Showing papers on "Dosage compensation published in 1987"

Proposed Mechanism of Inheritance and Expression of the Human Fragile-X Syndrome of Mental Retardation

Abstract: A mechanism is proposed for the inheritance and expression of the fragile-X-linked syndrome of mental retardation in humans. Two independent events are required for expression of the syndrome: the fragile-X mutation, and X chromosome inactivation in pre-oogonial cells. The fragile-X mutation at site Xq27 has little or no effect until the chromosome is inactivated in a female as part of the process of dosage compensation. At a stage where the inactivated X chromosome would normally be reactivated in preparation for oogenesis, the mutation results in a local block to the reactivation process. This block to reactivation leads to mental retardation in progeny by reducing the level of products from the unreactivated Xq27 region in male cells, and, for a heterozygous female, in somatic cells in which the normal X chromosome has been inactivated. Published data relevant to this proposed mechanism are discussed.

DNA methylation stabilizes X chromosome inactivation in eutherians but not in marsupials: evidence for multistep maintenance of mammalian X dosage compensation

Abstract: In marsupials and eutherian mammals, X chromosome dosage compensation is achieved by inactivating one X chromosome in female cells; however, in marsupials, the inactive X chromosomes is always paternal, and some genes on the chromosome are partially expressed. To define the role of DNA methylation in maintenance of X chromosome inactivity, we examined loci for glucose-6-phosphate dehydrogenase and hypoxanthine phosphoribosyltransferase in a North American marsupial, the opossum Didelphis virginiana, by using genomic hybridization probes cloned from this species. We find that these marsupial genes are like their eutherian counterparts, with respect to sex differences in methylation of nuclease-insensitive (nonregulatory) chromatin. However, with respect to methylation of the nuclease-hypersensitive (regulatory) chromatin of the glucose-6-phosphate dehydrogenase locus, the opossum gene differs from those of eutherians, as the 5' cluster of CpG dinucleotides is hypomethylated in the paternal as well as the maternal gene. Despite hypomethylation of the 5' CpG cluster, the paternal allele, identified by an enzyme variant, is at best partially expressed; therefore, factors other than methylation are responsible for repression. In light of these results, it seems that the role of DNA methylation in eutherian X dosage compensation is to "lock in" the process initiated by such factors. Because of similarities between dosage compensation in marsupials and trophectoderm derivatives of eutherians, we propose that differences in timing of developmental events--rather than differences in the basic mechanisms of X inactivation--account for features of dosage compensation that differ among mammals.

Carrier Detection in X-Linked Agammaglobulinemia by Analysis of X-Chromosome Inactivation

Abstract: We used a recently developed strategy to analyze patterns of X-chromosome inactivation in human cell populations in order to study female members of families with X-linked agammaglobulinemia--i.e., to detect the carrier state and to test the hypothesis that the disorder results from a defect intrinsic in the development of B cells. According to this strategy, recombinant-DNA probes simultaneously detect restriction-fragment-length polymorphisms and patterns of methylation of X-chromosome genes. Random X-inactivation patterns were observed in isolated peripheral-blood granulocytes, T lymphocytes, and B lymphocytes of women who were not carriers. In contrast, one of the two X chromosomes was preferentially active in the peripheral B cells, but not the T cells or granulocytes, of three carriers of the disorder. This observation strongly supports the hypothesis that X-linked agammaglobulinemia results from an intrinsic defect in B-cell development. Moreover, the analysis described here can be used for direct identification of carriers in families with this disease.

(dC-dA)n.(dG-dT)n sequences have evolutionarily conserved chromosomal locations in Drosophila with implications for roles in chromosome structure and function.

Abstract: In situ hybridization of (dC-dA)n(dG-dT)n to the polytene chromosomes of Drosophila melanogaster reveals a clearly non-random distribution of chromosomal sites for this sequence Sites are distributed over most euchromatic regions but the density of sites along the X chromosome is significantly higher than the density over the autosomes All autosomes show approximately equal levels of hybridization except chromosome 4 which has no detectable stretches of (dC-dA)n(dG-dT)n Another striking feature is the lack of hybridization of the beta-heterochromatin of the chromocenter The specific sites are conserved between different strains of D melanogaster The same overall chromosomal pattern of hybridization is seen for the other Drosophila species studied, including D simulans, a sibling species with a much lower content of middle repetitive DNA, and D virilis, a distantly related species The evolutionary conservation of the distribution of (dC-dA)n(dG-dT)n suggests that these sequences are of functional importance The distribution patterns seen for D pseudoobscura and D miranda raise interesting speculations about function In these species a chromosome equivalent to an autosomal arm of D melanogaster has been translocated onto the X chromosome and acquired dosage compensation In each species the new arm of the X also has a higher density of (dC-dA)n(dG-dT)n similar to that seen on other X chromosomes In addition to correlations with dosage compensation, the depletion of (dC-dA)n(dG-dT)n in beta-heterochromatin and chromosome 4 may also be related to the fact that these regions do not normally undergo meiotic recombination

Dosage Compensation in Drosophila: Evidence That daughterless and Sex-lethal Control X Chromosome Activity at the Blastoderm Stage of Embryogenesis.

Abstract: Dosage compensation is a mechanism that equalizes the expression of X chromosome linked genes in males, who have one X chromosome, with that in females, who have two. In Drosophila, this is achieved by the relative hyperactivation of X-linked genes in males, as was first shown by Muller using a phenotypic assay based on adult eye color. Several genes involved in regulating dosage compensation have been identified through the isolation of mutations that are sex-specific lethals. However, because of this lethality it is not straightforward to assay the relative roles of these genes using assays based on adult phenotypes. Here this problem is circumvented using an assay based on embryonic phenotypes. These experiments indicate that dosage compensation is established early in development and demonstrate that the daughterless and Sex-lethal gene products are involved in regulating X chromosome activity at the blastoderm stage of embryogenesis.

Primary sex determination in the nematode C. elegans.

Abstract: Most nematodes have XO male/XX female sex determination. C. elegans is anomalous, having XX hermaphrodites rather than females. The hermaphrodite condition appears to result from the modification of a basic male/female sex-determination system, which permits both spermatogenesis and oogenesis to occur within a female soma. This modification is achieved by a germ-line-specific control acting at one step in a cascade of autosomal regulatory genes, which respond to X-chromosome dosage and direct male, female, or hermaphrodite development. Mutations of one of these genes can be used to construct artificial strains with ZZ male/WZ female sex determination. Primary sex determination normally depends on the ratio of X chromosomes to autosomes, as in Drosophila, and there appear to be multiple sites on the X chromosome that contribute to this ratio. Also, as in Drosophila, X-chromosome expression is compensated to equalize gene activity in XX and XO animals. Interactions between dosage compensation and sex determination are described and discussed.

Cloning and characterization of a dispersed, multicopy, X chromosome sequence in Drosophila melanogaster.

Abstract: We have isolated and characterized a dispersed middle repetitive DNA sequence from Drosophila melanogaster that is concentrated on the euchromatic portion of the X chromosome. In situ hybridization of the repeat unit to salivary gland chromosomes shows the sequence is distributed among approximately 10 major and 20 minor X chromosomal sites. Based on DNA sequence analysis of homologous sequences from three different cytogenetic regions, the 372-base-pair repeat unit appears to be (A + T)-rich and noncoding and shows strong sequence conservation among units from different chromosomal regions. The nature and distribution of this sequence are suggestive of the hypothetical X chromosome DNA sequences thought to be involved in the primary establishment of sex determination and dosage compensation in Drosophila.

X-linked gene expression and X-chromosome inactivation: marsupials, mouse, and man compared

Abstract: The existence of paternal X inactivation in Australian and American marsupial species suggests that this feature of X-chromosome dosage compensation is not a recent adaptation, but probably predates the evolutionary separation of the Australian and American marsupial lineages. Although it is theoretically possible that the marsupial system is one of random X inactivation with p greater than 0.99 and q less than 0.01 and dependent on parental source, no instance of random X inactivation (p = q or p not equal to q) has ever been verified in any tissue or cell type of any marsupial species. Therefore, we conclude that the most fundamental difference in X inactivation of marsupials and eutherians is whether the inactive X is the paternal one or is determined at random (with p = q in most but not all cases). The only other unequivocal difference between eutherians and marsupials is that both X chromosomes are active in mice and human oocytes, but not in kangaroo oocytes. Apparently, the inactive X is reactivated at a later meiotic stage or during early embryogenesis in kangaroos. X-chromosome inactivation takes place early in embryogenesis of eutherians and marsupials. Extraembryonic membranes of mice exhibit paternal X inactivation, whereas those of humans seem to exhibit random X inactivation with p greater than q (i.e., preferential paternal X inactivation). In general, extraembryonic membranes of marsupial exhibit paternal X inactivation, but the Gpd locus is active on both X chromosomes in at least some cells of kangaroo yolk sac. It is difficult to draw any general conclusion because of major differences in embryogeny of mice, humans, and marsupials, and uncertainties in interpreting the data from humans. Other differences between marsupials and eutherians in patterns of X-linked gene expression and X-chromosome inactivation seem to be quantitative rather than qualitative. Partial expression of some genes on the inactive X is characteristic of marsupials, with species variation in the behavior of specific loci; some X-linked human genes on the inactive chromosome also are known to exhibit partial activity in vivo and in cultured cells. The X chromosomes of marsupials do not behave as units with respect to transcriptional activity, nor does the human X chromosome. In addition, Barr bodies have recently been detected at interphase in some marsupials, establishing that this manifestation of X chromosome inactivity is not restricted to eutherians.(ABSTRACT TRUNCATED AT 400 WORDS)

Molecular genetic aspects of sex determination in Drosophila.

Abstract: Analysis of the mechanisms underlying sex determination and sex differentiation in Drosophila has provided evidence for a complex but comprehensible regulatory hierarchy governing these developmental decisions It is suggested here that the pattern of sexual differentiation and dosage compensation characteristic of the male is a default regulatory state Recent results have provided, in addition, some surprising and intriguing conclusions: (1) that several of the critical controlling genes produce more transcripts than was predicted from the genetic analyses; (2) that setting of the alternative sex-specific states of the doublesex (dsx) locus involves differential transcript processing; and (3) that some aspects of sexual differentation require the prolonged action of certain elements of the regulatory hierarchy These findings are discussed in connection with the current model of sex determination in Drosophila

Poly(dC—dA/dG—dT) repeats in the Drosophila genome: a key function for dosage compensation and position effects?

Abstract: In situ hybridization experiments demonstrate the wide distribution of (CA/GT)n repeats within the genome of Drosophila hydei. (CA/GT)n sequences are evenly distributed in the euchromatin of autosomes and the X chromosome but are not present in most of the heterochromatin of the sex chromosomes. Both sex chromosomes carry one large block of (CA/GT)n. At least part of this (CA/GT)n cluster in the Y chromosome is transcribed in a strandspecific manner and at a high rate in primary spermatocyte nuclei. Also, in polytene chromosomes, specific transcription of (CA/GT)n sequences is found in certain puffs as demonstrated by transcript in situ hybridization. The X chromosomal euchromatin carries approximately twice as much (CA/GT)n over its entire length as the autosomes. These observations are discussed with respect to the mechanisms of dosage compensation and position-effect variegation. A possible biological role of (CA/GT)n sequences, which has been the subject of controversy among various investigators, is their involvement in the control of the rate of transcription.

Genetic analysis of X-chromosome dosage compensation in Caenorhabditis elegans.

Abstract: We have shown that the phenotypes resulting from hypomorphic mutations (causing reduction but not complete loss of function) in two X-linked genes can be used as a genetic assay for X-chromosome dosage compensation in Caenorhabditis elegans between males ( XO) and hermaphrodites (XX). In addition we show that recessive mutations in two autosomal genes, dpy-21 V and dpy-26 IV, suppress the phenotypes resulting from the X-linked hypomorphic mutations, but not the phenotypes resulting from comparable autosomal hypomorphic mutations. This result strongly suggests that the dpy-21 and dpy-26 mutations cause increased X expression, implying that the normal function of these genes may be to lower the expression of X-linked genes. Recessive mutations in two other dpy genes, dpy-22 X and dpy-23 X, increase the severity of phenotypes resulting from some X-linked hypomorphic mutations, although dpy-23 may affect the phenotypes resulting from the autosomal hypomorphs as well. The mutations in all four of the dpy genes show their effects in both XO and XX animals, although to different degrees. Mutations in 18 other dpy genes do not show these effects.

X-Linked Gene Expression in the Virginia Opossum: Differences Between the Paternally Derived Gpd and Pgk-A Loci

Abstract: Expression of X-linked glucose-6-phosphate dehydrogenase (G6PD) and phosphoglycerate kinase-A (PGK-A) in the Virginia opossum ( Didelphis virginiana) was studied electrophoretically in animals from natural populations and those produced through controlled laboratory crosses. Blood from most of the wild animals exhibited a common single-banded phenotype for both enzymes. Rare variant animals, regardless of sex, exhibited single-banded phenotypes different in mobility from the common mobility class of the respective enzyme. The laboratory crosses confirmed the allelic basis for the common and rare phenotypes. Transmission of PGK-A phenotypes followed the pattern of determinate (nonrandom) inactivation of the paternally derived Pgk-A allele, and transmission of G6PD also was consistent with this pattern. A survey of tissue-specific expression of G6PD phenotypes of heterozygous females revealed, in almost all tissues, three-banded patterns skewed in favor of the allele that was expressed in blood cells. Three-banded patterns were never observed in males or in putatively homozygous females. These patterns suggest simultaneous, but unequal, expression of the maternally and paternally derived Gpd alleles within individual cells (i.e., partial paternal allele expression). The absence of such partial expression was noted in a parallel survey of females heterozygous at the Pgk-A locus. Thus, it appears that Gpd and Pgk-A are X-linked in D. virginiana and subject to preferential paternal allele inactivation, but that dosage compensation may not be complete for all paternally derived X-linked genes. The data establish the similarity between the American and Australian marsupial patterns of X-linked gene regulation and, thus, support the hypothesis that this form of dosage compensation was present in the early marsupial lineage that gave rise to these modern marsupial divisions. In addition, the data provide the first documentation of the differential expression of two X-linked genes in a single marsupial species. Because of its combination of X-linked variation, high fecundity, and short generation time, D. virginiana is a unique model for pursuing questions about marsupial gene regulation that have been difficult to approach through studies of Australian species.

Assessment of X Chromosome Dosage Compensation in Caenorhabditis elegans by Phenotypic Analysis of lin-14

Abstract: Caenorhabditis elegans compensates for the difference in X chromosome gene dose between males (XO) and hermaphrodites (XX) through a mechanism that equalizes the levels of X-specific mRNA transcripts between the two sexes. We have devised a sensitive and quantitative genetic assay to measure perturbations in X chromosome gene expression caused by mutations that affect this process of dosage compensation. The assay is based on quantitating the precocious alae phenotype caused by a mutation that reduces but does not eliminate the function of the X-linked gene lin-14. We demonstrate that in diploid animals the lin-14 gene is dosage compensated between XO and XX animals. In XXX diploid animals, however, lin-14 expression is not compensated, implying that the normal dosage compensation mechanism in C. elegans lacks the capacity to compensate completely for the additional X chromosome in triplo-X animals. Using the lin-14 assay we compare the effects of mutations in the genes dpy-21, dpy-26, dpy-27, dpy-28, and dpy-22 on X-linked gene expression. Additionally, in the case of dpy-21 we correlate the change in phenotypic expression of lin-14 with a corresponding change in the lin-14 mRNA transcript level.

Upstream sequences of dosage-compensated and non-compensated alleles of the larval secretion protein gene Sgs-4 in Drosophila.

Abstract: The X chromosomal gene Sgs-4 coding for a larval secretion protein of Drosophila melanogaster is expressed stage and tissue specifically and is hyperexpressed in male larvae of most Drosophila stocks which show dosage compensation. We analysed three Sgs-4 alleles which differ in the size of their coding region, in the intensity of their expression and in the level of dosage compensation. The size and amount of Sgs-4 proteins directly reflect those of RNAs. Different RNA sizes result from different numbers of 21 bp repeats within the structural genes. We sequenced 2.8 kb of DNA upstream of the transcription initiation site of the three alleles. Sequences known to be essential for correct gene expression were located. The only difference within DNA sequences from −1 to −1200 between two alleles with different degrees of expression and differing in dosage compensation is a C to T transition at −344 within a supposed consensus sequence for ecdysone receptor complex binding (ECR). This mutation is partly located within a region of dyad symmetry. Alleles with identical expression show identical mutations within a GTT-rich region at −1.2 kb, but differ within a GT-rich region at −2.0 kb. A polyadenylated 0.5 kb RNA was found to be transcribed from the GTT-rich region of the strand opposite to that of Sgs-4. The corresponding gene is active only in larval salivary glands and, therefore, is named gland specific gene, gsg.

Regulatory sequences of the Sgs-4 gene of Drosophila melanogaster analysed by P element-mediated transformation

Abstract: The X chromosomally located allele Sgs-4 c for a larval secretion protein of Drosophila melanogaster is normally expressed in female larvae of the strain Oregon R and is hyperexpressed in male larvae exhibiting dosage compensation; the allele Sgs-4 d in the strain Samarkand is weakly expressed and is not hyperexpressed in male larvae showing a dosage effect. P element-mediated transformation of upstream DNA sequences from both alleles combined with Sgs-4 d coding and downstream sequences was performed to localize sequences which are responsible for the level of gene expression and for hyperexpression of Sgs-4 c in male larvae. Our results demonstrate that weak expression and dosage effect are inherited with the upstream region from −1 to −838. This Samarkand fragment differs from the homologous Oregon R region only by a C to T transiion at −344 which lies within an assumed binding sequence for the ecdysone receptor complex of dyad base symmetry. Replacing the Samarkand upstream region from −1 to −838 by the Oregon R region restores normal Sgs-4 expression and dosage compensation. Hyperexpression in male larvae displays high sensitivity to position effect and is nearly completely inhibited in one transformed line under heterozygous conditions. The integration of an Sgs-4 d transposon into a weak spot of polytene chromosome 2L results in a decrease in gene expression. The GTT- and GT-rich regions at −1.2 and −2.0 kb do not obviously influence Sgs-4 expression but possibly play a role in induction of stage-specific chromosome puffing.

Structural organization of the alpha-amylase gene locus in Drosophila melanogaster and Drosophila miranda.

Abstract: Chromosomal sites belonging to the alpha-amylase gene family have been identified in D. melanogaster and D. miranda and in the sibling species of miranda, pseudoobscura, and persimilis. Two sites occur in chromosome 2 of melanogaster; one contains the Amy gene locus (54A) and the other an amylase "pseudogene" (53CD). Two sites of homology exist at 73A and 78C and perhaps another at 81BC in chromosome 3 of pseudoobscura and persimilis and in the homologous regions of the X2 chromosome in miranda. The active Amy locus is apparently at 73A. The structural organization of cloned sequences from this multigene family in melanogaster and miranda is under analysis, with emphasis on the functional Amy gene region. Electrophoretic variants of amylase have served as invaluable tools in these studies. For melanogaster, their use as genetic markers enabled us to positively identify our lambda Dm65 clone of the Amy locus and to show that it contains two functional copies of the structural gene for alpha-amylase. Amylase isozymes are now being used in P element-mediated transformation experiments aimed at defining regulatory elements for the temporal and spatial control of amylase expression during development and in response to dietary glucose. In miranda, electrophoretic variants of amylase were useful in assigning the Amy locus to chromosome X2, and they continue to serve as essential markers in our study of the evolution of dosage compensation for amylase expression in males of this species. Restriction maps of the Amy locus in 7 strains of D. melanogaster indicate that despite the worldwide origins of the chromosome samples, all contain a duplication of the amylase structural gene at this locus regardless of whether they produce two alpha-amylase isozymes, a single variant, or none. We have aligned these maps with the genetic and cytological maps of chromosome 2R in melanogaster and assigned alleles for different amylase isozymes to either the proximal or distal Amy gene copy in a number of strains. Restriction site polymorphism is relatively limited at the Amy locus, but some strain-specific rearrangements exist. The locus of two strains with reduced amylase activity, Amy1 (CA 1) and Amy "null", contain anomalies--an insertion in the former and an inversion in the latter. Causal relationships are being sought between the level of amylase expression in these strains and the position of their respective anomalies.(ABSTRACT TRUNCATED AT 400 WORDS)

Differential methylation of the ornithine carbamoyl transferase gene on active and inactive mouse X chromosomes.

Abstract: Ornithine carbamoyl transferase (Oct) is an X-linked gene which exhibits tissue-specific expression. To determine whether methylation of specific CpG sequences plays a role in dosage compensation or tissue-specific expression of the gene, 13 potentially methylatable sites were identified over a 30-kilobase (kb) region spanning from approximately 15 kb upstream to beyond exon II. Fragments of the Mus hortulanus Oct gene were used as probes to establish the degree of methylation at each site. By considering the methylation status in liver (expressing tissue) versus kidney (nonexpressing tissue) from male and female mice, the active and inactive genes could be investigated on active and inactive X-chromosome backgrounds. One MspI site, 12 kb 5' of the Oct-coding region, was cleaved by HpaII in liver DNA from males but not in kidney DNA from males and thus exhibited complete correlation with tissue-specific expression of the gene. Six other sites showed partial methylation, reflecting incomplete correlation with tissue-specific expression.

Molecular analysis of X chromosome dosage compensation in Caenorhabditis elegans

Abstract: We used a convenient quantitative dot blot assay to measure transcript levels for two X chromosome-linked genes, myo-2 and act-4, in the nematode Caenorhabditis elegans. We show that there is dosage compensation of transcript levels for these two genes between XX hermaphrodites and X0 males and that a mutation in the dpy-21 gene, postulated from genetic analysis to be involved in control of X chromosome expression, can affect these transcript levels in the manner predicted. However, we observe the dpy-21 effects only at some stages of the life cycle and not at others. These results are generally consistent with earlier genetic and molecular evidence.

Conservation in toto of the mammalian X-linkage group as a frozen accident

Abstract: It now seems that the X-linkage group has indeed been conserved in toto throughout evolution of not only placental mammals but also of marsupials as well (Ohno et al.,1964a). The unique dosage compensation mechanism that relies on inactivation of one or the other X in females (Lyon,1961) has often been invoked as the cause of this extreme evolutionary conservation. I see no merit in this widely held belief. There apparently exists no dosage compensation mechanism for avian Z-linked genes. In fact, avian species seem to expolit dosage effects of their Z-linked genes for the manifestation of sexual dimorphism. I had a personal experience with a flock of pigeons. A single dose of a particular Z-linked mutant gene in hemizygous ZW females merely diluted the plumage color to creamy yellow, whereas double dose of the same gene in homozygous males made them almost white. In fact, hemizyous females were phenotypic eqivalents of heterozygous males. Similarly in the barred Plymouth Rock breed of chickens, roosters appear considerably lighter in color than hens, for the double dose of Z-linked Barred caused white bands in individual feathers to be much wider than white bands of hemizygous females. Yet the Z-linkage group of birds has been conserved as dilligently as the mammalian X-linkage group (Ohno et al,1964b, Baverstock et al,1982).