furthermore, methylation is surprisingly unstable at least at some sites on the X. Studies of derepressed loci on the inactive X-chromosome in these clonal cell populations show that the expression of loci that escape inactivation is greater on XA than XI. G6PD variants have provided significant insights into the consequences of X-chromosome inactivation, revealing the role of intercellular communication and cell selection in determining female phenotype. The evidence that is now available indicates that the regulation of the X-chromosome is complex. The mechanisms involved include: 1) initiation, an event that may be mediated through an autosomal product and 2) maintenance which is stable but subject to programmed derepression of the entire chromosome during ontogeny of germ cells and occasional unprogrammed localized derepression in somatic cells. In addition, there may be transcriptional differences between XI and XA to compensate for monosomy of X-linked genes. The molecular basis for this multilevel regulation is unclear, but it seems certain that the cloned DNA probe for G6PD [Persico et al, 1981] will provide further insights.

Alleles↗
PubMed1982

Differential expression of steroid sulphatase locus on active and inactive human X chromosome.

The X chromosome in mammalian somatic cells is subject to unique regulation--usually genes on a single X chromosome are expressed while those on other X chromosomes are inactivated. The X-locus for steroid sulphatase (STS; EC 3.1.6.2), the microsomal enzyme that catalyses the hydrolysis of various 3 beta-hydroxysteroid sulphates, is exceptional because it seems to escape inactivation. Evidence for this comes from fibroblast clones in females heterozygous for mutations that result in a severe deficiency of this enzyme in affected males; all clones from these heterozygotes have STS activity, and enzyme-deficient clones that are expected if the locus were subject to inactivation, have not been found. Further evidence that the STS locus escapes inactivation is that the human inactive X chromosomes contributes STS activity to mouse-human hybrid cells. On the basis of these hybrid studies the STS locus has been mapped to the distal half of the short arm (p22-pter) of the human X chromosome. Although the STS locus on both X chromosomes in human female cells is expressed, quantitative measurements of STS activity in males and females do not accurately reflect the sex differences in number of X chromosomes (Table 1). The ratio of mean values for normal females to that of normal males is greater than 1:1 but less than the ratio of 2:1 expected if STS loci on all X chromosomes were equally expressed. The incomplete dosage effect suggests that the STS locus on the inactive X chromosome might not be fully expressed. To test this hypothesis, we examine two heterozygotes for X-linked STS deficiency who were also heterozygous for the common electrophoretic variants of glucose-6-phosphate dehydrogenase (G6PD A and B). Studies of fibroblast clones from these females provide evidence, presented here, for differential expression of STS loci on the active and inactive X chromosome.

Dosage Compensation, Genetic↗
PubMed1982

Studies of X chromosome DNA methylation in normal human cells.

Studies of methylation along 28 kilobases of X-chromosome DNA, assayed by Southern blot analysis using cloned X-chromosome-specific probes, indicates that X DNA methylation in normal human cells changes with replication, is not correlated with number of X chromosomes or transcriptional activity, and is less stable and more prevalent than when the human X is in the foreign environment of mouse-human hybrid cells. In contrast with observations of others in heteroploid cells, we observed no derepression of the inactive X in clonal populations of normal human fibroblasts treated with 5-azacytidine. This may reflect the differences in stability of the methylation of the human X in a foreign environment. Our observations preclude ubiquitous methylation differences as the molecular basis for X-chromosome inactivation.

Azacitidine↗
PubMed1982

Derepression with decreased expression of the G6PD locus on the inactive X chromosome in normal human cells.

Studies of a unique clone of skin fibroblasts from a normal 46 XX female reveal that the G6PD locus on the inactive X chromosome has been derepressed. The reactivation event occurs spontaneously, and is associated with normal karyotype, including the presence of a late-replicating X chromosome. Analysis of mouse-human hybrids with the relevant chromosome provides evidence that the derepressed locus is on the inactive X, and that reactivation is not extensive (the PGK locus is not derepressed). Nor is any general change in DNA methylation of this chromosome detectable with Hpa II and an X-specific DNA probe. Studies of the glucose-6-phosphate dehydrogenase phenotype in these heterozygous cells indicate that the reactivated X produces only half the enzyme subunits as are produced by the active X. Although this dosage difference may be related to the mutational event responsible for derepression of the locus, these observations along with other evidence suggest that loci on the inactive X, when expressed, have less activity than corresponding loci on the active X.

Clone Cells↗
PubMed1982

The prenatal diagnosis of adrenoleukodystrophy. Demonstration of increased hexacosanoic acid levels in cultured amniocytes and fetal adrenal gland.

Amniocentesis was performed in two women heterozygous for adrenoleukodystrophy (ALD). One fetus was male, and the hexacosanoic acid (C26) level in the cultured amniotic cells was 0.808 microgram per mg of protein, compared to 0.104 +/- 0.069 (S.D.) in controls. Pregnancy was interrupted at 22 wk gestation. The fetal adrenal cortex showed the ultrastructural inclusions characteristic of ALD and C26 accounted for 35% of the fatty acids in the cholesterol esters extracted from this tissue, more than one thousand times control. The second amniocentesis was performed in a woman who was also heterozygous for an electrophoretic variant of glucose 6- phosphate dehydrogenase (G6PD), and a member of a kindred showing genetic linkage of loci for ALD and G6PD. The fetus was female and the C26 level in cultured amniotic cells was 0.577. Pregnancy was interrupted at 11 wk for reasons unrelated to ALD. Study of C26 level and G6PD type in cultured fetal tissues confirmed heterozygosity for ALD.

Adolescent↗
PubMed1981

Studies of the locus for androgen receptor: localization on the human X chromosome and evidence for homology with the Tfm locus in the mouse.

We have established a cell line from mouse kidney cells expressing the tfm mutation and showed that these cells lack androgen binding activity. A subclone of these simian virus 40 (SV40)-transformed cells (6TGR-SV-tfm) selected in 6-thioguanine and lacking hypoxanthine phosphoribosyltransferase was used to produce a series of mouse--human hybrids containing the normal human X chromosome or various X autosome-translocation chromosomes (expressing only segments of the human X chromosome). When the androgen receptor locus (AR) was present in the hybrid, the number of receptor sites and kinetics of binding were similar to that in the human parental cells. Analysis of hybrids with partial human X chromosomes by using assays for X chromosome-linked enzymes and for the androgen receptor protein indicate that the AR locus on the human X chromosome is near the centromere between Xq13 and Xp11 and is proximal to the locus for phosphoglycerate kinase. Hybrids derived from 6TGR-SV-tfm mouse cells and human labial fibroblasts from an XY individual with the ar- form of androgen insensitivity have no binding activity. The lack of complementation indicates that the X chromosome-linked mutations in mouse and man affect homologous loci and supports the evolutionary conservation of X chromosomal loci in mammals; however, the position of the locus on the human X chromosome indicates that intrachromosomal rearrangement has occurred.

Animals↗
PubMed1981

Adrenoleukodystrophy: evidence for X linkage, inactivation, and selection favoring the mutant allele in heterozygous cells.

Skin fibroblasts of human males affected with adrenoleukodystrophy (ALD) have previously been shown to be abnormal with respect to C26 fatty acid content. Skin fibroblast clones from heterozygotes in three families segregating this mutation have been analyzed and are of two types: clones with normal ratios of C26 to C22 fatty acids and clones with an excess of C26 fatty acids similar to that found in cells of affected males. This indicates not only that the locus is X linked but also that it is subject to inactivation. In most of the heterozygotes there were significantly more clones of abnormal type than those expressing the normal allele, indicating a proliferative advantage in vitro for skin fibroblasts of mutant type. The increased levels of fatty acids in plasma in most heterozygotes and the phenotype of blood cells of women heterozygous for both ALD and glucose-6-phosphate dehydrogenase (G6PD) in one family are evidence that selection favoring the mutant allele may occur in vivo as well as in vitro and may explain why many heterozygotes manifest clinical symptoms of the disease. These studies have also revealed the close linkage between ALD and G6PD loci, because there are no recombinants among 18 informative offspring of doubly heterozygous mothers. Therefore, the ALD locus can be mapped on the human X chromosome near the G6PD locus at Xq28.

Adrenal Insufficiency↗
PubMed1981

Fragile X syndrome: search for phenotypic manifestations at loci for hypoxanthine phosphoribosyltransferase and glucose-6-phosphate dehydrogenase.

The subjects of this study were individuals with the form of X-linked mental retardation that is associated with the presence of a cytologically variant X chromosome having a secondary constriction or "fragile site" at Xq 27-28 (Fra X). Studies were carried out to test the hypothesis that deletions or modifications at neighboring loci occur as a consequence of events at the fragile site. Skin fibroblasts and peripheral blood lymphocytes from affected males were analyzed with respect to the expression of two X-lined enzymes: glucose-6-phosphate dehydrogenase (G6PD) and hypoxanthine phosphoribosyltransferase (HPRT); loci for these enzymes are known to be located in the region of the fragile site. Although the number of cells resistant to thioguanine (HPRT-deficient) obtained from some cultures from one Fra X male and blood cells of another was greater than expected, the frequency of these cells was not increased in cultures from other Fra X males. Furthermore, our results indicate that the G6PD activity and electrophoretic mobility in Fra X males is similar to that in normal cells, thus providing no evidence for the loss of the long-arm telomere in the fragile X syndrome.

Adult↗
PubMed1981

Phenotypic heterogeneity within clones of fetal human cells.

The heterogeneity of cell morphology characteristics of some colonies of human fetal kidney and amniotic fluid cells has been analyzed by biochemical and cell-cloning techniques. All the presumed subclones derived from dimorphic colonies were initially epithelioid, but some cells became fibroblastlike as the cell density increased. To determine if the observed heterogeneity occurred within clonal populations of cells, we determined the isozyme phenotype of dimers from renal cells heterozygous for glucose-6-phosphate dehydrogenase (G6PD). Colonies showing mixed cellular morphology expressed only a single G6PD isozyme, thus revealing their single-cell origin. Our results indicate that cell morphology is influenced by the cellular density within the clone, and that a single human renal cell in vitro can yield progeny of two morphological types.

Amniotic Fluid↗
PubMed1980

Mechanism of origin of complete hydatidiform moles.

Complete or 'true' hydatidiform mole, an abnormality of human gestation, is characterized by hydropic degeneraton of all placental villi, marked hypertrophy of the trophoblast, absence of a fetus and a propensity to become malignant. The chromosome constitution of complete moles is usually 46,XX, and Kajii et al. reported that the entire genome in seven out of seven cases was paternal in origin, with all centromere markers homozygous for paternal heteromorphisms. These observations, since confirmed, can be explained by the fertilization of an 'empty' egg--no effective genome--by either a haploid sperm that then duplicates without cytokinesis, to restore the diploid number, or by a diploid sperm resulting from failure of the second meiotic division. We report here a study of a series of complete moles that shows the first alternative to be correct in the majority of cases.

Female↗
PubMed1980

Localization of loci for hypoxanthine phosphoribosyltransferase and glucose-6-phosphate dehydrogenase and biochemical evidence of nonrandom X chromosome expression from studies of a human X-autosome translocation.

We report a unique and complex karyotypic rearrangement involving chromosomes X, 3, 7, and 21. Blood cells and fibroblasts from the proband do not express the maternal allele for glucose-6-phosphate dehydrogenase (G6PD), providing biochemical evidence for nonrandom expression of X-linked genes in balanced X-autosome translocations. The break point on the X chromosome, at the junction of Xq27-Xq28, separates the loci for hypoxanthine phosphoribosyltransferase (HPRT) and G6PD. Studies of mouse-human hybrids derived from the proband's cells indicate that G6PD, at q28, is clearly distal to all other X loci now assigned. From these and previous studies, we can localize HPRT to that segment between Xq26 and Xq27. The studies also provide further evidence for the stability of the inactive X phenotype in hybrid cells.

Alleles↗
PubMed1980

Complex chromosome rearrangements. Report of a new case and literature review.

A complex and unique, apparently balanced translocation involving three autosomes and an X in a phenotypically abnormal child is described. Family studies using glucose 6 phosphate dehydrogenase as a marker provided biochemical evidence of non-random expression of this Xq locus and suggested that this de novo abnormality in the proband could be paternal in origin--the first such instance to be recorded.

Abnormalities, Multiple↗
PubMed1980

Androgen receptors and metabolism in cultured human fetal fibroblasts.

Testosterone metabolism and dihydrotestosterone (DHT) receptor activity were studied in fibroblasts cultured from genital and non-genital tissues of 8- to 22-week old human fetuses. As early as the eighth week of gestation, DHT receptor activity was detected in non-genital skin. The binding capacity (Bmax) was greater in genital skin fibroblasts (mean +/- SD = 474 +/- 32 moles x 10--18/micrograms DNA) than non-genital skin (mean +/- SD = 124 +/- 42 moles x 10(-18)/micrograms DNA). DHT receptor binding (Bmax) was found in fibroblasts derived from testes (112 moles x 10(-18)/micrograms DNA), but not intestine (less than 10 moles x 10(-18)/micrograms DNA). The DHT receptor activity of fetal skin fibroblasts of genital origin was similar to that of fibroblasts derived from the foreskin of normal newborns. DHT receptors from fetal and newborn fibroblast cultures had similar sedimentation coefficients in sucrose density gradient centrifugation, but there were small differences in their relative affinities for 17 beta-estradiol and cyproterone. Low, but detectable 5 alpha-reductase activity was observed at 8 weeks gestation in non-genital skin fibroblasts and was present in fibroblasts from a variety of tissues of older fetuses, including testes, kidneys and lungs. The highest 5 alpha-reductase activity of 210 pg/hour/micrograms DNA was found in fibroblasts cultured from clitoral tissue from a 10-week old fetus. In all but one specimen, the 5 alpha-reduced products were either DHT or 5 alpha-androstanedione. The demonstration of 5 alpha-reductase activity and specific DHT receptors in fetal tissues suggests that the intracellular mechanism for androgen action is present in the fetus, similar to that after birth.

3-Oxo-5-alpha-Steroid 4-Dehydrogenase↗
PubMed1979

Human and mouse hypoxanthine-guanine phosphoribosyltransferase: dimers and tetramers.

Human and mouse hypoxanthine-guanine phosphoribosyltransferase subunits combine to form an active heteropolymer. Dimers form the basic subunit structure of the enzymes, yet the dimers can readily associate to form tetramers. The equilibrium between dimers and tetramers is significantly influenced by the ionic strength of the enzyme solvent.

Animals↗
PubMed1979

Enrichment of human heterokaryons by Ficoll gradient for complementation analysis of iduronate sulfatase deficiency.

Ficoll gradients have been used to enrich for heterokaryons in cultures of human skin fibroblasts following polyethylene glycol (PEG) induced fusion. These gradients provide a simple and consistent method for obtaining populations of multinucleated cells, at least twofold greater than those resulting from fusion alone. Formation of glucose-6-phosphate dehydrogenase (G6PD) heteropolymers has been used as a functional assay for the presence of heterokaryons. Analysis of cell populations enriched for multinucleated cells has revealed complementation leading to iduronate sulfatase activity in heterokaryons derived from iduronate sulfatase-deficient fibroblasts expressing the Hunter and multiple sulfatase-deficiency mutations.

Centrifugation, Density Gradient↗