A Role for siRNA in X-Chromosome Dosage Compensation in <i>Drosophila melanogaster</i>

Menon, Debashish U.; Meller, Victoria H.

doi:10.1534/genetics.112.140236

Cited by 40 publications

(58 citation statements)

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“…Additionally, the constant level of expression of testis-specific paralogs under DCV infection (Figure 2) suggests that they have not evolved an inducible response to viral infection, either restricted to the testis or in other tissues. In contrast, one paralog in each species has retained the ubiquitous expression pattern seen in D. melanogaster (D. subobscura Ago2a, D. obscura Ago2a, and D. pseudoobscura Ago2c; Figure 3), suggesting that these paralogs have retained roles in antiviral defense (Li et al 2002;van Rij et al 2006), dosage compensation (Menon and Meller 2012), and/or somatic TE suppression Czech et al 2008).…”

Section: Discussionmentioning

confidence: 99%

“…This function imposes strong selective constraint on Ago1, resulting in slow evolution and very few adaptive substitutions (Obbard et al 2006(Obbard et al , 2009aKolaczkowski et al 2011). Finally, Ago2 binds siRNAs from viruses (viRNAs) and TEs (endo-siRNAs), and functions in gene regulation (Wen et al 2015), dosage compensation (Menon and Meller 2012), and the ubiquitous suppression of viruses (Li et al 2002;van Rij et al 2006) and TEs (Chung et al 2008;Czech et al 2008). Ago2 also evolves under strong positive selection, with frequent selective sweeps (Obbard et al 2006(Obbard et al , 2009a(Obbard et al ,b, 2011Kolaczkowski et al 2011), possibly driven by an arms race with virus-encoded suppressors of RNAi (VSRs) (Obbard et al 2006;Marques and Carthew 2007;van Mierlo et al 2014).…”

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confidence: 99%

“…Given the roles of D. melanogaster Ago2 in antiviral defense (Li et al 2002;van Rij et al 2006), TE suppression Czech et al 2008), dosage compensation (Menon and Meller 2012), and gene regulation (Wen et al 2015), we hypothesized that these D. pseudoobscura Ago2 paralogs may have diverged in function. To elucidate the evolution and function of Ago2 paralogs in D. pseudoobscura and its relatives, we identified and dated Ago2 duplication events across available Drosophila genomes and transcriptomes, tested for divergence in expression patterns between the Ago2 paralogs in D. subobscura, D. obscura, and D. pseudoobscura, and quantified the evolutionary rate and positive selection acting on each of these paralogs.…”

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Repeated duplication of Argonaute2 is associated with strong selection and testis specialization inDrosophila

Lewis

Webster

Salmela

et al. 2016

Preprint

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Argonaute2 (Ago2) is a rapidly evolving nuclease in the Drosophila melanogaster RNA interference (RNAi) pathway that targets viruses and transposable elements in somatic tissues. Here we reconstruct the history of Ago2 duplications across the D. obscura group and use patterns of gene expression to infer new functional specialization. We show that some duplications are old, shared by the entire species group, and that losses may be common, including previously undetected losses in the lineage leading to D. pseudoobscura. We find that while the original (syntenic) gene copy has generally retained the ancestral ubiquitous expression pattern, most of the novel Ago2 paralogs have independently specialized to testis-specific expression. Using population genetic analyses, we show that most testis-specific paralogs have significantly lower genetic diversity than the genome-wide average. This suggests recent positive selection in three different species, and model-based analyses provide strong evidence of recent hard selective sweeps in or near four of the six D. pseudoobscura Ago2 paralogs. We speculate that the repeated evolution of testis specificity in obscura group Ago2 genes, combined with their dynamic turnover and strong signatures of adaptive evolution, may be associated with highly derived roles in the suppression of transposable elements or meiotic drive. Our study highlights the lability of RNAi pathways, even within well-studied groups such as Drosophila, and suggests that strong selection may act quickly after duplication in RNAi pathways, potentially giving rise to new and unknown RNAi functions in nonmodel species. KEYWORDS Argonaute; RNAi; Drosophila; duplication; testis A RGONAUTE genes are found in almost all eukaryotes, where they play a key role in antiviral immune defense, gene regulation, and genome stability. They perform this diverse range of functions through their role in RNA interference (RNAi) mechanisms, an ancient system of nucleic acid manipulation in which small RNA (sRNA) molecules guide Argonaute proteins to nucleic acid targets through base complementarity (reviewed in Meister 2013). Gene duplication has occurred throughout the evolution of the Argonaute gene family, with ancient duplication events characteristic of some lineages-such as three duplications early in plant evolution (Singh et al. 2015) and multiple expansions and losses throughout the evolution of nematodes (reviewed in Buck and Blaxter 2013) and the Diptera . After duplication, Argonautes have often undergone functional divergence, involving changes in expression patterns and altered sRNA binding partners (Lu et al. 2011;Leebonoi et al. 2015;Miesen et al. 2015). Duplication early in eukaryotic evolution produced two distinct Argonaute subfamilies, Ago and Piwi, which have since been retained in the vast majority of Metazoa (Cerutti and Casas-Mollano 2006). Members of the Ago subfamily are expressed in both somatic and germline tissue, and variously bind sRNAs derived from host transcripts (miRNAs, endo-siRNAs) ...

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Section: Discussionmentioning

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Repeated duplication of Argonaute2 is associated with strong selection and testis specialization inDrosophila

Lewis

Webster

Salmela

et al. 2016

Preprint

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“…The siRNA pathway contributes to X chromosome recognition during dosage compensation (9). This suggests that siRNAproducing sequences on the X chromosome might participate in the identification of X chromatin.…”

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siRNAs from an X-linked satellite repeat promote X-chromosome recognition in Drosophila melanogaster

Menon

Coarfa

Xiao

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

109

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Highly differentiated sex chromosomes create a lethal imbalance in gene expression in one sex. To accommodate hemizygosity of the X chromosome in male fruit flies, expression of X-linked genes increases twofold. This is achieved by the male-specific lethal (MSL) complex, which modifies chromatin to increase expression. Mutations that disrupt the X localization of this complex decrease the expression of X-linked genes and reduce male survival. The mechanism that restricts the MSL complex to X chromatin is not understood. We recently reported that the siRNA pathway contributes to localization of the MSL complex, raising questions about the source of the siRNAs involved. The X-linked 1.688 g/cm 3 satellite related repeats (1.688 X repeats) are restricted to the X chromosome and produce small RNA, making them an attractive candidate. We tested RNA from these repeats for a role in dosage compensation and found that ectopic expression of singlestranded RNAs from 1.688 X repeats enhanced the male lethality of mutants with defective X recognition. In contrast, expression of double-stranded hairpin RNA from a 1.688 X repeat generated abundant siRNA and dramatically increased male survival. Consistent with improved survival, X localization of the MSL complex was largely restored in these males. The striking distribution of 1.688 X repeats, which are nearly exclusive to the X chromosome, suggests that these are cis-acting elements contributing to identification of X chromatin.ales and females of many species have an unequal number of X chromosomes, producing a potentially fatal imbalance in X-linked gene expression (1). The process by which balance is restored is called dosage compensation. In the male fruit fly, Drosophila melanogaster, the male-specific lethal (MSL) complex modifies the chromatin of X-linked genes to increase expression by twofold, equalizing expression between XX females and XY males (2). The long noncoding roX RNAs assemble with the MSL proteins to form the intact MSL complex. roX RNA is required for exclusive X-chromosome binding of the complex and for increased expression of X-linked genes (3, 4).How the MSL complex selectively recognizes X chromatin is not fully understood, but an elegant model for X recognition proposes that the complex is first recruited to chromatin entry sites (CESs), then spreads into nearby active genes through a cotranscriptional mechanism (5). The CESs, defined by elevated affinity for the MSL proteins, are limited to the X chromosome (6). A 21-bp motif, termed the MSL recognition element (MRE), is enriched within the CES and binds CLAMP, a protein essential for MSL recruitment (7, 8); however, MREs are only modestly enriched on the X chromosome, and CLAMP binds autosomal MREs without recruiting the MSL complex. These observations indicate that additional factors must contribute to X identification.The siRNA pathway contributes to X chromosome recognition during dosage compensation (9). This suggests that siRNAproducing sequences on the X chromosome might participate in the ...

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“…These scaffolds are often comprised of repetitive sequences and represent in large part, heterochromatic regions (Smith et al 2007). siRNAs have been implicated in Xchromosome dosage compensation in D. melanogaster where knockout of the siRNA specific Ago2 gene affects male survival (Menon and Meller 2012). In contrast to mammals where dosage compensation occurs through the inactivation of one copy of the X chromosome in females, dosage compensation in D. melanogaster occurs through the enhancement of the transcription of genes located on the single male X chromosome (Gilfillan et al 2004, Conrad andAkhtar 2012).…”

Section: High-uniformity Index-possessing Non-mirna Clusters In D Mementioning

confidence: 99%

RNA interference in early branching metazoans

Calcino¹

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The RNA interference (RNAi) repertoire of metazoans is principally composed of three independent but related systems -endogenous small interfering RNAs (endosiRNAs), micro RNAs (miRNAs) and Piwi-interacting RNAs (piRNAs). Although the endosiRNA pathway has been demonstrated in all five eukaryotic supergroups, the miRNA and piRNA pathways of metazoans likely evolved subsequent to metazoan divergence from other eukaryotes. These innovations evolved in a 100-200 million year period between the emergence of animal multicellularity and the divergence of the sponges. It is not yet known if the organisation and function of these systems, as characterised in bilaterian model species (vertebrates, Drosophila and C. elegans), is shared between early branching phyla (Porifera, Ctenophora, Cnidaria and Placozoa) and bilaterians. Preliminary evidence is mixed with commonalities like the existence of 'ping-pong' piRNA biogenesis in sponges and cnidarians contrasted with the lack of sequence and secondary structure homology of miRNAs between sponges and bilaterians.This thesis focuses on the RNAi systems of three early branching metazoan phyla that represent critical branches in early animal evolution (ie. Porifera, Ctenophora and Cnidaria). As no such tool was available, I developed a bioinformatic pipeline to annotate the RNAi components of mapped small RNA libraries from the demosponge Amphimedon queenslandica, the ctenophore Mnemiopsis leidyi and the cnidarian Nematostella vectensis. Detailed in Chapter 2, this pipeline, named RNAiTool, clusters mapped small RNAs from high-throughput sequencing data and then annotates those clusters based on a simple set of criteria, primarily focused on the read-length of the constituent small RNAs.RNAiTool was also used in the annotation of small RNA datasets from the well-studied bilaterian Drosophila melanogaster, providing a point of comparison for the less wellunderstood non-bilaterian species. As well as interrogating individual datasets, RNAiTool can be used to compare the expression dynamics of endo-siRNA and piRNA cluster between datasets. Although it was used here to investigate the expression dynamics of endo-siRNA and piRNA cluster through development, RNAiTool can be used to assess the expression dynamics of clusters between any test and control datasets.The second major innovation presented in this thesis is the introduction of the uniformity index. This property of small RNA clusters describes the uniformity of small RNA coverage across the length of that cluster. This simple index provides a rapid method 3 for the identification of potential miRNAs and other highly expressed small RNA producing loci from large pools of small RNA clusters. This thesis takes the first steps towards an understanding of the evolution and function of RNAi systems in early branching metazoans, in particular the demosponge A.queenslandica. It is hoped that the framework for RNAi annotation developed here will prove useful for other studies of emerging model RNAi systems. 4

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A Role for siRNA in X-Chromosome Dosage Compensation in Drosophila melanogaster

Cited by 40 publications

References 47 publications

Repeated duplication of Argonaute2 is associated with strong selection and testis specialization inDrosophila

Repeated duplication of Argonaute2 is associated with strong selection and testis specialization inDrosophila

siRNAs from an X-linked satellite repeat promote X-chromosome recognition in Drosophila melanogaster

RNA interference in early branching metazoans

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A Role for siRNA in X-Chromosome Dosage Compensation in Drosophila melanogaster

Cited by 40 publications

References 47 publications

Repeated duplication of Argonaute2 is associated with strong selection and testis specialization inDrosophila

Repeated duplication of Argonaute2 is associated with strong selection and testis specialization inDrosophila

siRNAs from an X-linked satellite repeat promote X-chromosome recognition in Drosophila melanogaster

﻿RNA interference in early branching metazoans

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RNA interference in early branching metazoans