Showing papers on "Dosage compensation published in 1994"

A dosage sensitive locus at chromosome Xp21 is involved in male to female sex reversal

Abstract: Male to female sex reversal has been observed in individuals with duplications of the short arm of the X chromosome. Here we demonstrate that sex reversal results from the presence of two active copies of an Xp locus rather than from its rearrangement and that alterations at this locus constitute one of the causes of sex reversal in individuals with a normal 46,XY karyotype. We have named this locus DSS (Dosage Sensitive Sex reversal) and localized it to a 160 kilobase region of chromosome Xp21, adjacent to the adrenal hypoplasia congenita locus. The identification of male individuals deleted for DSS suggests that this locus is not required for testis differentiation. We propose that DSS has a role in ovarian development and/or functions as a link between ovary and testis formation.

Acetylated histone H4 on the male X chromosome is associated with dosage compensation in Drosophila.

Abstract: Dosage compensation in Drosophila occurs by an increase in transcription of genes on the X chromosome in males. This elevated expression requires the function of at least four loci, known collectively as the male-specific lethal (msl) genes. The proteins encoded by two of these genes, maleless (mle) and male-specific lethal-1 (msl-1), are found associated with the X chromosome in males, suggesting that they act as positive regulators of dosage compensation. A specific acetylated isoform of histone H4, H4Ac16, is also detected predominantly on the male X chromosome. We have found that MLE and MSL-1 bind to the X chromosome in an identical pattern and that the pattern of H4Ac16 on the X is largely coincident with that of MLE/MSL-1. We fail to detect H4Ac16 on the X chromosome in homozygous msl males, correlating with the lack of dosage compensation in these mutants. Conversely, in Sxl mutants, we detect H4Ac16 on the female X chromosomes, coincident with an inappropriate increase in X chromosome transcription. These data suggest that synthesis or localization of H4Ac16 is controlled by the dosage compensation regulatory hierarchy. Dosage compensation may involve H4Ac16 function, potentially through interaction with the product of the msl genes.

An SRY-related sequence on the marsupial X chromosome : implications for the evolution of the mammalian testis-determining gene

Abstract: The SRY gene on the human, mouse, and marsupial Y chromosomes is the testis-determining gene that initiates male development in mammals. The SRY protein has a DNA-binding domain (high mobility group or HMG box) similar to those found in the high-mobility-group proteins. SRY is specific for the Y chromosome, but many autosomal genes have been identified that possess a similar HMG box region; those with the most closely SRY-related box regions form a gene family now referred to as SOX genes. We have identified a sequence on the marsupial X chromosome that shares homology with SRY. Sequence comparisons show near-identity with the mouse and human SOX3 gene (formerly called a3), the SOX gene which is the most closely related to SRY. We suggest here that the highly conserved X chromosome-linked SOX3 represents the ancestral SOX gene from which the sex-determining gene SRY was derived. In this model SOX3/SRY divergence and the acquisition of a testis-determining role by SRY might have preceded (and initiated) sex chromosome differentiation or, alternatively, might have been a consequence of X chromosome-Y chromosome differentiation initiated at the locus of an original sex-determining gene(s), later superseded by SRY.

The human X-inactivation centre is not required for maintenance of X-chromosome inactivation.

Abstract: X-chromosome inactivation occurs early in mammalian female development to achieve dosage compensation with males. Although it is widely accepted that this inactivation requires the presence in cis of the X-inactivation centre (XIC), it is not known whether the XIC is required for the initiation, promulgation or maintenance of X inactivation. The XIST gene, which is localized within the XIC interval on both the human and mouse X chromosomes, is constitutively expressed from inactive X chromosomes, suggesting a possible role in the maintenance of X inactivation. To address whether the presence of the XIC, including the XIST gene, is continuously required for the maintenance of X-chromosome inactivation, we have analysed the transcriptional activity of a number of X-linked genes in mouse/human somatic cell hybrids retaining an intact human inactive X chromosome or derivatives of the inactive X chromosome lacking the XIC. Genes subject to X inactivation remain transcriptionally silent despite the loss of the XIC, demonstrating that the presence of the XIC is not required for the maintenance of X inactivation in somatic cells.

Trans-acting dosage effects on the expression of model gene systems in maize aneuploids.

Abstract: The reduction in vigor of aneuploids was classically thought to be due to the imbalance of gene products expressed from the varied chromosome relative to those from the remainder of the genome. In this study, the dosage of chromosomal segments was varied, but the transcript level of most genes encoded therein showed compensation for the number of copies of the gene. Genes whose dosage was not altered were affected by aneuploidy of unlinked chromosomal segments. The phenotypic effects of aneuploidy and of a substantial fraction of quantitative variation are hypothesized to be the consequence of an altered dosage-sensitive regulatory system.

Sex determination and dosage compensation: Lessons from flies and worms

Abstract: In both Drosophila melanogaster and Caenorhabditis elegans somatic sex determination, germline sex determination, and dosage compensation are controlled by means of a chromosomal signal known as the X:A ratio. A variety of mechanisms are used for establishing and implementing the chromosomal signal, and these do not appear to be similar in the two species. Instead, the study of sex determination and dosage compensation is providing more general lessons about different types of signaling pathways used to control alternative developmental states of cells and organisms.

Sex-specific regulation of the male-specific lethal-1 dosage compensation gene in Drosophila

Abstract: Dosage compensation in Drosophila occurs by a twofold increase in transcription per copy of X-linked genes in males (XY) compared with females (XX). msl-1 is one of four genes that are essential for dosage compensation in males, and MSL-1 protein is associated specifically with the male X chromosome. To explore the basis for the sex specificity of dosage compensation, we examined MSL-1 expression in males, females, and dosage compensation mutants. MSL-1 protein levels are negatively regulated by Sxl in females, resulting in male-specific expression of MSL-1. In addition, msl-2 is required for translation and/or stability of MSL-1 in males. Furthermore, the wild-type pattern of MSL-1 localization to the X chromosome is dependent on mle and msl-3 function, although a subset of sites are stained with MSL-1 antibodies in these mutants. Collectively, these data provide the first evidence for an order of msl gene function and suggest that male-specific expression of MSL-1 plays a key role in the sex specificity of dosage compensation.

The dpy-30 gene encodes an essential component of the Caenorhabditis elegans dosage compensation machinery.

Abstract: The need to regulate X chromosome expression in Caenorhabditis elegans arises as a consequence of the primary sex-determining signal, the X/A ratio (the ratio of X chromosomes to sets of autosomes), which directs 1X@A animals to develop as males and 2X/2A animals to develop as hermaphrodites. C. elegans possesses a dosage compensation mechanism that equalizes X chromosome expression between the two sexes despite their disparity in X chromosome dosage. Previous genetic analysis led to the identification of four autosomal genes, dpy-21, dpy-26, dpy-27 and dpy-28, whose products are essential in XX animals for proper dosage compensation, but not for sex determination. We report the identification and characterization of dpy-30, an essential component of the dosage compensation machinery. Putative null mutations in dpy-30 disrupt dosage compensation and cause a severe maternal-effect, XX-specific lethality. Rare survivors of the dpy-30 lethality are dumpy and express their X-linked genes at higher than wild-type levels. These dpy-30 mutant phenotypes superficially resemble those caused by mutations in dpy-26, dpy-27 and dpy-28; however, detailed phenotypic analysis reveals important differences that distinguish dpy-30 from these genes. In contrast to the XX-specific lethality caused by mutations in the other dpy genes, the XX-specific lethality caused by dpy-30 mutations is completely penetrant and temperature sensitive. In addition, unlike the other genes, dpy-30 is required for the normal development of XO animals. Although dpy-30 mutations do not significantly affect the viability of XO animals, they do cause them to be developmentally delayed and to possess numerous morphological and behavioral abnormalities. Finally, dpy-30 mutations can dramatically influence the choice of sexual fate in animals with an ambiguous sexual identity, despite having no apparent effect on the sexual phenotype of otherwise wild-type animals. Paradoxically, depending on the genetic background, dpy-30 mutations cause either masculinization or feminization, thus revealing the complex regulatory relationship between the sex determination and dosage compensation processes. The novel phenotypes caused by dpy-30 mutations suggest that in addition to acting in the dosage compensation process, dpy-30 may play a more general role in the development of both XX and XO animals.

The SMC Family: Novel Motor Proteins for Chromosome Condensation? Midreview

Abstract: Craig L. Peterson Program in Molecular Medicine and Department of Biochemistry and Molecular Biology University of Massachusetts Medical Center Worcester, Massachusetts 01605 As eukaryotic cells progress from interphase to mitosis, they undergo a programmed series of morphological events that culminate in the faithful separation of each chromosome pair and in the formation of two individual cells. One of the most dramatic events that occurs during this process is the compaction of interphase chromatin to generate the familiar metaphase chromosome. Although several groups over the past 30 years have proposed mod- els that describe how chromatin fibers are organized in these condensed structures, less work has focused on the biochemical steps that generate such compaction. Re- cently, a protein family has been identified whose mem- bers may directly facilitate prokaryotic and eukaryotic chromosome condensation. In this issue of Cell, Hirano and Mitchison (1994) describe a biochemical analysis of two family members, XCAP-C and XCAP-E, that are re- quired for mitotic chromosome condensation in axenopus in vitro system. Similarly, Saka et al. (1994) recently showed that two fission yeast family members, cut3 and cutl4, are required for chromosome condensation and segregation in vivo. Although many family members may play a general role in chromosome compaction, Chuang et al. (1994 [this issue of Cell]) describe a Caenorhabditis elegans family member, DPY-27, that regulates dosage compensation presumably by catalyzing the compaction of only the X chromosomes of XX hermaphrodites. The SMC Protein Family This fami@of putative compaction proteins currently con- tains eight members: budding yeast SMCl (Strunnikov et al., 1993) and SMCP (cited in Hirano and Mitchison, 1994), Mycoplasma hyorhinis 115~ (Notarnicola et al., 1991), fis- sion yeast cut3 and cut14 (Saka et al., 1994), Xenopus XCAP-E and XCAP-C (Hirano and Mitchison, 1994), and 6. elegans DPY-27 (Chuang et al., 1994). Two genes, one from the purple bacterium Rhodospirillum rubrum (Falk and Walker, 1988) and one from mouse (Varnum et al., 1991), have onl,y been partially sequenced, but they proba- bly encode additional family members. These proteins are all quite large (- 1000-1200 amino acids), and they share a common head-rod-tail structural organization (Figure 1). The head, which’ contains an NTP-binding domain, and the tail, whfch contains a helix-loop-helix motif, are con- served among ‘ all family members (360/o-49% identity in a 59 residue window surrounding the NTP-binding domain and 29%-48% identity in a 79 residue window sur- rounding the helix-loop-helix element, compared with bud- ding yeast SMCl). SMC2, XCAP-E, c:utl4 are more related to each other than to other family Imembers, with >70% identity in the NTP-binding and helix-loop-helix do- mains, indicating that they may define a distinct subfamily. The central rod domain, which comprises >50% of each protein, is conserved at the structural level, containing two long stretches of

The severe phenotype of females with tiny ring X chromosomes is associated with inability of these chromosomes to undergo X inactivation.

Abstract: Mental retardation and a constellation of congenital malformations not usually associated with Turner syndrome are seen in some females with a mosaic 45,X/46,X,r(X) karyotype. Studies of these females show that the XIST locus on their tiny ring X chromosomes is either not present or not expressed. As XIST transcription is well correlated with inactivation of the X chromosome in female somatic cells and spermatogonia, nonexpression of the locus even when it is present suggests that these chromosomes are transcriptionally active. We examined the transcriptional activity of ring X chromosomes lacking XIST expression (XISTE-), from three females with severe phenotypes. The two tiny ring X chromosomes studied with an antibody specific for the acetylated isoforms of histone H4 marking transcribed chromatin domains were labeled at a level consistent with their being active. We also examined tow of the XISTE- ring chromosomes to determine whether genes that are normally silent on an inactive X are expressed from these chromosomes. Analyses of hybrid cells show that TIMP, ZXDA, and ZXDB loci on the proximal short arm, and AR and PHKA1 loci on the long arm, are well expressed from the tiny ring X chromosome lacking XIST DNA. Studies of the ring chromosome that has XIST DNA but does not transcribe it show that its AR allele is transcribed along with the one on the normal X allele.(ABSTRACT TRUNCATED AT 250 WORDS)

The Sex-lethal amino terminus mediates cooperative interactions in RNA binding and is essential for splicing regulation.

Abstract: Sex-lethal (Sxl) acts as a binary switch that regulates Drosophila sexual differentiation and dosage compensation and also maintains a stable female state through autoregulation. As part of a cascade of genes that are regulated by sex-specific splicing, Sxl controls the sex-specific splicing of transformer (tra) RNA and also its own RNA. Sxl contains two RNP-CS (RNA-binding) domains and is known to bind tra pre-mRNA near the alternative 3' splice site, thus blocking use of that site to give the female-specific splicing pattern. Here, we test how Sxl protein interacts with Sxl RNA during autoregulation. We show that Sxl not only binds Sxl pre-mRNA near the alternative 3' splice site but also at distant, multiple sites surrounding the Sxl alternative exon. Moreover, Sxl binds cooperatively at these multiple sites. The Sxl amino terminus is essential for the cooperative interaction and is also required for regulatory activity in vivo. It appears that this region of Sxl protein, which resembles regions in some other RNA-binding proteins, is a domain that mediates protein-protein interactions during RNA binding and plays an important role in splicing regulation.

DNA replication analysis of FMR1, XIST, and factor 8C loci by FISH shows nontranscribed X-linked genes replicate late.

Abstract: The relationship between the transcriptional state of a locus and the time when it replicates during DNA synthesis is increasingly apparent. Active autosomal genes tend to replicate early, whereas inactive ones are more permissive and frequently replicate later. Although the inactive X chromosome replicates later than its active homologue, little is known about the replication of X-linked genes. We have used FISH to examine the replication of loci on the active X chromosome that are not transcribed, either because the tissue analyzed was not the expressing tissue (F8C), because the locus is silent on all active X chromosomes (XIST), or because it has been mutated by expansion and methylation of a CpG island (FMR1). In this assay, an unreplicated locus is characterized by a single hybridization signal, and a replicated locus is characterized by a doublet hybridization signal. The percentage of doublets is used as a measure of relative time of replication in S phase. The validity of this approach has been established elsewhere, since results compare favorably with those obtained using traditional methods for studying DNA replication. Our results show that the FMR1 gene replicates relatively later in fragile X (fraX) males with the full mutation than in normal males, irrespective of the probe used. The F8C locus is late replicating in both normal and fraX males and replicates at nearly the same time on active and inactive X in females. The XIST locus replicates late in all the males studied and asynchronously in female cells.(ABSTRACT TRUNCATED AT 250 WORDS)

The Lighten up (Lip) gene of Drosophila melanogaster, a modifier of retroelement expression, position effect variegation and white locus insertion alleles.

Abstract: We are interested in identifying single gene mutations that are involved in trans-acting dosage regulation in order to understand further the role of such genes in aneuploid syndromes, various types of dosage compensation as well as in regulatory mechanisms. The Lighten up (Lip) gene in Drosophila melanogaster was identified in a mutagenic screen to detect dominant second site modifiers of white-blood (wbl), a retrotransposon induced allele of the white eye color locus. Lip specifically enhances the phenotype of wbl as well as a subset of other retroelement insertion alleles of white, including the copia-induced allele, white-apricot (wa), and six alleles caused by insertion of I elements. We isolated six alleles of Lip which are all recessive lethal, although phenotypically additive heteroallelic escapers were recovered in some combinations. Lip also suppresses position effect variegation, indicating that it may have a role in chromatin configuration. Additionally, Lip modifies the total transcript abundance of both the blood and copia retrotransposons, having an inverse effect on the steady state level of blood transcripts, while showing a non-additive effect on copia RNA.

Allele-specific gene expression in mammals: the curious case of the imprinted RNAs.

Abstract: Mammals exhibit two epigenetic phenomena whose consequence is the silencing of one of two wild-type copies of a gene X chromosome inactivation in females and parental imprinting of selected autosomal genes. As the mechanisms underlying these forms of gene dosage control are being elucidated, some striking similarities between them are being revealed. Of these similarities, none is more curious than the involvement of two noncoding RNAs in these processes, Xist, in the case of X chromosome inactivation, and H19, an imprinted RNA on chromosome 7. This article will explore the potential evolutionary relationship between X chromosome inactivation and imprinting by focusing on these unusual RNAs.

CpG island promoter region methylation patterns of the inactive-X-chromosome hypoxanthine phosphoribosyltransferase (Hprt) gene.

Abstract: Inactive-X-chromosome genes in mammalian females have methylated CpG islands. We have questioned whether there are variable levels of cytosine methylation at different CpG sites within the island that might indicate the presence of primary sites of methylation which may be critical for the maintenance of gene repression and candidate sites for the initiation of inactivation. To address these questions, we have analyzed the methylation patterns of 32 CpG sites of the X-linked hypoxanthine phosphoribosyltransferase (Hprt) gene on the active and inactive X chromosomes of mouse tissues and cell lines, using genomic sequencing of bisulfite-treated genomic DNA. Cytosine is deaminated by bisulfite, but methylcytosine is not affected. Cell lines that were heterozygous for the Hprt deletion mutation (Hprtb-m3) and a functional Hprt allele were selected with 6-thioguanine. The resulting cell populations uniformly carry the intact Hprt allele on the inactive X chromosome. The methylation of these CpG sites was determined either by the direct sequence analysis of bisulfite-treated and amplified DNA or by the sequence analysis of clones derived from the amplified DNA. No CpG methylation was detected on the active Hprt genes from either males or the active X chromosome of females. On average, 22 CpGs were methylated in the other 50% of female DNA, and the level of methylation at individual sites varied from 42 to 100%. Analysis of the inactive Hprt gene in two cell lines showed that averages of 14 and 18 CpGs were methylated and that the frequency of methylation at 32 individual sites ranged from 3 to 100%. The highest frequency of methylation in cell lines coincided with the sequences flanking transcription initiation sites. These results suggest that methylation patterns are heterogeneous within a tissue and even in clonal cell populations and that specific subsets of CpG sites sustain high methylation frequencies which may be critical for the maintenance of X-chromosome inactivation. The bisulfite method identified which CpG sites were methylated on the inactive X chromosome, and it provided a quantitative estimate of the frequency of methylation of these sites in genomic DNA.

Identification of X chromosome regions in Caenorhabditis elegans that contain sex-determination signal elements.

Abstract: The primary sex-determination signal of Caenorhabditis elegans is the ratio of X chromosomes to sets of autosomes (X/A ratio). This signal coordinately controls both sex determination and X chromosome dosage compensation. To delineate regions of X that contain counted signal elements, we examined the effect on the X/A ratio of changing the dose of specific regions of X, using duplications in XO animals and deficiencies in XX animals. Based on the mutant phenotypes of genes that are controlled by the signal, we expected that increases (in males) or decreases (in hermaphrodites) in the dose of X chromosome elements could cause sex-specific lethality. We isolated duplications and deficiencies of specific X chromosome regions, using strategies that would permit their recovery regardless of whether they affect the signal. We identified a dose-sensitive region at the left end of X that contains X chromosome signal elements. XX hermaphrodites with only one dose of this region have sex determination and dosage compensation defects, and XO males with two doses are more severely affected and die. The hermaphrodite defects are suppressed by a downstream mutation that forces all animals into the XX mode of sex determination and dosage compensation. The male lethality is suppressed by mutations that force all animals into the XO mode of both processes. We were able to subdivide this region into three smaller regions, each of which contains at least one signal element. We propose that the X chromosome component of the sex-determination signal is the dose of a relatively small number of genes.

Isolation of new genes in distal Xq28: transcriptional map and identification of a human homologue of the ARD1 N-acetyl transferase of Saccharomyces cerevisiae

Abstract: In this paper, we describe the physical and transcriptional organization of a region of 140 kb in Xq28, 5' to the L1CAM gene. By isolation and mapping of CpG islands to the physical map of the region, isolation of cDNAs, determination of partial nucleotide sequences and study of the pattern of expression and of the orientation of the transcripts identified we have established a transcriptional map of this region. In this map, previously identified genes (L1CAM, V2R, HCF1 and RnBP) have been positioned as well as 3 new genes. All genes in the region are rather small, ranging in size from 2 to 30 kb, and very close to one another. With the exception of the V2R gene, they are housekeeping, have a CpG island at their 5' end and the same orientation of transcription. This kind of organization is consistent with the one previously described for the more distal portion of Xq28, between the Color Vision (CV) and the G6PD genes and indicates that genes with housekeeping and tissue specific pattern of expression are interspersed in the genome but they are probably found in different 'transcriptional domains'. Among the new genes, TE2 demonstrated 40% identity with the protein N-acetyl transferase ARD1 of S. cerevisiae: TE2 may be the human homologue of the S. cerevisiae gene.

Expression of an X-linked HMG-lacZ transgene in mouse embryos: implication of chromosomal imprinting and lineage-specific X-chromosome activity.

Abstract: X-chromosome activity in female mouse embryos was studied at the cellular level using an X-linked lacZ transgene which encodes beta-galactosidase (beta-Gal). Translation of maternal RNA in oocytes is seen as beta-Gal activity that persists into early cleavage-stages. Zygotic transcription of the transgene from the maternal X chromosome (Xm) is first found at about the 8-cell stage. By contrast, expression of the lacZ transgene on the paternal X chromosome (Xp) is not seen until later at the 16-32-cell stage. Preferential inactivation of Xp occurs in the mural trophectoderm, the primitive endoderm, and derivatives of the polar trophectoderm, but a small number of cells in these lineages may still retain an active paternal X chromosome. X inactivation begins at 3.5 days in the inner cell mass but contrary to previous findings the process is not completed in the embryonic ectoderm by 5.5 to 6.0 days. Regional variation in beta-Gal activity is also observed in the embryonic ectoderm during gastrulation which may be related to the specification of cell fates. Random inactivation of Xp and Xm ensues in all somatic tissues but the process is completed at different times in different tissues. The slower progression of X inactivation in tissues such as the notochord, the heart, and the embryonic gut is primarily due to the persistent maintenance of two active X chromosomes in a significant fraction of cells in these tissues. Recent findings on the methylation of endogenous X-linked genes suggest that the prolonged expression of beta-Gal might also be due to the different rate of spreading of inactivation along the X chromosome to the lacZ transgene locus in different tissues.

Xq-Yq interchange resulting in supernormal X-linked gene expression in severely retarded males with 46,XYq- karyotype.

Abstract: The critical importance of dosage compensation is underscored by a novel human syndrome (“XYXq syndrome”) in which we have detected partial X disomy, demonstrated supernormal gene expression resulting from the absence of X inactivation, and correlated this overexpression with its phenotypic consequences. Studies of three unrelated boys with 46,XYq- karyotypes and anomalous phenotypes (severe mental retardation, generalized hypotonia and microcephaly) show the presence of a small portion of distal Xq on the long arm of the Y derivative. Cells from these boys exhibit twice-normal activity of glucose-6-phosphate dehydrogenase, a representative Xq28 gene product. In all three cases, the presence of Xq DMA on a truncated Y chromosome resulted from an aberrant Xq–Yq interchange occurring in the father's germline.

Differential Effects of Sex-Lethal Mutations on Dosage Compensation Early in Drosophila Development

Abstract: In response to the primary sex determination signal, X chromosome dose, the Sex-lethal gene controls all aspects of somatic sex determination and differentiation, including X chromosome dosage compensation. Two complementary classes of mutations have been identified that differentially affect Sxl somatic functions: (1) those impairing the "early" function used to set developmental pathway choice in response to the sex determination signal and (2) those impairing "late" functions involved in maintaining the pathway choice independent of the initiating signal and/or in directing differentiation. This "early vs. late" distinction correlates with a switch in promoter utilization from SxlPe to SxlPm at the blastoderm stage and a corresponding switch from transcriptional to RNA splicing control. Here we characterize five partial-loss-of-function Sxl alleles to explore a distinction between "early vs. late" functioning of Sxl in dosage compensation. Assaying for dosage compensation during the blastoderm stage, we find that the earliest phase of the dosage compensation process is controlled by products of the early Sxl promoter, SxlPe. Hence, in addition to triggering the sexual pathway decision of cells, products derived from SxlPe also control early dosage compensation, the first manifestation of sexually dimorphic differentiation. The effects of mutant Sxl alleles on early dosage compensation are consistent with their previous categorization as early vs. late defective with respect to their effects on pathway initiation. Results reported here suggest that the dosage compensation regulatory genes currently known to function downstream of Sxl, genes known as the "male-specific lethals," do not control all aspects of dosage compensation either at the blastoderm stage or later in development. In the course of this study, we also discovered that the canonical early defective allele, Sxlf9, which is impaired in its ability to establish the female developmental pathway commitment, is likely to be defective in the stability and/or functioning of products derived from SxlPe, rather than in the ability of SxlPe to respond to the chromosomal sex determination signal.

Effects of the maleless mutation on X and autosomal gene expression in Drosophila melanogaster.

Abstract: The mutational effect of the maleless (mle) gene in Drosophila has been reexamined. Earlier work had suggested that mle along with other male-lethal genes was responsible for hypertranscription of the X chromosome in males to bring about dosage compensation. Prompted by studies on dosage sensitive regulatory genes, we tested for effects of mlets on the phenotypes of 16 X or autosomal mutations in adult escapers of lethality. In third instar larvae, prior to the major lethal phase of mle, we examined activities of 6 X or autosomally encoded enzymes, steady state mRNA levels of 15 X-linked or autosomal genes and transcripts from two large genomic segments derived from either the X or from chromosome 2 and present in yeast artificial chromosomes. In contrast to the previously hypothesized role, we detected pronounced effects of mle on the expression of both X-linked and autosomal loci such that a large proportion of the tested genes were increased in expression, while only two X-linked loci were reduced. The most prevalent consequence was an increase of autosomal gene expression, which can explain previously observed reduced X:autosome transcription ratios. These observations suggest that if mle plays a role in the discrimination of the X and the autosomes, it may do so by modification of the effects of dosage sensitive regulatory genes.

Dosage compensation in Drosophila: the X-chromosomal binding of MSL-1 and MLE is dependent on Sxl activity.

Abstract: In Drosophila, dosage compensation, i.e. the equalization of levels of X-linked gene products in the two sexes, is achieved by the hypertranscription of most X-linked genes in males relative to females. The products of at least four genes, collectively termed male-specific lethal (msl) genes, are required for this process and, at least in the case of three of them, mediate this function through an association with the X chromosome in males. We have studied some of the parameters that affect the association of the msl-1 gene product and found that its presence is dependent on the wild-type function of the other three genes, leading to the conclusion that these gene products contribute to the formation of a multi-subunit complex. Furthermore, the X-chromosomal association of the msl-1 and mle gene products is negatively correlated with the level of function of the master regulatory gene Sxl and can assume either a mosaic or a uniform distribution in the tissues of mutant XX individuals. Surprisingly, we also found that the association of these two msl gene products with the two X chromosomes in females of certain mutant genotypes does not result in the hypertranscription of X-linked genes or in any apparent reduction in viability.

Sex determination in Drosophila melanogaster: X-linked genes involved in the initial step of Sex-lethal activation

Abstract: Sex determination is the commitment of an embryo to either the female or the male developmental pathway. The ratio of X chromosomes to sets of autosomes is the primary genetic signal that determines sex in Drosophila, by triggering the functional state of the gene Sex-lethal: in females (2X;2A) Sxl will be ON, whereas in males (X;2A) Sxl will be OFF. Genetic and molecular studies have defined a set of genes involved in the formation of the X:A signal, as well as other genes, with either maternal or zygotic effects, which are also involved in regulating the initial step of Sex-lethal activation. We review these data and present new data on two more regions of the X chromosome that define other genes needed for Sxl activation. In addition, we report on the interaction between some of the genes regulating Sxl activation.

Histone H4 acetylated at lysine 16 and proteins of theDrosophila dosage compensation pathway co-localize on the male X chromosome through mitosis

Abstract: In the fruit flyDrosophila, dosage compensation involves several proteins acting in concert to double the transcriptional activity of genes on the single male X chromosome. Three of these proteins, MLE, MSL-1 and histone H4 acetylated at lysine 16 (H4Ac16), have recently been shown to be located almost exclusively on the male X chromosome in interphase (polytene) cells. We show here that in neuroblasts from third instarDrosophila larvae antisera to H4Ac16, MLE and MSL-1 uniquely label the distal, euchromatic region of the male X chromosome through mitosis. The centromere-proximal, heterochromatic region of the male X is not labelled with these antisera, nor are male autosomes or any chromosomes in female cells. That the association of H4Ac16 with the male X chromosome persists, even when the chromosome is maximally compacted and transcriptionally quiescent, argues that this modified histone is an integral component of the dosage compensation pathway. In the nuclei of interphase neuroblasts from male (but never female) larvae, antibodies to H4Ac16 revealed a small, brightly labelled patch against a background of generally weak nuclear staining. In double-labelling experiments, this patch was also labelled, albeit comparatively weakly, with antibodies to MSL-1. These results strongly suggest that the distal, euchromatic region of the X chromosome in male cells occupies a limited and relatively compact nuclear domain.