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Imprinting of the Y Chromosome Influences Dosage Compensation in Rox1 Rox2 Drosophila Melanogaster

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Imprinting of the Y Chromosome Influences Dosage Compensation in Rox1 Rox2 Drosophila Melanogaster Copyright Ó 2009 by the Genetics Society of America DOI: 10.1534/genetics.109.107219 Imprinting of the Y Chromosome Influences Dosage Compensation in roX1 roX2 Drosophila melanogaster Debashish U. Menon and Victoria H. Meller1 Department of Biological Sciences, Wayne State University, Detroit, Michigan 48202 Manuscript received July 09, 2009 Accepted for publication August 14, 2009 ABSTRACT Drosophila melanogaster males have a well-characterized regulatory system that increases X-linked gene expression. This essential process restores the balance between X-linked and autosomal gene products in males. A complex composed of the male-specific lethal (MSL) proteins and RNA is recruited to the body of transcribed X-linked genes where it modifies chromatin to increase expression. The RNA components of this complex, roX1 and roX2 (RNA on the X1, RNA on the X2), are functionally redundant. Males mutated for both roX genes have dramatically reduced survival. We show that reversal of sex chromosome inheritance suppresses lethality in roX1 roX2 males. Genetic tests indicate that the effect on male survival depends upon the presence and source of the Y chromosome, revealing a germ line imprint that influences dosage compensation. Conventional paternal transmission of the Y chromosome enhances roX1 roX2 lethality, while maternal transmission of the Y chromosome suppresses lethality. roX1 roX2 males with both maternal and paternal Y chromosomes have very low survival, indicating dominance of the paternal imprint. In an otherwise wild-type male, the Y chromosome does not appreciably affect dosage compensation. The influence of the Y chromosome, clearly apparent in roX1 roX2 mutants, thus requires a sensitized genetic background. We believe that the Y chromosome is likely to act through modulation of a process that is de- fective in roX1 roX2 mutants: X chromosome recognition or chromatin modification by the MSL complex. ODIFICATION of genetic material in the parental Golic 2002). Rearrangements of the X chromosome M germ line can affect the structure, segregation, or that move euchromatic genes into the vicinity of an expression of chromosomes in the zygote (reviewed by imprinting center in proximal heterochromatin also Lloyd 2000; de la Casa-Esperon and Sapienza 2003). display imprinted PEV (Anaka et al. 2009). A common Parent-of-origin effects mediated by epigenetic marks theme in Drosophila imprinting is the central role of on chromosomes are called germ line imprints. The heterochromatin. Imprinted marks reside in hetero- importance of imprints for mammalian embryonic de- chromatin, and formation of heterochromatin in the velopment is illustrated by the early lethality of uniparental zygote is required for the maintenance of imprints diploids (Surani et al. 1986). Unlike mammals, Drosoph- (Lloyd et al. 1999). Because PEV requires rearranged ila uniparental diploids are viable and without apparent chromosomes or insertion of a transgene into hetero- defect, suggesting that the role of imprinting in flies is chromatin, another recurring motif is that the affected minor (Lindsley and Zimm 1992). In spite of this, chromosome is structurally abnormal, or a reporter has imprinting does occur in Drosophila and is detected been moved into an abnormal chromatin environment. through its effect on gene expression. Euchromatic Sex chromosomes are frequent targets of germ line genes that are moved to heterochromatic environments imprints, perhaps because their fate is unusually pre- allrath by inversion or transposition are silenced (W dictable. Fathers always donate a Y to their sons (YP) and Elgin 1995). Silencing, detected by variegated and an X (XP) to their daughters. Maternally derived expression, is termed position effect variegation (PEV). X chromosomes (XM) are hemizygous when passed to a With few exceptions, imprinting in Drosophila is son. Parents may thus anticipate the genetic and detected through modulation of PEV. Although most developmental environment that these chromosomes variegating insertions are not affected by transmission, a will encounter in the zygote. This is exploited in few are influenced by their parent of origin (Golic et al. regulation of several processes. Germ line imprinting 1998; Haller and Woodruff 2000; Maggert and of mammalian X-linked genes has been implicated in neural development and determination of sex-specific Supporting information is available online at http://www.genetics.org/ behaviors (reviewed in Wilkinson et al. 2007). Germ cgi/content/full/genetics.109.107219/DC1. line imprinting directs inactivation of the paternal X 1Corresponding author: Department of Biological Sciences, Wayne State University, 5047 Gullen Mall, Detroit, MI 48202. chromosome in female marsupials, and in the extra E-mail: [email protected] embryonic tissues of female rodents (Migeon 1998). Genetics 183: 811–820 (November 2009) 812 D. U. Menon and V. H. Meller Inactivation of a single X chromosome equalizes ex- stable association of the MSL complex with the X pression between females that carry two copies of the chromosome. Furthermore, integration of roX into the X chromosome and males that carry single X and Y MSL complex is required for normal chromatin mod- chromosomes, a process termed dosage compensation. ification by the complex (Park et al. 2008). In spite of Drosophila also compensate for unequal X chromo- the importance of the roX genes in dosage compensa- some dosage in males and females, but accomplish this tion, how roX RNA regulates changes in the localization by increasing transcription from genes on the single and activity of the MSL complex is poorly understood. male X chromosome (Lucchesi et al. 2005). Male flies We observed that reversal of sex chromosome in- dosage compensate normally regardless of the origin of heritance is a potent suppressor of roX1 roX2 male their X chromosome. Imprinting therefore does not lethality. Males carrying a paternal roX1 roX2 chromo- identify the X chromosome in male Drosophila. Both some and a maternal Y chromosome have dramatically sex determination and dosage compensation in flies is higher survival than males that inherit identical sex determined by the number of X chromosomes present chromosomes conventionally. Surprisingly, this effect (Baker and Belote 1983). While the Y chromosome car- can be attributed solely to the presence, and parent of ries genes necessary for male fertility, it is not believed to origin, of the Y chromosome. We find that a maternally play a regulatory role in either sex determination or transmitted Y chromosome suppresses roX1 roX2 lethal- dosage compensation. In flies, dosage compensation is ity, a paternally transmitted Y chromosome enhances accomplished by the male-specific lethal (MSL) com- roX1 roX2 lethality, and absence of the Y chromosome plex, composed of proteins and RNA. The MSL complex produces an intermediate level of male survival. Males binds within the body of X-linked genes and alters with both maternal and paternal Y chromosomes have chromatin to enhance transcription (Alekseyenko et al. very low survival, suggesting that the effect of the 2006; Gilfillan et al. 2006; Legube et al. 2006). The paternal Y chromosome is dominant. In spite of the protein-coding components of the MSL complex were widely held view that the Y chromosome has little genetic identified by the male-specific lethality of mutations in information or importance, Y chromosomes from dif- these genes. mle (maleless), msl1, -2, and -3 (male specific ferent Drosophila strains have unexpectedly large lethals 1, -2, and -3), and mof (males absent on first) effects on expression throughout the genome, particu- together define a set of genes essential for compensation larly the expression of male-biased genes (Lemos et al. (Mendjan and Akhtar 2007). Mutation of any one of 2008). However, the Y chromosome is not necessary for these genes causes male lethality as third instar larvae or dosage compensation and is not believed to influence pupae, but none is essential in females. Elimination of this process in otherwise normal males (reviewed in an individual protein not only blocks transcriptional Lucchesi 1973). The effect we observe thus requires a upregulation in males, but also lowers the levels of the roX1 roX2 mutant background. A dose-sensitive X-linked remaining MSL proteins and disrupts their association reporter and quantitative reverse transcription–PCR with the X. (qRT–PCR) of X-linked genes reveals higher expression The large, noncoding roX1 and roX2 RNAs (RNA on in roX1 roX2 males with a maternal Y chromosome than the X) are essential but redundant components of the with a paternal Y chromosome. We conclude that a MSL complex. Both roX RNAs are highly male prefer- maternally imprinted Y chromosome suppresses roX1 ential in expression. Both genes are X linked, and their roX2 lethality through a process that culminates in transcripts assemble with the MSL proteins and localize increased expression of X-linked genes. along the male X chromosome (Meller et al. 1997; Akhtar et al. 2000; Smith et al. 2000). Simultaneous mutation of roX1 and roX2 causes male-specific lethality, MATERIALS AND METHODS although males have normal survival with a single intact roX gene (Meller and Rattner 2002). Localization of Fly culture and genetics: Flies were maintained at 25° on standard cornmeal–agar fly food in a humidified incubator. the MSL complex to the X chromosome is disrupted in Unless otherwise noted, all mutations are described in Lindsley roX1 roX2 males. In polytene preparations
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