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L J Shapiro

Publications and source records attributed to L J Shapiro.

At least 73 records · Page 4Linked to original sources
PubMed1989

X-chromosome inactivation in cultured cells from human chorionic villi.

X-chromosome inactivation was investigated in human chorionic villi in the first trimester of pregnancy and cultured cells established from them. Expression of glucose-6-phosphate dehydrogenase (G6PD) was evaluated in these extraembryonic cells from four females heterozygous for the electrophoretic variants (AB) of G6PD. In each case the uncultured villi as well as derived cultured cells expressed the AB phenotype for G6PD with about equal intensity for the A and B bands. Single-cell-derived clones established from two of the four cases expressed either G6PD A or B. One clone expressing G6PD B was fused with mouse cells, and a hybrid clone retaining the inactive human X chromosome was isolated; there was no evidence of human G6PD expression in this clone retaining an inactive human X. DNA methylation in the first intron of the human gene for hypoxanthine phosphoribosyltransferase (HPRT) was evaluated in the four pairs of cultured villi and fetal cells. No differences were detected between the cultured villi and fetal cells as they all showed bands characteristic of an inactive X from somatic cells. These results show that there is no preferential inactivation of an X in the majority of cells that constitute human tertiary chorionic villi or in cultured cells derived from them. Long-term cultures established from chorionic villi appear to be no different from somatic cells with respect to X-chromosome inactivation.

Animals↗
PubMed1989

Human and mouse amelogenin gene loci are on the sex chromosomes.

Enamel is the outermost covering of teeth and is the hardest tissue in the vertebrate body. The enamel matrix is composed of enamelin and amelogenin classes of protein. We have determined the chromosomal locations for the human and mouse amelogenin (AMEL) loci using Southern blot analyses of DNA from human, mouse, or somatic cell hybrids by hybridization to a characterized mouse amelogenin cDNA. We have determined that human AMEL sequences are located on the distal short arm of the X chromosome in the p22.1----p22.3 region and near the centromere on the Y chromosome, possibly at the proximal long arm (Yq11) region. These chromosomal assignments are consistent with the hypothesis that perturbation of the amelogenin gene is involved in X-linked types of amelogenesis imperfecta, as well as with the Y-chromosomal locations for genes that participate in regulating tooth size and shape. Unlike the locus in humans, the mouse AMEL locus appears to be assigned solely to the X chromosome. Finally, together with the data on other X and Y chromosome sequences, these data for AMEL mapping support the notion of a pericentric inversion occurring in the human Y chromosome during primate evolution.

Amelogenin↗
PubMed1989

Molecular studies of deletions at the human steroid sulfatase locus.

The human steroid sulfatase gene (STS) is located on the distal X chromosome short arm close to the pseudoautosomal region but in a segment of DNA that is unique to the X chromosome. In contrast to most X chromosome-encoded genes, STS expression is not extinguished during the process of X chromosome inactivation. Deficiency of STS (steryl-sulfatase; steryl-sulfate sulfohydrolase, EC 3.1.6.2) activity produces the syndrome of X chromosome-linked ichthyosis, which is one of the most common inborn errors of metabolism in man. Approximately 90% of STS- individuals have large deletions at the STS locus. We and others have found that the end points of such deletions are heterogeneous in their location. One recently ascertained subject was observed to have a 40-kilobase deletion that is entirely intragenic, permitting the cloning and sequencing of the deletion junction. Studies of this patient and of other X chromosome sequences in other subjects permit some insight into the mechanism(s) responsible for generating frequent deletions on the short arm of the X chromosome.

Arylsulfatases↗
PubMed1989

In situ hybridization of prepro-epidermal growth factor mRNA in the mouse kidney.

The presence of prepro-epidermal growth factor (prepro-EGF) mRNA was studied in the mouse kidney by in situ hybridization using [3H]prepro-EGF cDNA and 35S-labeled prepro-EGF cRNA probes. In addition, anti-EGF serum was utilized to immunolocalize the peptide by the avidin-biotin complex immunoperoxidase method. Both EGF immunoreactivity and prepro-EGF mRNA hybridization were localized to the thick ascending limb of Henle (TAL) and the distal convoluted tubule (DCT), whereas the macula densa was negative. The glomerulus, the proximal portion of the nephron, and the collecting system were negative. Computer-assisted image analysis of the optical density of the autoradiographic and immunocytochemical signals were performed. The medullary TAL expressed significantly less prepro-EGF than the cortical TAL and the DCT.

Animals↗
PubMed1988

The human X-linked steroid sulfatase gene and a Y-encoded pseudogene: evidence for an inversion of the Y chromosome during primate evolution.

The mammalian X and Y chromosomes are thought to have evolved from a common, nearly homologous chromosome pair. Although there is little sequence similarity between the mouse or the human X and Y, there are several regions in which moderate to extensive sequence homologies have been found, including, but not limited to, the so-called pseudoautosomal segment, in which X-Y pairing and recombination take place. The steroid sulfatase gene is in the pseudoautosomal region of the mouse, but not in man. We have cloned and characterized the human STS X-encoded locus and a pseudogene that is present on the long arm of the Y chromosome. Our data in humans and other primates suggest that there has been a pericentric inversion of the Y chromosome during primate evolution that has disrupted the former pseudoautosomal arrangement of these genes. These results provide additional insight into the evolution of the sex chromosomes and into the nature of this interesting portion of the human genome.

Animals↗
PubMed1988

Back muscle function during bipedal walking in chimpanzee and gibbon: implications for the evolution of human locomotion.

The evolution of erect posture and locomotion continues to be a major focus of interest among paleoanthropologists and functional morphologists. To date, virtually all of our knowledge about the functional role of the back muscles in the evolution of bipedalism is based on human experimental data. In order to broaden our evolutionary perspective on the vertebral region, we have undertaken an electromyographic (EMG) analysis of three deep back muscles (multifidus, longissimus thoracis, iliocostalis lumborum) in the chimpanzee (Pan troglodytes) and gibbon (Hylobates lar) during bipedal walking. The recruitment patterns of these three muscles seen in the chimpanzee closely parallel those observed in the gibbon. The activity patterns of multifidus and longissimus are more similar to each other than either is to iliocostalis. Iliocostalis recruitment is clearly related to contact by the contralateral limb during bipedal walking in both species. It is suggested that in both the chimpanzee and gibbon, multifidus controls trunk movement primarily in the sagittal plane, iliocostalis responds to and adjusts movement in the frontal plane, while longissimus contributes to both of these functions. In many respects, the activity patterns shared by the chimpanzee and gibbon are quite consistent with recent human experimental data. This suggests a basic similarity in the mechanical constraints placed on the back during bipedalism among these three hominoids. Thus, the acquisition of habitual bipedalism in humans probably involved not so much a major change in back muscle action or function, but rather an improvement in the mechanical advantages and architecture of these muscles.

Animals↗
PubMed1988

Metabolism of 3H-dehydroepiandrosterone sulphate by subjects with steroid sulphatase deficiency.

Patients with steroid sulphatase deficiency develop ichthyosis with accumulation of cholesterol sulphate in plasma and in the stratum corneum. The present study was undertaken to determine whether desulphation of the C19 steroid DHEAS is also impaired. The mean plasma concentrations of DHEA and androstenedione were significantly lower for patients than for controls (p less than 0.02 and 0.001) while the mean concentration of DHEAS was higher (p less than 0.002). Following intravenous administration of 3H-DHEAS, one patient failed to desulphate 3H-DHEAS as evidenced by an absence of urinary 3H-glucuronides. A second produced normal amounts of urinary 3H-glucuronides (indicative of desulphation capacity) in a baseline study but did not desulphate 3H-DHEAS following ampicillin treatment to alter gut microflora. A third patient had consistent sulphatase activity with and without ampicillin.

Adult↗
PubMed1988

In situ hybridization of nerve growth factor mRNA in the mouse submandibular gland.

We studied the presence of beta-nerve growth factor (NGF) mRNA in the submandibular gland of the mouse by in situ hybridization using 35S-labeled prepro-beta-NGF antisense RNA. Female and male mice were studied at different stages of postnatal development, ranging from 3 to 12 weeks. Although NGF mRNA was detectable in the granular convoluted tubules of the submandibular gland in all the age and sex groups studied, the abundance of the signal dramatically increased after 5 weeks during the development of the submandibular gland. In addition, a conspicuous sexual dimorphism became increasingly apparent in the 6-, 7-, 10-, and 12-week-old animals, due to the remarkable development of the granular convoluted tubules in the adult male mouse, that expressed abundant NGF mRNA.

Animals↗
PubMed1987

Cloning and expression of steroid sulfatase cDNA and the frequent occurrence of deletions in STS deficiency: implications for X-Y interchange.

Human STS is a microsomal enzyme important in steroid metabolism. The gene encoding STS is pseudoautosomal in the mouse but not in humans, and escapes X inactivation in both species. We have prepared monoclonal and polyclonal antibodies to the protein which has been purified and from which partial amino acid sequence data have been obtained. cDNA clones containing the entire coding sequence were isolated, sequenced, and expressed in heterologous cells. Variable length transcripts have been shown to be present and due to usage of alternative poly(A) addition sites. The functional gene maps to Xp22.3-Xpter and there is a pseudogene on Yq suggesting a recent pericentric inversion. Absence of STS enzymatic activity occurs frequently in human populations and produces a visible phenotype of scaly skin or ichthyosis. Ten patients with inherited STS deficiency were studied and eight had complete gene deletions. The possibility that STS deficiency results from aberrant X-Y interchange is discussed.

Amino Acid Sequence↗
PubMed1987

Cytogenetic and molecular studies on a recombinant human X chromosome: implications for the spreading of X chromosome inactivation.

A pericentric inversion of a human X chromosome and a recombinant X chromosome [rec(X)] derived from crossing-over within the inversion was identified in a family. The rec(X) had a duplication of the segment Xq26.3----Xqter and a deletion of Xp22.3----Xpter and was interpreted to be Xqter----Xq26.3::Xp22.3----Xqter. To characterize the rec(X) chromosome, dosage blots were done on genomic DNA from carriers of this rearranged X chromosome using a number of X chromosome probes. Results showed that anonymous sequences from the distal end of the long arm to which probes 4D8, Hx120A, DX13, and St14 bind as well as the locus for glucose-6-phosphate dehydrogenase (G6PD) were duplicated on the rec(X). Mouse-human cell hybrids were constructed that retained the rec(X) in the active or inactive state. Analyses of these hybrid clones for markers from the distal short arm of the X chromosome showed that the rec(X) retained the loci for steroid sulfatase (STS) and the cell surface antigen 12E7 (MIC2); but not the pseudoautosomal sequence 113D. These molecular studies confirm that the rec(X) is a duplication-deficiency chromosome as expected. In the inactive state in cell hybrids, STS and MIC2 (which usually escape X chromosome inactivation) were expressed from the rec(X), whereas G6PD was not. Therefore, in the rec(X) X chromosome inactivation has spread through STS and MIC2 leaving these loci unaffected and has inactivated G6PD in the absence of an inactivation center in the q26.3----qter region of the human X chromosome. The mechanism of spreading of inactivation appears to operate in a sequence-specific fashion. Alternatively, STS and MIC2 may have undergone inactivation initially but could not be maintained in an inactive state.

Abnormalities, Multiple↗
PubMed1986

A new syndrome of anosmia, ichthyosis, hypogonadism, and various neurological manifestations with deficiency of steroid sulfatase and arylsulfatase C.

We describe a family consisting of 3 affected men with congenital ichthyosis, anosmia, hypogonadism, nystagmus with decreased visual acuity, strabismus, hypopigmentation of the iris, and mirror movements of the hands and feet. Two of them had limitation of ocular movement and unilateral renal agenesis or hypoplasia. The condition appears to be inherited as an X-linked recessive trait. Clinical, pathological, and biochemical evaluations were compatible with a diagnosis of X-linked ichthyosis. Steroid sulfatase and arylsulfatase C activities in leukocytes and fibroblasts were markedly diminished in the affected patients. Their hypogonadism was due to decreased luteinizing hormone-releasing hormone secretion (hypogonadotropic). Hyposecretion of antidiuretic hormone was also recognized. Chromosome analysis of leukocytes and skin fibroblasts revealed a normal 46,XY male karyotype in all of the patients.

Adolescent↗
PubMed1986

Stability of DNA methylation of the human hypoxanthine phosphoribosyltransferase gene.

Methylation sensitive restriction enzymes were used to evaluate the methylation level of several restriction sites near human hypoxanthine phosphoribosyltransferase (HPRT) genes on active and inactive X chromosomes. DNA samples from leukocytes, from clonally derived fibroblasts, and from independent mouse-human hybrid lines isolated from the fusion of A-9 cells and these clonally derived human cells were studied. Comparison of the methylation patterns shows that restriction sites may show variable or constant methylation among tissues and clones, and heritability of methylation is also different among restriction sites. Methylation is more stable at sites whose methylation status correlate well with HPRT activity. Our results suggest that the methylation of certain cytosine residues may critically affect gene activity and that the methylation pattern of these sites is stably inherited.

Animals↗
PubMed1986

New frontiers in genetic medicine.

Disorders determined wholly or in part by genetic factors constitute a substantial number of human diseases. This realization has grown during the past 2 decades with the recognition of many specific heritable conditions and the identification of familial risk factors for common disorders. New technologies, such as fetal visualization, chorionic villus sampling, molecular cloning methods, and gene transfer technology, provides a framework for dealing with genetically determined illness in unprecedented ways. Several current and potential applications of these methods are examined, as is the use of restriction fragment length polymorphisms to survey the variability within the genome and to generate markers permitting the prospective detection of genetic disorders. The promise and limitations of chorionic villus biopsy sampling are considered for early prenatal diagnosis. The future of gene therapy in hereditary diseases is examined, and some of the substantial social and ethical considerations engendered by these new developments are explored.

Abortion, Spontaneous↗
PubMed1986

Fine mapping of the distal short arm of the human X chromosome using X/Y translocations.

The loci for steroid sulfatase (STS), the deficiency of which causes X-linked ichthyosis, the cell surface antigen 12E7 (MIC2X), and the blood group antigen Xg (Xg) have been mapped to Xp22.3. These loci are of particular interest since they do not appear to undergo X-chromosome inactivation. In an attempt to establish the relative order of STS and MIC2X, fibroblasts from carriers of four different X/Y translocations and an X/10 translocation were obtained and fused with mouse cell lines deficient in hypoxanthine phosphoribosyltransferase. The breakpoints on the X chromosome in these five translocations are in Xp22. Several independent clones from each fusion were isolated in HAT medium. The clones were examined cytogenetically, and in each case at least two independent clones were identified that have an active X/Y or X/10 translocation chromosome in the absence of other X or Y material. These clones were then tested for STS and 12E7 expression. In two of the X/Y translocations, the markers, STS and 12E7, were both absent. In the X/10 and a third X/Y translocation, both markers were retained. In each of three clones containing the fourth X/Y translocation, STS activity was retained but 12E7 antigenicity was lost. Assuming that this is a simple translocation and does not represent a more complex rearrangement, these results suggest that MIC2X is distal to STS.

Animals↗
PubMed1985

Activator protein required for the enzymatic hydrolysis of cerebroside sulfate. Deficiency in urine of patients affected with cerebroside sulfatase activator deficiency and identity with activators for the enzymatic hydrolysis of GM1 ganglioside and globotriaosylceramide.

Urine specimens from two sibs affected with cerebroside sulfatase activator deficiency were examined to ascertain whether the deficiency of the supplementary activator protein required for the enzymatic hydrolysis of cerebroside sulfate was also evident in urine. Material from chromatographic fractionations was examined for the activator activity to avoid ambiguities resulting from protein inhibition. There were substantial deficits in all chromatographic fractions corresponding to activator-containing fractions of control urines. Since patient urines contained elevated amounts of lactosylceramide, digalactosylceramide, and globotriaosylceramide and since similarities between activators for cerebroside sulfate and GM1 ganglioside hydrolyses had been noted previously, the chromatographic fractions were also examined for activators in other glycosphingolipid hydrolase systems. There was coincidence of activators for the GM1 ganglioside/beta-galactosidase and the globotriaosylceramide/alpha-galactosidase A reactions with the cerebroside sulfatase activator in control urine fractions, and the patients' urines were deficient in activator activities for the three reactions. Identity of the three activators was suggested and antiserum to purified GM1 ganglioside activator was used to test this possibility. There were depressed levels of cross-reacting material in fractions of patient urines by Ouchterlony double diffusion and in unfractionated urine by enzyme-linked immunosorbent assay. Purified activators for the cerebroside sulfate and GM1 ganglioside systems showed lines of identity with no spurring on Ouchterlony double diffusion, identical mobility on immunoelectrophoresis, and similar stimulatory activities toward hydrolysis of the three glycosphingolipid species by their respective enzymes. Finally, the three activator activities were retained by anti-GM1-activator IgG coupled to Sepharose 4B. The results suggest strongly that the same protein entity serves as activator for the enzymatic hydrolysis of cerebroside sulfate, GM1 ganglioside, and globotriaosylceramide.

Animals↗
PubMed1985

Assignment of the gene for human DNA polymerase alpha to the X chromosome.

We have applied an assay based on a monoclonal antibody that discriminates the activity of human DNA polymerase alpha in rodent-human somatic cell hybrid clones to identify a single genetic locus that is both necessary and sufficient for the expression of DNA polymerase alpha. We have mapped this locus to the short arm of the human X chromosome, near the junction of bands Xp21.3 and Xp22.1, and demonstrated that it is not expressed from an inactive X chromosome.

Animals↗
PubMed1985

Familial X-linked ichthyosis, steroid sulfatase deficiency, mental retardation, and nullisomy for Xp223-pter.

Steroid sulfatase (STS)-deficient X-linked ichthyosis was diagnosed in a man with short stature and mental retardation. His generation includes five similarly affected male members. A translocation chromosome is segregating in this Newfoundland kindred. The proband's mother and grandmother have normal skin and are of normal intelligence. From his carrier mother, the proband inherited an X short arm (Xp) to Y long arm (Yq) translocation chromosome, with the entire Y short arm and the X short arm terminal segment deleted (Xp223-pter). His cells are completely deficient in STS activity, confirming assignment of the STS locus to Xp223-pter. Effective management of his ichthyosis included treatment with 6% salicylic acid gel under plastic occlusion and removal of the scales by scrubbing.

Adult↗