Expression in Aneuploid Drosophila S2 Cells

Laboratory of Cellular and Developmental Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, United States of America.
PLoS Biology (Impact Factor: 9.34). 02/2010; 8(2):e1000320. DOI: 10.1371/journal.pbio.1000320
Source: PubMed


Extensive departures from balanced gene dose in aneuploids are highly deleterious. However, we know very little about the relationship between gene copy number and expression in aneuploid cells. We determined copy number and transcript abundance (expression) genome-wide in Drosophila S2 cells by DNA-Seq and RNA-Seq. We found that S2 cells are aneuploid for >43 Mb of the genome, primarily in the range of one to five copies, and show a male genotype ( approximately two X chromosomes and four sets of autosomes, or 2X;4A). Both X chromosomes and autosomes showed expression dosage compensation. X chromosome expression was elevated in a fixed-fold manner regardless of actual gene dose. In engineering terms, the system "anticipates" the perturbation caused by X dose, rather than responding to an error caused by the perturbation. This feed-forward regulation resulted in precise dosage compensation only when X dose was half of the autosome dose. Insufficient compensation occurred at lower X chromosome dose and excessive expression occurred at higher doses. RNAi knockdown of the Male Specific Lethal complex abolished feed-forward regulation. Both autosome and X chromosome genes show Male Specific Lethal-independent compensation that fits a first order dose-response curve. Our data indicate that expression dosage compensation dampens the effect of altered DNA copy number genome-wide. For the X chromosome, compensation includes fixed and dose-dependent components.

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Available from: Eric P Spana, Oct 06, 2015
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    • "One important scenario for adaptive regulation of gene expression is when there are changes or differences in gene dose (Veitia et al. 2008, 2013). Maintaining ancestral protein levels to sustain stoichiometric relationships in networks and pathways might in such cases be critical for proper function of protein–protein interactions (Zhang et al. 2010). A classical question in genetics to which this applies is the potential need for sex-linked dosage compensation upon sex chromosome evolution and the concomitant degeneration of nonrecombining genes (Ohno 1967; Disteche 2012). "
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    ABSTRACT: There is increasing evidence that dosage compensation is not a ubiquitous feature following sex chromosome evolution, especially not in organisms where females are the heterogametic sex, like in birds. Even when it occurs, compensation can be incomplete and limited to dosage-sensitive genes. However, previous work has mainly studied transcriptional regulation of sex-linked genes, which may not reflect expression at the protein level. Here we used liquid chromatography - tandem mass spectrometry (LC-MS/MS) to detect and quantify expressed levels of more than 2,400 proteins in 10 different tissues of male and female chicken embryos. For comparison, transcriptome sequencing was performed in the same individuals, five of each sex. The proteomic analysis revealed that dosage compensation was incomplete, with a mean male-to-female (M:F) expression ratio of Z-linked genes of 1.32 across tissues, similar to that at the RNA level (1.29). The mean Z chromosome-to-autosome expression ratio was close to one in males and lower than one in females, consistent with partly reduced Z chromosome expression in females. While our results exclude a general mechanism for chromosome-wide dosage compensation at translation, 30% of all proteins encoded from Z-linked genes showed a significant change in the M:F ratio compared to the corresponding ratio at the RNA level. This resulted in a pattern where some genes showed balanced expression between sexes and some close to twofold higher expression in males. This suggests that proteomic analyses will be necessary to reveal a more complete picture of gene regulation and sex chromosome evolution. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.
    Molecular Biology and Evolution 06/2015; DOI:10.1093/molbev/msv147 · 9.11 Impact Factor
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    • "In eutherian mammals, dosage compensation occurs through X-inactivation: the ncRNA XIST suppresses most expression on one X copy in females 48, 49. In contrast to mammals, Drosophila have a macromolecular complex thought to increase expression of genes on the single male X chromosome by about 2-fold, thus making male and female expression effectively the same; it is worth noting, however, that complex network dynamics and transcriptional control are also involved 50-53. Other compensation systems, such as incomplete and gene-specific inactivation, have been found in the monotreme platypus and chickens (an “atypical” XY system and ZW, respectively; 54). "
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    ABSTRACT: Frog sex chromosomes offer an ideal system for advancing our understanding of genome evolution and function because of the variety of sex determination systems in the group, the diversity of sex chromosome maturation states, the ease of experimental manipulation during early development. After briefly reviewing sex chromosome biology generally, we focus on what is known about frog sex determination, sex chromosome evolution, and recent, genomics-facilitated advances in the field. In closing we highlight gaps in our current knowledge of frog sex chromosomes, and suggest priorities for future research that can advance broad knowledge of gene dose and sex chromosome evolution.
    03/2014; 2:68-76. DOI:10.7150/jgen.8044
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    • "This was unexpected given that the ChIP-Seq data was generated in a cell model system rather than a sexually dimorphic brain. Chromosome analysis of S2 cells have shown that they are male in origin and have an X/A ration of 0.5, as is found in male Drosophila, demonstrating similar dosage compensation as is seen for X-linked genes in the male fly38 This might, in part, explain the bias towards target genes that are specifically regulated in the male brain. In any case, we demonstrate here that the in vitro S2 model system represents a powerful starting point providing ease of manipulation for investigating the activity of transcription factors such as Fru, that are involved in complex behavioural programs. "
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    ABSTRACT: The fruitless gene (fru) encodes a set of transcription factors (Fru) that display sexually dimorphic gene expression in the brain of the fruit-fly; Drosophila melanogaster. Behavioural studies have demonstrated that fru is essential for courtship behaviour in the male fly and is thought to act by directing the development of sex-specific neural circuitry that encodes this innate behavioural response. This study reports the identification of direct regulatory targets of the sexually dimorphic isoforms of the Fru protein using an in vitro model system. Genome wide binding sites were identified for each of the isoforms using Chromatin Immunoprecipitation coupled to deep sequencing (ChIP-Seq). Putative target genes were found to be involved in processes such as neurotransmission, ion-channel signalling and neuron development. All isoforms showed a significant bias towards genes located on the X-chromosome, which may reflect a specific role for Fru in regulating x-linked genes. Taken together with expression analysis carried out in Fru positive neurons specifically isolated from the male fly brain, it appears that the Fru protein acts as a transcriptional activator. Understanding the regulatory cascades induced by Fru will help to shed light on the molecular mechanisms that are important for specification of neural circuitry underlying complex behaviour.
    Scientific Reports 03/2014; 4:4412. DOI:10.1038/srep04412 · 5.58 Impact Factor
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