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We mapped regulatory loci for nearly all protein-coding genes in mammals using comparative genomic hybridization and expression array measurements from a panel of mouse-hamster radiation hybrid cell lines. The large number of breaks in the mouse chromosomes and the dense genotyping of the panel allowed extremely sharp mapping of loci. As the regulatory loci result from extra gene dosage, we call them copy number expression quantitative trait loci, or ceQTLs. The -2log10P support interval for the ceQTLs was 4, including 13 hotspots each regulating >100 genes in trans. Further, this work identifies 2,761 trans ceQTLs harboring no known genes, and provides evidence for a mode of gene expression autoregulation specific to the X chromosome.

X chromosome inactivation (XCI) serves as a paradigm for RNA-mediated regulation of gene expression, wherein the long non-coding RNA XIST spreads across the X chromosome in cis to mediate gene silencing chromosome-wide. In female naive human pluripotent stem cells (hPSCs), XIST is in a dispersed configuration, and XCI does not occur, raising questions about XIST's function. We found that XIST spreads across the X chromosome and induces dampening of X-linked gene expression in naive hPSCs. Surprisingly, XIST also targets specific autosomal regions, where it induces repressive chromatin changes and gene expression dampening. Thereby, XIST equalizes X-linked gene dosage between male and female cells while inducing differences in autosomes. The dispersed Xist configuration and autosomal localization also occur transiently during XCI initiation in mouse PSCs. Together, our study identifies XIST as the regulator of X chromosome dampening, uncovers an evolutionarily conserved trans -acting role of XIST/Xist, and reveals a correlation between XIST/Xist dispersal and autosomal targeting. [Display omitted] • XIST localizes across the X and spreads to autosomal regions in female naive hPSCs • XIST dampens the expression of X-linked and autosomal genes in female naive hPSCs • XIST mediates chromatin changes at target regions on the X and autosomes • Xist spreads to autosomal regions during XCI initiation in differentiating mouse PSCs During X chromosome dampening, when XIST expression is uncoupled from complete gene silencing and displays a non-typical dispersed configuration, XIST spreads beyond the X chromosome to downregulate gene expression on autosomes. [ABSTRACT FROM AUTHOR]

RNA unwinding by DExH-type helicases underlies most RNA metabolism and function. It remains unresolved if and how the basic unwinding reaction of helicases is regulated by auxiliary domains. We explored the interplay between the RecA and auxiliary domains of the RNA helicase maleless (MLE) from Drosophila using structural and functional studies. We discovered that MLE exists in a dsRNA-bound open conformation and that the auxiliary dsRBD2 domain aligns the substrate RNA with the accessible helicase tunnel. In an ATP-dependent manner, dsRBD2 associates with the helicase module, leading to tunnel closure around ssRNA. Furthermore, our structures provide a rationale for blunt-ended dsRNA unwinding and 3′-5′ translocation by MLE. Structure-based MLE mutations confirm the functional relevance of our model for RNA unwinding. Our findings contribute to our understanding of the fundamental mechanics of auxiliary domains in DExH helicase MLE, which serves as a model for its human ortholog and potential therapeutic target, DHX9/RHA. [Display omitted] • Cryo-EM reveals autoregulatory function of dsRBD2 domain in DExH RNA helicase MLE • Auxiliary domain dsRBD2 aligns dsRNA with MLE's RNA tunnel • MLE unwinds blunt-end dsRNA and translocates from 3′ to 5′ • These findings reveal essential roles of auxiliary domains in MLE/DHX9 function Jagtap et al., combining structural and functional studies, show how the auxiliary domain dsRBD2 in DExH RNA helicase MLE regulates the helicase activity by undergoing large-scale conformation changes. These findings contribute to understanding the fundamental mechanics of auxiliary domains in MLE, which serves as a model for its human ortholog and potential therapeutic target, DHX9. [ABSTRACT FROM AUTHOR]

BackgroundIn animals with heteromorphic sex chromosomes, dosage compensation of sex-chromosome genes is thought to be critical for species survival. Diverse molecular mechanisms have evolved to effectively balance the expressed dose of X-linked genes between XX and XY animals, and to balance expression of X and autosomal genes. Dosage compensation is not understood in birds, in which females (ZW) and males (ZZ) differ in the number of Z chromosomes.ResultsUsing microarray analysis, we compared the male:female ratio of expression of sets of Z-linked and autosomal genes in two bird species, zebra finch and chicken, and in two mammalian species, mouse and human. Male:female ratios of expression were significantly higher for Z genes than for autosomal genes in several finch and chicken tissues. In contrast, in mouse and human the male:female ratio of expression of X-linked genes is quite similar to that of autosomal genes, indicating effective dosage compensation even in humans, in which a significant percentage of genes escape X-inactivation.ConclusionBirds represent an unprecedented case in which genes on one sex chromosome are expressed on average at constitutively higher levels in one sex compared with the other. Sex-chromosome dosage compensation is surprisingly ineffective in birds, suggesting that some genomes can do without effective sex-specific sex-chromosome dosage compensation mechanisms.

BackgroundMost Z chromosome genes in birds are expressed at a higher level in ZZ males than in ZW females, and thus are relatively ineffectively dosage compensated. Some Z genes are compensated, however, by an unknown mechanism. Previous studies identified a non-coding RNA in the male hypermethylated (MHM) region, associated with sex-specific histone acetylation, which has been proposed to be involved in dosage compensation.ResultsUsing microarray mRNA expression analysis, we find that dosage compensated and non-compensated genes occur across the Z chromosome, but a cluster of compensated genes are found in the MHM region of chicken chromosome Zp, whereas Zq is enriched in non-compensated genes. The degree of dosage compensation among Z genes is predicted better by the level of expression of Z genes in males than in females, probably because of better compensation of genes with lower levels of expression. Compensated genes have different functional properties than non-compensated genes, suggesting that dosage compensation has evolved gene-by-gene according to selective pressures on each gene. The group of genes comprising the MHM region also resides on a primitive mammalian (platypus) sex chromosome and, thus, may represent an ancestral precursor to avian ZZ/ZW and monotreme XX/XY sex chromosome systems.ConclusionThe aggregation of dosage compensated genes near the MHM locus may reflect a local sex- and chromosome-specific mechanism of dosage compensation, perhaps mediated by the MHM non-coding RNA.

Polycomb group (PcG) proteins belonging to the polycomb (Pc) repressive complexes 1 and 2 (PRC1 and PRC2) maintain homeotic gene silencing. In Drosophila, PRC2 methylates histone H3 on lysine 27, and this epigenetic mark facilitates recruitment of PRC1. Mouse PRC2 (mPRC2) has been implicated in X inactivation, as mPRC2 proteins transiently accumulate on the inactive X chromosome (Xi) at the onset of X inactivation to methylate histone H3 lysine 27 (H3-K27). In this study, we demonstrate that mPRC1 proteins localize to the Xi, and that different mPRC1 proteins accumulate on the Xi during initiation and maintenance of X inactivation in embryonic cells. The Xi accumulation of mPRC1 proteins requires Xist RNA and is not solely regulated by the presence of H3-K27 methylation, as not all cells that exhibit this epigenetic mark on the Xi show Xi enrichment of mPRC1 proteins. Our results implicate mPRC1 in X inactivation and suggest that the regulated assembly of PcG protein complexes on the Xi contributes to this multistep process.

Abstract Background Heterogametic species display a differential number of sex chromosomes resulting in imbalanced transcription levels for these chromosomes between males and females. To correct this disequilibrium, dosage compensation mechanisms involving gene expression and chromatin accessibility regulations have emerged throughout evolution. In insects, these mechanisms have been extensively characterized only in Drosophila but not in insects of agronomical importance. Aphids are indeed major pests of a wide range of crops. Their remarkable ability to switch from asexual to sexual reproduction during their life cycle largely explains the economic losses they can cause. As heterogametic insects, male aphids are X0, while females (asexual and sexual) are XX. Results Here, we analyzed transcriptomic and open chromatin data obtained from whole male and female individuals to evaluate the putative existence of a dosage compensation mechanism involving differential chromatin accessibility of the pea aphid’s X chromosome. Transcriptomic analyses first showed X/AA and XX/AA expression ratios for expressed genes close to 1 in males and females, respectively, suggesting dosage compensation in the pea aphid. Analyses of open chromatin data obtained by Formaldehyde-Assisted Isolation of Regulatory Elements (FAIRE-seq) revealed a X chromosome chromatin accessibility globally and significantly higher in males than in females, while autosomes’ chromatin accessibility is similar between sexes. Moreover, chromatin environment of X-linked genes displaying similar expression levels in males and females—and thus likely to be compensated—is significantly more accessible in males. Conclusions Our results suggest the existence of an underlying epigenetic mechanism enhancing the X chromosome chromatin accessibility in males to allow X-linked gene dose correction between sexes in the pea aphid, similar to Drosophila. Our study gives new evidence into the comprehension of dosage compensation in link with chromatin biology in insects and newly in a major crop pest, taking benefits from both transcriptomic and open chromatin data.

Abstract Background The ubiquitin protein E3A ligase gene (UBE3A) gene is imprinted with maternal-specific expression in neurons and biallelically expressed in all other cell types. Both loss-of-function and gain-of-function mutations affecting the dosage of UBE3A are associated with several neurodevelopmental syndromes and psychological conditions, suggesting that UBE3A is dosage-sensitive in the brain. The observation that loss of imprinting increases the dosage of UBE3A in brain further suggests that inactivation of the paternal UBE3A allele evolved as a dosage-regulating mechanism. To test this hypothesis, we examined UBE3A transcript and protein levels among cells, tissues, and species with different imprinting states of UBE3A. Results Overall, we found no correlation between the imprinting status and dosage of UBE3A. Importantly, we found that maternal Ube3a protein levels increase in step with decreasing paternal Ube3a protein levels during neurogenesis in mouse, fully compensating for loss of expression of the paternal Ube3a allele in neurons. Conclusions Based on our findings, we propose that imprinting of UBE3A does not function to reduce the dosage of UBE3A in neurons but rather to regulate some other, as yet unknown, aspect of gene expression or protein function.

Theory makes several predictions concerning differences in genetic variation between the X chromosome and the autosomes due to male X hemizygosity. The X chromosome should: (i) typically show relatively less standing genetic variation than the autosomes, (ii) exhibit more variation in males compared to females because of dosage compensation, and (iii) potentially be enriched with sex-specific genetic variation. Here, we address each of these predictions for lifespan and aging in Drosophila melanogaster. To achieve unbiased estimates of X and autosomal additive genetic variance, we use 80 chromosome substitution lines; 40 for the X chromosome and 40 combining the two major autosomes, which we assay for sex-specific and cross-sex genetic (co)variation. We find significant X and autosomal additive genetic variance for both traits in both sexes (with reservation for X-linked variation of aging in females), but no conclusive evidence for depletion of X-linked variation (measured through females). Males display more X-linked variation for lifespan than females, but it is unclear if this is due to dosage compensation since also autosomal variation is larger in males. Finally, our results suggest that the X chromosome is enriched for sex-specific genetic variation in lifespan but results were less conclusive for aging overall. Collectively, these results suggest that the X chromosome has reduced capacity to respond to sexually concordant selection on lifespan from standing genetic variation, while its ability to respond to sexually antagonistic selection may be augmented.

Proteins MSL1 and MSL2 form the core of the Drosophila dosage compensation complex, which specifically binds to the X chromosome of males. Phosphorylation of certain amino acid residues was previously shown to regulate MSL1 activity. In the present work, transgenic lines of Drosophila expressing mutant variants of the MSL1 protein were obtained, in which amino acids undergoing phosphorylation were replaced. As a result, it was shown that inactivation of phosphorylation sites does not affect the efficiency of specific binding of the dosage compensation complex to the X chromosome of males and its functional activity. [ABSTRACT FROM AUTHOR]

Sex chromosomes originate as a pair of homologus autosomes that then follow a general pattern of divergence. This is evident in mammalian sex chromosomes, which have undergone stepwise recombination suppression events that left footprints of evolutionary strata on the X chromosome. The loss of genes on the Y chromosome led to Ohno’s hypothesis of dosage equivalence between XY males and XX females, which is achieved through X-chromosome inactivation (XCI). This process transcriptionally silences all but one X chromosome in each female cell, although 15–30% of human X-linked genes still escape inactivation. There are multiple evolutionary pathways that may lead to a gene escaping XCI, including remaining Y chromosome homology, or female advantage to escape. The conservation of some escape genes across multiple species and the ability of the mouse inactive X to recapitulate human escape status both suggest that escape from XCI is controlled by conserved processes. Evolutionary pressures to minimize dosage imbalances have led to the accumulation of genetic elements that favor either silencing or escape; lack of dosage sensitivity might also allow for the escape of flanking genes near another escapee, if a boundary element is not present between them. Delineation of the elements involved in escape is progressing, but mechanistic understanding of how they interact to allow escape from XCI is still lacking. Although increasingly well-studied in humans and mice, non-trivial challenges to studying escape have impeded progress in other species. Mouse models that can dissect the role of the sex chromosomes distinct from sex of the organism reveal an important contribution for escape genes to multiple diseases. In humans, with their elevated number of escape genes, the phenotypic consequences of sex chromosome aneuplodies and sexual dimorphism in disease both highlight the importance of escape genes.

We previously showed that Guy1, a primary signal expressed from the Y chromosome, is a strong candidate for a male-determining factor that confers female-specific lethality in Anopheles stephensi (Criscione et al., 2016). Here, we present evidence that Guy1 increases X gene expression in Guy1-transgenic females from two independent lines, providing a mechanism underlying the Guy1-conferred female lethality. The median level gene expression (MGE) of X-linked genes is significantly higher than autosomal genes in Guy1-transgenic females while there is no significant difference in MGE between X and autosomal genes in wild-type females. Furthermore, Guy1 significantly upregulates at least 40% of the 996 genes across the X chromosome in transgenic females. Guy1-conferred female-specific lethality is remarkably stable and completely penetrant. These findings indicate that Guy1 regulates dosage compensation in An. stephensi and components of dosage compensation may be explored to develop novel strategies to control mosquito-borne diseases.

Arabidopsis contains hundreds of ribosomal DNA copies organized within the nucleolar organizing regions (NORs) in chromosomes 2 and 4. There are four major types of variants of rDNA, VAR1-4, based on the polymorphisms of 3' external transcribed sequences. The variants are known to be differentially expressed during plant development. We created a mutant by the CRISPR-Cas9-mediated excision of ~ 25 nt from predominantly NOR4 ribosomal DNA copies, obtaining mosaic mutational events on ~ 5% of all rDNA copies. The excised region consists of P-loop and Helix-82 segments of 25S rRNA. The mutation led to allelic, dosage-dependent defects marked by lateral root inhibition, reduced size, and pointy leaves, all previously observed for defective ribosomal function. The mutation in NOR4 led to dosage compensation from the NOR2 copies by elevated expression of VAR1 in mutants and further associated single-nucleotide variants, thus, resulting in altered rRNA sub-population. Furthermore, the mutants exhibited rRNA maturation defects specifically in the minor pathway typified by 32S pre-rRNA accumulation. Density-gradient fractionation and subsequent RT-PCR of rRNA analyses revealed that mutated copies were not incorporated into the translating ribosomes. The mutants in addition displayed an elevated autophagic flux as shown by the autophagic marker GFP-ATG8e, likely related to ribophagy.

lncRNAs, genes transcribed but not translated, longer than 200 nucleotides, are classified as a separate class of nonprotein coding genes. Since their discovery, largely from RNAseq data, a number of pioneer studies have begun to unravel its myriad functions, centered on gene expression regulation, suggesting developmental and evolutionary conservation. Since they do not code for proteins and have no open reading frames, their functional constraints likely differ from that of protein coding genes, or of genes where the majority of the nucleotide sequence is required for function, like tRNAs. This has complicated assessment of both developmental and evolutionary conservation, and the identification of homologs in different species. Here we argue that other characteristics: general synteny and particular chromosomal placement regardless of sequence, sequence micro‐motifs, and secondary structure allow for "homologs" to be identified and compared, confirming developmental and evolutionary conservation of lncRNAs. We conclude exemplifying a case in point: that of the evolutionarily conserved lncRNA acal, characterized and required for embryogenesis in Drosophila. Key Findings: lncRNAs are a heterogeneous class of RNA genes.Homology in lncRNAs relies on several points: synteny, micro‐domains, and secundary structure.Common function may imply common evolutionary origin. [ABSTRACT FROM AUTHOR]

Hundreds of genome-wide association studies were conducted to map the disease genes on autosomes in human beings. It is known that many complex diseases are sex-determined and X chromosome is expected to play an important role. However, only a few single-nucleotide polymorphisms on X chromosome were found to be significantly associated with the diseases under study. On the other hand, to balance the genetic effect between two sexes, X chromosome inactivation occurs in most of X-linked genes by silencing one copy of two X chromosomes in females and dosage compensation is achieved. A few association studies on X chromosome incorporated the information on dosage compensation. However, some of them require the assumption of Hardy-Weinberg equilibrium and some need to specify the underlying genetic model. Therefore, in this article, we propose a novel method for association by taking account of different dosage compensation patterns. The proposed test is a robust approach because it requires neither specifying the underlying genetic models nor the assumption of Hardy-Weinberg equilibrium. Further, the proposed method allows for different deviations from Hardy-Weinberg equilibrium between cases and controls. Simulation results demonstrate that our proposed method generally outperforms the existing methods in terms of controlling the size and the test power. Finally, we apply the proposed test to the meta-analysis of the Graves' disease data for its practical use. [ABSTRACT FROM AUTHOR]

In mammals, dosage compensation of sex chromosomal genes between females (XX) and males (XY) is achieved through X-chromosome inactivation (XCI). The X-linked X-inactive-specific transcript (Xist) long noncoding RNA is indispensable for XCI and initiates the process early during development by spreading in cis across the X chromosome from which it is transcribed. During XCI, Xist RNA triggers gene silencing, recruits a plethora of chromatin modifying factors, and drives a major structural reorganization of the X chromosome. Here, we review our knowledge of the multitude of epigenetic events orchestrated by Xist RNA to allow female mammals to survive through embryonic development by establishing and maintaining proper dosage compensation. In particular, we focus on recent studies characterizing the interaction partners of Xist RNA, and we discuss how they have affected the field by addressing long-standing controversies or by giving rise to new research perspectives that are currently being explored. This review is dedicated to the memory of Denise Barlow, pioneer of genomic imprinting and functional long noncoding RNAs (lncRNAs), whose work has revolutionized the epigenetics field and continues to inspire generations of scientists. [ABSTRACT FROM AUTHOR]

Once recombination is halted between the X and Y chromosomes, sex chromosomes begin to differentiate and transition to heteromorphism. While there is a remarkable variation across clades in the degree of sex chromosome divergence, far less is known about the variation in sex chromosome differentiation within clades. Here, we combined whole-genome and transcriptome sequencing data to characterize the structure and conservation of sex chromosome systems across Poeciliidae, the livebearing clade that includes guppies. We found that the Poecilia reticulata XY system is much older than previously thought, being shared not only with its sister species, Poecilia wingei, but also with Poecilia picta, which diverged roughly 20 million years ago. Despite the shared ancestry, we uncovered an extreme heterogeneity across these species in the proportion of the sex chromosome with suppressed recombination, and the degree of Y chromosome decay. The sex chromosomes in P. reticulata and P. wingei are largely homomorphic, with recombination in the former persisting over a substantial fraction. However, the sex chromosomes in P. picta are completely nonrecombining and strikingly heteromorphic. Remarkably, the profound degradation of the ancestral Y chromosome in P. picta is counterbalanced by the evolution of functional chromosome-wide dosage compensation in this species, which has not been previously observed in teleost fish. Our results offer important insight into the initial stages of sex chromosome evolution and dosage compensation. [ABSTRACT FROM AUTHOR]

Aphids present an ideal system to study epigenetics as they can produce diverse, but genetically identical, morphs in response to environmental stimuli. Here, using whole genome bisulphite sequencing and transcriptome sequencing of the green peach aphid (Myzus persicae), we present the first detailed analysis of cytosine methylation in an aphid and investigate differences in the methylation and transcriptional landscapes of male and asexual female morphs. We found that methylation primarily occurs in a CG dinucleotide (CpG) context and that exons are highly enriched for methylated CpGs, particularly at the 3′ end of genes. Methylation is positively associated with gene expression, and methylated genes are more stably expressed than unmethylated genes. Male and asexual female morphs have distinct methylation profiles. Strikingly, these profiles are divergent between the sex chromosome and the autosomes; autosomal genes are hypomethylated in males compared to asexual females, whereas genes belonging to the sex chromosome, which is haploid in males, are hypermethylated. Overall, we found correlated changes in methylation and gene expression between males and asexual females, and this correlation was particularly strong for genes located on the sex chromosome. Our results suggest that differential methylation of sex‐biased genes plays a role in aphid sexual differentiation. [ABSTRACT FROM AUTHOR]

In species with genetic sex determination, dosage compensation can evolve to equal expression levels of sex-linked and autosomal genes. Current knowledge about dosage compensation has mainly been derived from male-heterogametic (XX/XY) model organisms, whereas less is understood about the process in female-heterogametic systems (ZZ/ZW). In moths and butterflies, downregulation of Z-linked expression in males (ZZ) to match the expression level in females (ZW) is often observed. However, little is known about the underlying regulatory mechanisms, or if dosage compensation patterns vary across ontogenetic stages. In this study, we assessed dynamics of Z-linked and autosomal expression levels across developmental stages in the wood white (Leptidea sinapis). We found that although expression of Z-linked genes in general was reduced compared with autosomal genes, dosage compensation was actually complete for some categories of genes, in particular sex-biased genes, but equalization in females was constrained to a narrower gene set. We also observed a noticeable convergence in Z-linked expression between males and females after correcting for sex-biased genes. Sex-biased expression increased successively across developmental stages, and male-biased genes were enriched on the Z-chromosome. Finally, all five core genes associated with the ribonucleoprotein dosage compensation complex male-specific lethal were detected in adult females, in correspondence with a reduction in the expression difference between autosomes and the single Z-chromosome. We show that tuning of gene dosage is multilayered in Lepidoptera and argue that expression balance across chromosomal classes may predominantly be driven by enrichment of male-biased genes on the Z-chromosome and cooption of available dosage regulators. [ABSTRACT FROM AUTHOR]

The Masculinizer (Masc) gene encodes a CCCH‐tandem zinc finger protein that controls both masculinization and dosage compensation in the silkworm Bombyx mori. We previously measured the masculinizing activity of the lepidopteran Masc proteins using B. mori ovary‐derived cell line BmN‐4. Here, we established an RNA‐seq data‐based assay system in which the level of B. mori Masc (BmMasc)‐induced dosage compensation can be estimated in BmN‐4 cells. Using this system, we found that a cysteine residue at position 301, which was shown to be essential for the masculinizing activity of BmMasc, is also required for dosage compensation. We further investigated the relationships between Masc‐induced cell growth inhibition, masculinizing activity, and the level of dosage compensation, using Masc genes from three lepidopteran insects. In summary, we have established a cell‐based system to monitor levels of Masc‐induced dosage compensation. [ABSTRACT FROM AUTHOR]

Differentiated sex chromosomes are accompanied by a difference in gene dose between X/Z-specific and autosomal genes. At the transcriptomic level, these sex-linked genes can lead to expression imbalance, or gene dosage can be compensated by epigenetic mechanisms and results into expression level equalization. Schistosoma mansoni has been previously described as a ZW species (i.e. female heterogamety, in opposition to XY male heterogametic species) with a partial dosage compensation, but underlying mechanisms are still unexplored. Here, we combine transcriptomic (RNA-Seq) and epigenetic data (ChIP-Seq against H3K4me3, H3K27me3, and H4K20me1 histone marks) in free larval cercariae and intravertebrate parasitic stages. For the first time, we describe differences in dosage compensation status in ZW females, depending on the parasitic status: free cercariae display global dosage compensation, whereas intravertebrate stages show a partial dosage compensation. We also highlight regional differences of gene expression along the Z chromosome in cercariae, but not in the intravertebrate stages. Finally, we feature a consistent permissive chromatin landscape of the Z chromosome in both sexes and stages. We argue that dosage compensation in schistosomes is characterized by chromatin remodeling mechanisms in the Z-specific region. [ABSTRACT FROM AUTHOR]

In birds, the female is heterogametic (ZW) and the male homogametic (ZZ). The small W chromosome comprises only 28 protein coding genes (homologues to Z chromosome counterparts) and a number of repeat regions. Here, we report our analysis of one of these genes, BTF3 (basic transcription factor 3), which exhibits differential expression during gonadogenesis. We measured RNA levels of both Z and W homologues and BTF3 protein levels in male and female gonads during development of the chicken embryo. In addition, BTF3 RNA and protein levels were compared in female gonads (ovary) and in female gonads following treatment to induce sex reversal (testis). Combined BTF3 RNA levels were higher in female gonads than male gonads, while BTF3-Z was expressed at similar levels in males and females. Surprisingly, BTF3 protein levels were higher in male gonads than female gonads at embryonic day 6 (E6), suggesting translational rather than transcriptional regulation. BTF3 protein was expressed in both somatic and germ cells and was restricted to the medulla of the developing ovary in females and the sex cords of the developing testis in males. In addition, in gonadal sex-reversed females, RNA and protein levels of BTF3 were similar to those normally found in male gonads, suggesting that BTF3 expression correlated with the gonadal phenotype. [ABSTRACT FROM AUTHOR]

Condensins are evolutionarily conserved protein complexes that are required for chromosome segregation during cell division and genome organization during interphase. In Caenorhabditis elegans, a specialized condensin, which forms the core of the dosage compensation complex (DCC), binds to and represses X chromosome transcription. Here, we analyzed DCC localization and the effect of DCC depletion on histone modifications, transcription factor binding, and gene expression using chromatin immunoprecipitation sequencing and mRNA sequencing. Across the X, the DCC accumulates at accessible gene regulatory sites in active chromatin and not heterochromatin. The DCC is required for reducing the levels of activating histone modifications, including H3K4me3 and H3K27ac, but not repressive modification H3K9me3. In X-to-autosome fusion chromosomes, DCC spreading into the autosomal sequences locally reduces gene expression, thus establishing a direct link between DCC binding and repression. Together, our results indicate that DCC-mediated transcription repression is associated with a reduction in the activity of X chromosomal gene regulatory elements. [ABSTRACT FROM AUTHOR]

Sex determination in varanids, Gila monsters, beaded lizards, and other anguimorphan lizards is still poorly understood. Sex chromosomes were reported only in a few species based solely on cytogenetics, which precluded assessment of their homology. We uncovered Z-chromosome-specific genes in varanids from their transcriptomes. Comparison of differences in gene copy numbers between sexes across anguimorphan lizards and outgroups revealed that homologous differentiated ZZ/ZW sex chromosomes are present in Gila monsters, beaded lizards, alligator lizards, and a wide phylogenetic spectrum of varanids. However, these sex chromosomes are not homologous to those known in other amniotes. We conclude that differentiated sex chromosomes were already present in the common ancestor of Anguimorpha living in the early Cretaceous or even in the Jurassic Period, 115–180 Ma, placing anguimorphan sex chromosomes among the oldest known in vertebrates. The analysis of transcriptomes of Komodo dragon (Varanus komodoensis) showed that the expression levels of genes linked to anguimorphan sex chromosomes are not balanced between sexes. Besides expanding our knowledge on vertebrate sex chromosome evolution, our study has important practical relevance for breeding and ecological studies. We introduce the first, widely applicable technique of molecular sexing in varanids, Gila monsters, and beaded lizards, where reliable determination of sex based on external morphology is dubious even in adults. [ABSTRACT FROM AUTHOR]

Males and females of Artemia franciscana, a crustacean commonly used in the aquarium trade, are highly dimorphic. Sex is determined by a pair of ZW chromosomes, but the nature and extent of differentiation of these chromosomes is unknown. Here, we characterize the Z chromosome by detecting genomic regions that show lower genomic coverage in female than in male samples, and regions that harbor an excess of female-specific SNPs. We detect many Z-specific genes, which no longer have homologs on the W, but also Z-linked genes that appear to have diverged very recently from their existing W-linked homolog. We assess patterns of male and female expression in two tissues with extensive morphological dimorphism, gonads, and heads. In agreement with their morphology, sex-biased expression is common in both tissues. Interestingly, the Z chromosome is not enriched for sex-biased genes, and seems to in fact have a mechanism of dosage compensation that leads to equal expression in males and in females. Both of these patterns are contrary to most ZW systems studied so far, making A. franciscana an excellent model for investigating the interplay between the evolution of sexual dimorphism and dosage compensation, as well as Z chromosome evolution in general. [ABSTRACT FROM AUTHOR]

Described as a 'rare genius' by one of his close friends, Hermann Joseph Muller was one of the pioneers of modern genetics in the early decades of the twentieth century. Although his work on mutagenesis and mutations fetched him widespread recognition and glory, thanks to the Nobel Prize he received in 1946, the depth and breadth of his contributions to genetics and allied areas of biology are far more impressive. To many, he brought about a paradigm shift in the way experimental genetics was done. Even the way in which geneticists name genes — the standard genetics symbols and notation is largely as proposed by Muller. From school textbooks to advanced genetics research in genetics labs, from mutation to the mechanism of species formation, the intellectual outputs of Muller are omnipresent in the domain of genetics. His life is an inspiring story of a person waging an immense struggle against personal and political nightmares and yet, almost heroically solving some of the most challenging and fundamental problems in biology at the time. If the early work of William Bateson [1], Thomas Hunt Morgan [2] and others initiated the follow-up to the resurrection of Mendel's laws, it was largely completed by Muller. The purpose of this article is to summarize some of his key contributions to biology. [ABSTRACT FROM AUTHOR]

In eutherian mammals, dosage compensation arose to balance X-linked gene expression between sexes and relatively to autosomal gene expression in the evolution of sex chromosomes. Dosage compensation occurs in early mammalian development and comprises X chromosome upregulation and inactivation that are tightly coordinated epigenetic processes. Despite a uniform principle of dosage compensation, mechanisms of X chromosome inactivation and upregulation demonstrate a significant variability depending on sex, developmental stage, cell type, individual, and mammalian species. The review focuses on relationships between X chromosome inactivation and upregulation in mammalian early development. [ABSTRACT FROM AUTHOR]

The repeatability or predictability of evolution is a central question in evolutionary biology and most often addressed in experimental evolution studies. Here, we infer how genetically heterogeneous natural systems acquire the same molecular changes to address how genomic background affects adaptation in natural populations. In particular, we take advantage of independently formed neo-sex chromosomes in Drosophila species that have evolved dosage compensation by co-opting the dosage-compensation male-specific lethal (MSL) complex to study the mutational paths that have led to the acquisition of hundreds of novel binding sites for the MSL complex in different species. This complex recognizes a conserved 21-bp GA-rich sequence motif that is enriched on the X chromosome, and newly formed X chromosomes recruit the MSL complex by de novo acquisition of this binding motif. We identify recently formed sex chromosomes in the D. melanica and D. robusta species groups by genome sequencing and generate genomic occupancy maps of the MSL complex to infer the location of novel binding sites. We find that diverse mutational paths were utilized in each species to evolve hundreds of de novo binding motifs along the neo-X, including expansions of microsatellites and transposable element (TE) insertions. However, the propensity to utilize a particular mutational path differs between independently formed X chromosomes and appears to be contingent on genomic properties of that species, such as simple repeat or TE density. This establishes the “genomic environment” as an important determinant in predicting the outcome of evolutionary adaptations. [ABSTRACT FROM AUTHOR]

In the last decade, progress has been made in methods to identify the sex determination system in plants. This gives the opportunity to study sex chromosomes that arose independently at different phylogenetic scales, and thus allows the discovery and the understanding of early stages of sex chromosome evolution. In the genus Silene, sex chromosomes have evolved independently in at least two clades from a nondioecious ancestor, the Melandrium and Otites sections. In the latter, sex chromosomes could be younger than in the section Melandrium, based on phylogenetic studies and as no heteromorphic sex chromosomes have been detected. This section might also exhibit lability in sex determination, because male heterogamy and female heterogamy have been suggested to occur. In this study, we investigated the sex determination system of two dioecious species in the section Otite s (Silene otites and its close relative Silene pseudotites). Applying the new probabilistic method SEX-DETector on RNA-seq data from cross-controlled progenies, we inferred their most likely sex determination system and a list of putative autosomal and sex-linked contigs. We showed that the two phylogenetically close species differed in their sex determination system (XY versus ZW) with sex chromosomes that derived from two different pairs of autosomes. We built a genetic map of the sex chromosomes and showed that both pairs exhibited a large region with lack of recombination. However, the sex-limited chromosomes exhibited no strong degeneration. Finally, using the "ancestral" autosomal expression of sex-linked orthologs of nondioecious S. nutans, we found a slight signature of dosage compensation in the heterogametic females of S. otites. [ABSTRACT FROM AUTHOR]

Background: During the evolution of mammalian sex chromosomes, the degeneration of Y-linked homologs has led to a dosage imbalance between X-linked and autosomal genes. The evolutionary resolution to such dosage imbalance, as hypothesized by Susumu Ohno fifty years ago, should be doubling the expression of X-linked genes. Recent studies have nevertheless shown that the X to autosome expression ratio equals ~ 1 in haploid human parthenogenetic embryonic stem (pES) cells and ~ 0.5 in diploid pES cells, suggesting no doubled expression for X-linked genes and refuting Ohno's hypothesis. Results: Here, by reanalyzing an RNA-seq-based single-cell transcriptome dataset of human embryos, we found that from the 8-cell stage until the time-point just prior to implantation, the expression levels of X-linked genes are not two-fold upregulated in male cells and gradually decrease from two-fold in female cells. Additional analyses of gene expression noise further suggest that the dosage sensitivity of X-linked genes is weaker than that of autosomal genes in differentiated female cells, which contradicts a key assumption in Ohno's hypothesis, that most X-linked genes are dosage sensitive. Moreover, the dosage-sensitive housekeeping genes are preferentially located on autosomes, implying selection against X-linkage for dosage-sensitive genes. Conclusions: We observed dosage imbalance between X-linked and autosomal genes, as well as relatively high expression noise from X-linked genes. These results collectively suggest that X-linked genes are less dosage sensitive than autosomal genes, putting one primary assumption of Ohno's hypothesis in question. [ABSTRACT FROM AUTHOR]

Background: Hexaploid bread wheat (Triticum aestivum L) arose by two polyploidisation events from three diploid species with homoeologous genomes. Nullisomic-tetrasomic (nulli-tetra or NT) lines are aneuploid wheat plants lacking two and adding two of six homoeologous chromosomes. These plants can grow normally, but with significantly morphological variations because the adding two chromosomes or the remaining four chromosomes compensate for those absent. Despite these interesting phenomena, detailed molecular mechanisms underlying dosage deletion and compensation in these useful genetic materials have not been determined. Results: By sequencing the transcriptomes of leaves in two-week-old seedlings, we showed that the profiles of differentially expressed genes between NT stocks for homoeologous group 7 and the parent hexaploid Chinese Spring (CS) occurred throughout the whole genome with a subgenome and chromosome preference. The deletion effect of nulli-chromosomes was compensated partly by the tetra-chromosomes via the dose level of expressed genes, according to the types of homoeologous genes. The functions of differentially regulated genes primarily focused on carbon metabolic process, photosynthesis process, hormone metabolism, and responding to stimulus, and etc., which might be related to the defective phenotypes that included reductions in plant height, flag leaf length, spikelet number, and kernels per spike. Conclusions: The perturbation of the expression levels of transcriptional genes among the NT stocks for homoeologous group 7 demonstrated the gene dosage effect of the subgenome at the genome-wide level. The gene dosage deletion and compensation can be used as a model to elucidate the functions of the subgenomes in modern polyploid plants. [ABSTRACT FROM AUTHOR]

Extensive cell-to-cell variation exists even among putatively identical cells, and there is great interest in understanding how the properties of transcription relate to this heterogeneity. Differential expression from the two gene copies in diploid cells could potentially contribute, yet our ability to measure from which gene copy individual RNAs originated remains limited, particularly in the context of tissues. Here, we demonstrate quantitative, single molecule allele-specific RNA FISH adapted for use on tissue sections, allowing us to determine the chromosome of origin of individual RNA molecules in formaldehyde-fixed tissues. We used this method to visualize the allele-specific expression of Xist and multiple autosomal genes in mouse kidney. By combining these data with mathematical modeling, we evaluated models for allele-specific heterogeneity, in particular demonstrating that apparent expression from only one of the alleles in single cells can arise as a consequence of low-level mRNA abundance and transcriptional bursting. [ABSTRACT FROM AUTHOR]

Abstract The Masculinizer (Masc) gene encodes a novel lepidopteran-specific protein that controls both masculinization and dosage compensation in the silkworm Bombyx mori. The Masc protein possesses two CCCH-type zinc finger domains (ZFs), a nuclear localization signal, and an 11-amino-acid region that is highly conserved among lepidopteran insects. Using a cell-based assay system, we revealed that two cysteine residues localized in the conserved region, but not ZFs, are required for masculinization. In addition, nuclear localization of the Masc protein is not associated with masculinizing activity. Because dosage compensation is considered to occur in the nucleus, we inferred that the two ZFs play a role in the establishment of dosage compensation. To investigate this hypothesis at the organism level, we utilized the CRISPR/Cas9 system and established three B. mori strains whose Masc is partially deleted at different regions. The strain lacking the 210 C-terminal amino acids of the Masc protein showed male-specific embryonic lethality due to its low abundance and/or instability. The male embryos of this strain expressed the female-type splice variants of B. mori doublesex and did not express the male-type mRNA of B. mori IGF-II mRNA-binding protein. Furthermore, mRNA levels of Z-linked genes were abnormally enhanced only in male embryos. In contrast, the strain lacking both ZFs grew normally and did not show any defective phenotypes including sexual differentiation and the expression of Z-linked genes, demonstrating that the two CCCH-type ZFs, which are conserved in lepidopteran Masc homologs, are dispensable for masculinization and dosage compensation. Graphical abstract Image 1 Highlights • Three Bombyx mori strains with mutations in the Masc gene were established by CRISPR/Cas9-mediated mutagenesis. • Deletion of C-terminal 210 amino acids of the Masc protein causes defects in masculinization and dosage compensation. • Two CCCH-type zinc finger domains of the Masc protein are dispensable for masculinization and dosage compensation. [ABSTRACT FROM AUTHOR]

In Drosophila melanogaster, the male-specific lethal (MSL) complex plays a key role in dosage compensation by stimulating expression of male X-chromosome genes. It consists of MSL proteins and two long noncoding RNAs, roX1 and roX2, that are required for spreading of the complex on the chromosome and are redundant in the sense that loss of either does not affect male viability. However, despite rapid evolution, both roX species are present in diverse Drosophilidae species, raising doubts about their full functional redundancy. Thus, we have investigated consequences of deleting roX1 and/or roX2 to probe their specific roles and redundancies in D. melanogaster. We have created a new mutant allele of roX2 and show that roX1 and roX2 have partly separable functions in dosage compensation. In larvae, roX1 is the most abundant variant and the only variant present in the MSL complex when the complex is transmitted (physically associated with the X-chromosome) in mitosis. Loss of roX1 results in reduced expression of the genes on the X-chromosome, while loss of roX2 leads to MSL-independent upregulation of genes with male-biased testis-specific transcription. In roX1 roX2 mutant, gene expression is strongly reduced in a manner that is not related to proximity to high-affinity sites. Our results suggest that high tolerance of mis-expression of the X-chromosome has evolved. We propose that this may be a common property of sex-chromosomes, that dosage compensation is a stochastic process and its precision for each individual gene is regulated by the density of high-affinity sites in the locus. [ABSTRACT FROM AUTHOR]

In each somatic cell of a female mammal one X chromosome is transcriptionally silenced via X-chromosome inactivation (XCI), initiating early in development. Although XCI events are conserved in mouse and human postimplantation development, regulation of X-chromosome dosage in preimplantation development occurs differently. In preimplantation development, mouse embryos undergo imprinted form of XCI, yet humans lack imprinted XCI and instead regulate gene expression of both X chromosomes by dampening transcription. The long non-coding RNA Xist/XIST is expressed in mouse and human preimplantation and postimplantation development to orchestrate XCI, but its role in dampening is unclear. In this review, we discuss recent advances in our understanding of the role of Xist in X chromosome dosage compensation in mouse and human. Highlights X-inactive specific transcript (Xist) is a long non-coding RNA that remains associated with the X chromosome from which it is expressed. Xist is unequivocally required for the imprinted form of X-chromosome inactivation (XCI) in mice in vivo , but demonstration of its indisputable requirement for random XCI in vivo is yet to be shown. Loss of Xist expression in mice in vivo and in conventional human pluripotent stem cells correlates with partial reactivation of genes residing on the inactive X chromosome, suggesting an important role of Xist in maintenance of the silent state of genes on the inactive X chromosome. Human preimplantation embryos have a unique X-chromosome dosage compensation state called X-chromosome dampening (XCD), where transcriptional output is tuned down from both X chromosomes. Correlative observations suggest that XCD might be mediated by XIST. [ABSTRACT FROM AUTHOR]

Reciprocal cross effects with respect to larval salivary secretory protein levels were studied in the interspecific fertile reciprocal hybrids by crossing Drosophila nasuta nasuta, and Drosophila nasuta albomicans. These proteins are produced copiously during the third larval instar stage and are believed to play a role in the attachment of pupa to the substratum prior to pupariation as well as in insect immunity. Quantitative variations were encountered among the reciprocal hybrids. Significant heterosis was observed between D. n. nasuta and the F1 hybrid female of a cross between D. n. albomicans female and D. n. nasuta male (21.39%) while the F1 hybrids of a cross between D. n. nasuta female and D. n. albomicans male showed a marginal increase (4.24%) from the mid parent level. The glue secretions were correlated to total cell number but independent of gland size. SDS PAGE revealed a considerable heterosis with respect to X-linked protein fractions. Here we report sex specific biochemical heterosis. However the X-linked fractions undergo dosage compensation in both parents and hybrids indicating strict regulatory control.

Chromosome-specific regulatory mechanisms provide a model to understand the coordinated regulation of genes on entire chromosomes or on larger genomic regions. In fruit flies, two chromosome-wide systems have been characterized: The male-specific lethal (MSL) complex, which mediates dosage compensation and primarily acts on the male X-chromosome, and Painting of fourth (POF), which governs chromosome-specific regulation of genes located on the 4th chromosome. How targeting of one specific chromosome evolves is still not understood; but repeated sequences, in forms of satellites and transposable elements, are thought to facilitate the evolution of chromosome-specific targeting. The highly repetitive 1.688 satellite has been functionally connected to both these systems. Considering the rapid evolution and the necessarily constant adaptation of regulatory mechanisms, such as dosage compensation, we hypothesised that POF and/or 1.688 may still show traces of dosage-compensation functions. Here, we test this hypothesis by transcriptome analysis. We show that loss of Pof decreases not only chromosome 4 expression but also reduces the X-chromosome expression in males. The 1.688 repeat deletion, Zhr1 (Zygotic hybrid rescue), does not affect male dosage compensation detectably; however, Zhr1 in females causes a stimulatory effect on X-linked genes with a strong binding affinity to the MSL complex (genes close to high-affinity sites). Lack of pericentromeric 1.688 also affected 1.688 expression in trans and was linked to the differential expression of genes involved in eggshell formation. We discuss our results with reference to the connections between POF, the 1.688 satellite and dosage compensation, and the role of the 1.688 satellite in hybrid lethality.

XY systems usually show chromosome-wide compensation of X-linked genes, while in many ZW systems, compensation is restricted to a minority of dosage-sensitive genes. Why such differences arose is still unclear. Here, we combine comparative genomics, transcriptomics and proteomics to obtain a complete overview of the evolution of gene dosage on the Z-chromosome of Schistosoma parasites. We compare the Z-chromosome gene content of African (Schistosoma mansoni and S. haematobium) and Asian (S. japonicum) schistosomes and describe lineage-specific evolutionary strata. We use these to assess gene expression evolution following sex-linkage. The resulting patterns suggest a reduction in expression of Z-linked genes in females, combined with upregulation of the Z in both sexes, in line with the first step of Ohno's classic model of dosage compensation evolution. Quantitative proteomics suggest that post-transcriptional mechanisms do not play a major role in balancing the expression of Z-linked genes.

RNA is a critical component of chromatin in eukaryotes, both as a product of transcription, and as an essential constituent of ribonucleoprotein complexes that regulate both local and global chromatin states. Here, we present a proximity ligation and sequencing method called Chromatin-Associated RNA sequencing (ChAR-seq) that maps all RNA-to-DNA contacts across the genome. Using Drosophila cells, we show that ChAR-seq provides unbiased, de novo identification of targets of chromatin-bound RNAs including nascent transcripts, chromosome-specific dosage compensation ncRNAs, and genome-wide trans-associated RNAs involved in co-transcriptional RNA processing.