Genomic imprinting in Drosophila is maintained by the products of Suppressor of variegation and trithorax group,but not Polycomb group,genes

Genomic imprinting is a form of epigenetic inheritance that is characterized by differential expression of a gene depending on its parental origin. The mini-X chromosome Dp(1;f)LJ9 in Drosophila shows this type of classical imprinting; when transmitted by the maternal parent genes on this chromosome are fully expressed, but when the chromosome is transmitted by the male parent at least three genes are subject to silencing, resulting in a variegated expression pattern. Chemical and environmental modifiers of position-effect variegation have been shown to alter the somatic maintenance of the imprint. To extend these observations, several mutations in chromatin-associated proteins were examined for their effect on imprinting on the Dp(1;f)LJ9 mini-X chromosome. Effects on establishment and maintenance were independently assessed by genetically associating the mutations in chromatin modifiers with the mini-X chromosome in either the parents, where the imprint is established, or the progeny, in which the imprint must be maintained. Nine Suppressor of variegation [ Su(var)] mutations, including alleles of the Su(var)2-5 gene, which encodes the well characterized heterochromatin-associated protein HP1, abolished maintenance but not the establishment of the imprint. Mutant alleles of two genes in the trithorax group ( trx-G), brahma and trithorax, showed a maternal-effect enhancement of the paternal imprint. Surprisingly, however, with the exception of an Enhancer of Polycomb [ E(Pc)] allele, none of the Polycomb-group ( Pc-G) mutations tested affected the imprint. Thus, the maintenance of this imprint relies on the wild-type products of Su(var) and trx-G, but not Pc-G, genes. Finally, none of the mutations tested affected the maintenance of the maternal imprint or the establishment of either the maternal or paternal imprint, suggesting that the maternal and paternal imprints depend on different molecular processes and that imprint establishment and maintenance are independently regulated.     

The Effect of Modifiers of Position-Effect Variegation on the Variegation of Heterochromatic Genes of Drosophila Melanogaster

Dominant modifiers of position-effect variegation of Drosophila melanogaster were tested for their effects on the variegation of genes normally located in heterochromatin. These modifiers were previously isolated as strong suppressors of the variegation of euchromatic genes and have been postulated to encode structural components of heterochromatin or other products that influence chromosome condensation. While eight of the modifiers had weak or no detectable effects, six acted as enhancers of light (lt) variegation. The two modifiers with the strongest effects on lt were shown to also enhance the variegation of neighboring heterochromatic genes. These results suggest that the wild-type gene products of some modifiers of position-effect variegation are required for proper expression of genes normally located within or near the heterochromatin of chromosome 2. We conclude that these heterochromatic genes have fundamentally different regulatory requirements compared to those typical of euchromatic genes.     

Varied expression of a Y-linked P[w+] insert due to imprinting in Drosophila melanogaster.

During gametogenesis, a gene can become imprinted affecting its expression in progeny. We have used the expression of a Y-linked P[w+]YAL transposable DNA element as a reporter system to investigate the effect of parental origination on the expression of the w+ insert. Expression of w+ was greater in male progeny when the Y chromosome, harboring the insert, was inherited from the parental male rather than from the parental female. Imprinting was not due to a genetic background influence in the males, since the only difference among the males was the parental origin of the Y chromosome. It was also observed that the genetic background can affect imprinting, since w+ expression was also higher in males when the Y was derived from C(1)DX attached-X parental females rather than from C(1)RM attached-X parental females. Though the heterochromatic imprinting mechanism is unknown, a mutated Heterochromatin Protein 1 (HP1) gene, which is associated with suppression of position-effect variegation, increases expression of the w+ locus in the P[w+]YAL insert, indicating that HP1 may play a role in Y chromosome packaging.     

Genomic imprinting: male mice with uniparentally derived sex chromosomes

Although it has been known that there is an X-chromosome imprinting effect during early embryogenesis in female mammals, it remains unknown if parental origin of the X chromosome has an effect in males. Furthermore, it has not been possible to produce animals with normal sex chromosomes of uniparental origin to further evaluate such imprinting effects. We have devised a breeding scheme to produce male mice, designated XPYP males, in which both the X and Y chromosomes are paternally inherited. To our knowledge, these are the first mammals produced that have a normal sex chromosome constitution but with both sex chromosomes derived from one parent. Development and reproduction in these XPYP males and the sex ratio and chromosome constitution of their offspring appeared normal; thus there is no apparent effect in males of having both sex chromosomes derive from one parent or of having the X chromosome derived from an inappropriate parent. Although we have detected no X-chromosome imprinting effect in these males, evidence from other sources suggest that the X chromosome is parentally imprinted. Thus detection and definition of an imprint can depend on the assay used.     

Germ line imprinting in Drosophila: Epigenetics in search of function

Germ line imprinting produces parent-specific differences in the behavior of chromosomes or expression of genes. Epigenetic marks, placed on chromosomes in the parental germ line, govern classical imprinted effects such as chromosomal inactivation, chromosome elimination and mono-allelic expression. Germ line imprinting occurs in insects, plants and mammals. Several Drosophila systems display imprinted effects. In spite of this, many aspects of imprinting in flies, including the normal function of this process, remain mysterious. Transgenerational inheritance of epigenetic marks is a powerful force in genome regulation. Elucidation of the mechanism of imprint establishment and maintenance in a model organism, such as Drosophila, is thus of great interest. In this review we summarize the primary systems that have been used to study imprinting in flies and speculate on the origin and biological function of imprinting in Drosophila.     

Imprinting defects on human chromosome 15

The Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are two distinct neurogenetic diseases that are caused by the loss of function of imprinted genes on the proximal long arm of human chromosome 15. In a few percent of patients with PWS and AS, the disease is due to aberrant imprinting and gene silencing. In patients with PWS and an imprinting defect, the paternal chromosome carries a maternal imprint. In patients with AS and an imprinting defect, the maternal chromosome carries a paternal imprint. Imprinting defects offer a unique opportunity to identify some of the factors and mechanisms involved in imprint erasure, resetting and maintenance. In approximately 10% of cases the imprinting defects are caused by a microdeletion affecting the 5' end of the SNURF-SNRPN locus. These deletions define the 15q imprinting center (IC), which regulates imprinting in the whole domain. These findings have been confirmed and extended in knock-out and transgenic mice. In the majority of patients with an imprinting defect, the incorrect imprint has arisen without a DNA sequence change, possibly as the result of stochastic errors of the imprinting process or the effect of exogenous factors.     

Genetic and molecular characterization of a variegating hsp70-acZ fusion gene in the euchromatic 31 B region of Drosophila melanogaster.

A hsp70-lacZ fusion gene introduced into Drosophila melanogaster at the euchromatic 31B region by Pelement transformation displayed a variegated expression with respect to the lacZ fusion protein in the salivary gland cells under heat-shock conditions. The variegation is also reflected by the chromosome puffing pattern. Subsequent transposition of the 31B P element to other euchromatic positions restored wild-type activity, that is, a nonvariegated phenotype. A lower developmental temperature reduced the amount of expression under heat-shock conditions, similar to genes undergoing position-effect variegation (PEV). However, other modifiers of PEV did not affect the expression pattern of the gene. These results show a novel euchromatic tissue-specific variegation that is not associated with classical heterochromatic PEV.     

An unexpected function of the Prader-Willi syndrome imprinting center in maternal imprinting in mice

Genomic imprinting is a phenomenon that some genes are expressed differentially according to the parent of origin. Prader-Willi syndrome (PWS) and Angelman syndrome (AS) are neurobehavioral disorders caused by deficiency of imprinted gene expression from paternal and maternal chromosome 15q11-q13, respectively. Imprinted genes at the PWS/AS domain are regulated through a bipartite imprinting center, the PWS-IC and AS-IC. The PWS-IC activates paternal-specific gene expression and is responsible for the paternal imprint, whereas the AS-IC functions in the maternal imprint by allele-specific repression of the PWS-IC to prevent the paternal imprinting program. Although mouse chromosome 7C has a conserved PWS/AS imprinted domain, the mouse equivalent of the human AS-IC element has not yet been identified. Here, we suggest another dimension that the PWS-IC also functions in maternal imprinting by negatively regulating the paternally expressed imprinted genes in mice, in contrast to its known function as a positive regulator for paternal-specific gene expression. Using a mouse model carrying a 4.8-kb deletion at the PWS-IC, we demonstrated that maternal transmission of the PWS-IC deletion resulted in a maternal imprinting defect with activation of the paternally expressed imprinted genes and decreased expression of the maternally expressed imprinted gene on the maternal chromosome, accompanied by alteration of the maternal epigenotype toward a paternal state spread over the PWS/AS domain. The functional significance of this acquired paternal pattern of gene expression was demonstrated by the ability to complement PWS phenotypes by maternal inheritance of the PWS-IC deletion, which is in stark contrast to paternal inheritance of the PWS-IC deletion that resulted in the PWS phenotypes. Importantly, low levels of expression of the paternally expressed imprinted genes are sufficient to rescue postnatal lethality and growth retardation in two PWS mouse models. These findings open the opportunity for a novel approach to the treatment of PWS.     

Parental Imprinting in Drosophila

Genetic imprinting is a form of epigenetic silencing. But with a twist. The twist is that while imprinting results in the silencing of genes, chromosome regions or entire chromosome sets, this silencing occurs only after transmission of the imprinted region by one sex of parent. Thus genetic imprinting reflects intertwined levels of epigenetic and developmental modulation of gene expression. Imprinting has been well documented and studied in Drosophila, however, these studies have remained largely unknown due to nothing more significant than differences in terminology. Imprinting in Drosophilais invariably associated with heterochromatin or regions with unusual chromatin structure. The imprint appears to spread from imprinted centers that reside within heterochromatin and these are, seemingly, the only regions that are normally imprinted in Drosophila. This is significant as it implies that while imprinting occurs in Drosophila, it is generally without phenotypic consequence. Hence the evolution of imprinting, at least in Drosophila, is unlikely to be driven by the function of specific imprinted genes. Thus, the study of imprinting in Drosophilahas the potential to illuminate the mechanism and biological function of imprinting, and challenge models based solely on imprinting of mammalian genes. This revised version was published online in July 2006 with corrections to the Cover Date.     

Disruption of the bipartite imprinting center in a family with Angelman syndrome

Imprinting in 15q11-q13 is controlled by a bipartite imprinting center (IC), which maps to the SNURF-SNRPN locus. Deletions of the exon 1 region impair the establishment or maintenance of the paternal imprint and can cause Prader-Willi syndrome (PWS). Deletions of a region 35 kb upstream of exon 1 impair maternal imprinting and can cause Angelman syndrome (AS). So far, in all affected sibs with an imprinting defect, an inherited IC deletion was identified. We report on two sibs with AS who do not have an IC deletion but instead have a 1-1.5 Mb inversion separating the two IC elements. The inversion is transmitted silently through the male germline but impairs maternal imprinting after transmission through the female germline. Our findings suggest that the close proximity and/or the correct orientation of the two IC elements are/is necessary for the establishment of a maternal imprint.     

Genomic imprinting absent in Drosophila melanogaster adult females

Genomic imprinting occurs when expression of an allele differs based on the sex of the parent that transmitted the allele. In D. melanogaster, imprinting can occur, but its impact on allelic expression genome-wide is unclear. Here, we search for imprinted genes in D. melanogaster using RNA-seq to compare allele-specific expression between pools of 7- to 10-day-old adult female progeny from reciprocal crosses. We identified 119 genes with allelic expression consistent with imprinting, and these genes showed significant clustering within the genome. Surprisingly, additional analysis of several of these genes showed that either genomic heterogeneity or high levels of intrinsic noise caused imprinting-like allelic expression. Consequently, our data provide no convincing evidence of imprinting for D. melanogaster genes in their native genomic context. Elucidating sources of false-positive signals for imprinting in allele-specific RNA-seq data, as done here, is critical given the growing popularity of this method for identifying imprinted genes.     

Genomic imprinting of chromatin inDrosophila melanogaster

During gametogenesis, chromosomes may become imprinted with information which facilitates proper expression of the DNA in offspring. We have used a position effect variegation mutant as a reporter system to investigate the possibility of imprinting inDrosophila melanogaster. Genetic crosses were performed in which the variegating gene and a strong modifier of variegation were present either within the same parental genome or in opposite parental genomes in all possible combinations. Our results indicate that the presence of the variegating chromosome and a modifier chromosome in the same parental genome can alter the amount of variegation formed in progeny. The genomic imprinting we observed is not determined by the parental origin of the variegating chromosome but is instead determined by the genetic background the variegating chromosome is subjected to during gametogenesis.     

Minichromosomes inDrosophila melanogaster derived from the transposing elementTE1

A minichromosome has originated from the transposing elementTE1. This autonomously replicating chromosome contains the structural genes white and roughest, from theDrosophila X chromosome. It arose within a stock carryingTE1 at 45F on chromosome2. In addition to thew andrst genes, the minichromosome may carry section 45C–45F from chromosome2. It is inherited by 33%–47% of the offspring. By this criterion it carries a centromere, although the origin of the centromere is unknown. From this minichromosome a still smaller one has originated, probably through the loss of all material from chromosome2 together with some heterochromatin. At the same time a duplication of white and roughest could have taken place. This chromosome has a strange morphology and is more frequently lost in meiosis than the larger one, but is still transmitted to about 29%–37% of the progeny of one parent heterozygous for the minichromosome. In both cases the flies have variegated eyes, probably as a result of position-effect variegation. The variegation pattern is influenced by factors in theX chromosome. The size of the smaller minichromosome is little more than 1 Mb as determined by pulsed field gel electrophoresis.     

Euchromatic inversions causing mosaic gene expression in Drosophila hydei: A study of forked and Zebra

In D. hydei two new mutants, In(1)f³ and IN(5)Z, show obvious mosaic gene expression. Their phenotypic expression is susceptible to the breeding temperature and to the addition of a supernumerary Y chromosome to the chromosome set. In this respect the mutants resemble standard cases of position-effect variegation based on the action of heterochromatin. However, since neither centromeric nor sex chromosomal heterochromatin apparently are involved, the mutations point to a new type of variegation provoked by euchromatic sections. The mosaic patterns of these mutants, in particular those of In(1)f³, will be described.     

Reduced DNA polytenization of a minichromosome region undergoing position-effect variegation in Drosophila

Molecular analysis of a Drosophila minichromosome, Dp(1;f)1187, revealed a relationship between position-effect variegation and the copy number reductions of heterochromatic sequences that occur in polytene cells. Heterochromatin adjacent to a defined junction with euchromatin underpolytenized at least 60-fold. Lesser reductions were observed in euchromatic sequences up to 103 kb from the breakpoint. The copy number changes behaved in all respects like the expression of yellow, a gene located within the affected region. Both copy number and yellow expression displayed a cell-by-cell mosaic pattern of reduction, and adding a Y chromosome, a known suppressor of variegation, increased both substantially. We discuss the possibility that changes in replication alter copy number locally and also propose an alternative model of position-effect variegation based on the somatic elimination of heterochromatic sequences.     

Trans-effect of modifiers on position-effect variegation in a set of euchromatin-heterochromatin rearrangements in Drosophila melanogaster]

The effects of suppressors of position-effect variegation were studied in a set of euchromatin-heterochromatin rearrangements of the X chromosome accompanied by inactivation of the gene wapl. The rearrangements differed from one another in the size of the heterochromatic block adjacent to euchromatin, with the euchromatin-heterochromatin border remaining unchanged. In one rearrangement (r20), the position effect caused by a small block of adjacent heterochromatin may be determined by its interaction with the neighboring main heterochromatic region of the X chromosome. Chromosome 3 (the RT chromosome) was found to have a strong suppressing effect on all rearrangements, irrespective of the amount of heterochromatin adjacent to euchromatin. Su-var(3)9, a known suppressor of the position-effect variegation, had a considerably weaker suppressing effect. The RT chromosome had the strongest suppressing effect on the rearrangement r20.