Comparative Analysis of Position–Effect Variegation Mutations in Drosophila melanogaster Delineates the Targets of Modifiers

Abstract In Drosophila melanogaster, heterochromatin-induced silencing or position–effect variegation (PEV) of a reporter gene has provided insights into the properties of heterochromatin. Class I modifiers suppress PEV, and class II modifiers enhance PEV when the modifier gene is present in fewer than two doses. We have examined the effects of both class I and class II modifiers on four PEV mutations. These mutations include the inversions In(1)wm4 and In(2R)bwVDe2, which are classical chromosomal rearrangements that typify PEV mutations. The other mutations are a derivative of brownDominant, in which brown+ reporters are inactivated by a large block of heterochromatin, and a P[white+] transposon insertion associated with second chromosome heterochromatin. In general, we find that class I modifiers affect both classical and nonclassical PEV mutations, whereas class II modifiers affect only classical PEV mutations. We suggest that class II modifiers affect chromatin architecture in the vicinity of reporter genes, and only class I modifiers identify proteins that are potentially involved in heterochromatin formation or maintenance. In addition, our observations support a model in which there are different constraints on the process of heterochromatin-induced silencing in classical vs. nonclassical PEV mutations.

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Suppressors of position-effect variegation in Drosophila melanogaster affect expression of the heterochromatic gene light in the absence of a chromosome rearrangement

Genome ◽

10.1139/g98-041 ◽

1998 ◽

Vol 41 (4) ◽

pp. 495-503 ◽

Cited By ~ 20

Author(s):

N J Clegg ◽

B M Honda ◽

I P Whitehead ◽

T A Grigliatti ◽

B Wakimoto ◽

...

Keyword(s):

Drosophila Melanogaster ◽

Chromosomal Rearrangements ◽

Position Effect ◽

Centromeric Heterochromatin ◽

Position Effect Variegation ◽

Gene Products ◽

Affect Expression ◽

Effect Variegation ◽

Heterochromatic Genes ◽

Var Gene

Suppressors of position-effect variegation (Su(var)s) in Drosophila melanogaster are usually studied in the presence of chromosomal rearrangements, which exhibit variegated expression of euchromatic genes moved near to, or heterochromatic genes moved away from, centromeric heterochromatin. However, the effects of Su(var) mutations on heterochromatic gene expression in the absence of a variegating re-arrangement have not yet been defined. Here we present a number of results which suggest that Su(var) gene products can interact to affect the expression of the light gene in its normal heterochromatic location. We initially observed that eye pigment was reduced in several Su(var) double mutants; the phenotype resembled that of light mutations and was more severe when only one copy of the light gene was present. This reduced pigmentation could be alleviated by a duplication for the light gene or by a reduction in the amount of cellular heterochromatin. In addition, the viability of most Su(var) double mutant combinations tested was greatly reduced in a genetic background of reduced light gene dosage, when extra heterochromatin is present. We conclude that Su(var) gene products can affect expression of the heterochromatic light gene in the absence of any chromosomal rearrangements. However, it is noteworthy that mutations in any single Su(var) gene have little effect on light expression; we observe instead that different pairings of Su(var) mutations are required to show an effect on light expression. Interestingly, we have obtained evidence that at least two of the second chromosome Su(var) mutations are gain-of-function lesions, which also suggests that there may be different modes of interaction among these genes. It may therefore be possible to use this more sensitive assay of Su(var) effects on heterochromatic genes to infer functional relationships among the products of the 50 or more known Su(var) loci.Key words: heterochromatin, chromatin, gene interactions.

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The role of heterochromatin in the expression of a heterochromatic gene, the rolled locus of Drosophila melanogaster.

Genetics ◽

10.1093/genetics/134.1.277 ◽

1993 ◽

Vol 134 (1) ◽

pp. 277-292 ◽

Cited By ~ 8

Author(s):

D F Eberl ◽

B J Duyf ◽

A J Hilliker

Keyword(s):

Drosophila Melanogaster ◽

Chromosomal Rearrangements ◽

Position Effect ◽

Centromeric Heterochromatin ◽

Position Effect Variegation ◽

Position Effects ◽

Major Function ◽

Effect Variegation ◽

Heterochromatic Genes

Abstract Constitutive heterochromatic regions of chromosomes are those that remain condensed through most or all of the cell cycle. In Drosophila melanogaster, the constitutive heterochromatic regions, located around the centromere, contain a number of gene loci, but at a much lower density than euchromatin. In the autosomal heterochromatin, the gene loci appear to be unique sequence genes interspersed among blocks of highly repeated sequences. Euchromatic genes do not function well when brought into the vicinity of heterochromatin (position-effect variegation). We test the possibility that the blocks of centromeric heterochromatin provide an environment essential for heterochromatic gene function. To assay directly the functional requirement of autosomal heterochromatic genes to reside in heterochromatin, the rolled (rl) gene, which is normally located deep in chromosome 2R heterochromatin, was relocated within small blocks of heterochromatin to a variety of euchromatic positions by successive series of chromosomal rearrangements. The function of the rl gene is severely affected in rearrangements in which the rl gene is isolated in a small block of heterochromatin, and these position effects can be reverted by rearrangements which bring the rl gene closer to any large block of autosomal or X chromosome heterochromatin. There is some evidence that five other 2R heterochromatic genes are also affected among these rearrangements. These findings demonstrate that the heterochromatic genes, in contrast to euchromatic genes whose function is inhibited by relocation to heterochromatin, require proximity to heterochromatin to function properly, and they argue strongly that a major function of the highly repeated satellite DNA, which comprises most of the heterochromatin, is to provide this heterochromatic environment.

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Dosage-dependent modifiers of position effect variegation in Drosophila and a mass action model that explains their effect.

Genetics ◽

10.1093/genetics/120.1.181 ◽

1988 ◽

Vol 120 (1) ◽

pp. 181-198

Author(s):

J Locke ◽

M A Kotarski ◽

K D Tartof

Keyword(s):

Position Effect ◽

Class Ii ◽

Mass Action ◽

P Element ◽

Class I ◽

Gene Copy ◽

Position Effect Variegation ◽

Mass Action Model ◽

Effect Variegation ◽

Action Model

Abstract Twelve dominant enhancers of position effect variegation, representing four loci on the second and third chromosomes of Drosophila melanogaster, have been induced by P-element mutagenesis. Instead of simple transposon insertions, seven of these mutations are cytologically visible duplications and three are deficiencies. The duplications define two distinct regions, each coinciding with a locus that also behaves as a dominant haplo-dependent suppressor of variegation. Conversely, two of the deficiencies overlap with a region that contains a haplo-dependent enhancer of variegation while duplications of this same region act to suppress variegation. The third deficiency defines another haplo-dependent enhancer. These data indicate that loci capable of modifying variegation do so in an antipodal fashion through changes in the wild-type gene copy number and may be divided into two reciprocally acting classes. Class I modifiers enhance variegation when duplicated or suppress variegation when deficient. Class II modifiers enhance when deficient but suppress when duplicated. From our data, and those of others, we propose that in Drosophila there are about 20 to 30 dominant loci that modify variegation. Most appear to be of the class I type whereas only two class II modifiers have been identified so far. From these observations we put forth a model, based on the law of mass action, for understanding how such suppressor-enhancer loci function. We propose that each class I modifier codes for a structural protein component of heterochromatin and their effects on variegation are a consequence of their dosage dependent influence on the extent of the assembly of heterochromatin at the chromosomal site of the position effect. It is further proposed that class II modifiers may inhibit the class I products directly, bind to hypothetical termination sites that define heterochromatin boundaries or promote euchromatin formation. Consistent with our mass action model we find that combining two enhancers together produce additive and not epistatic effects. Also, since different enhancers have different relative strengths on different variegating mutants, we suggest that heterochromatic domains are constructed by a combinatorial association of proteins. The mass action model proposed here is of general significance for any assembly driven reaction and has implications for understanding a wide variety of biological phenomena.(ABSTRACT TRUNCATED AT 400 WORDS)

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CYTOGENETIC LOCALIZATION OF THE ACID PHOSPHATASE-1 GENE IN DROSOPHILA MELANOGASTER

Genetics ◽

10.1093/genetics/88.3.487 ◽

1978 ◽

Vol 88 (3) ◽

pp. 487-497

Author(s):

William J Morrison ◽

Ross J MacIntyre

Keyword(s):

Drosophila Melanogaster ◽

Acid Phosphatase ◽

Y Chromosome ◽

Chromosomal Rearrangements ◽

Position Effect ◽

Chromosome 3 ◽

Position Effect Variegation ◽

Effect Variegation

ABSTRACT A translocation in which a segment of chromosome 3 is inserted into the Y chromosome was found to contain the acid phosphatase-1 gene (Acph-1). In flies hyperploid for that gene, acid phosphatase-1 levels are proportional to the dose of the gene. The locus is placed within the salivary chromosome subdivisions 99D and 99E on the basis of its inclusion in the translocated segment and on the previous placement of the claret locus. Several chromosomal rearrangements involving heterochromatic breakpoints and euchromatic breakpoints adjacent to 99D-99E were tested for possible position-effect variegation of acid phosphatase-1. No decrease in the synthesis of the electrophoretic subunit encoded by the relocated gene was observed within any of the rearrangements.

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Mutational Analysis of a Histone Deacetylase in Drosophila melanogaster: Missense Mutations Suppress Gene Silencing Associated With Position Effect Variegation

Genetics ◽

10.1093/genetics/154.2.657 ◽

2000 ◽

Vol 154 (2) ◽

pp. 657-668 ◽

Cited By ~ 2

Author(s):

Randy Mottus ◽

Richard E Sobel ◽

Thomas A Grigliatti

Keyword(s):

Drosophila Melanogaster ◽

Histone Deacetylase ◽

Transcriptional Activity ◽

Mutational Analysis ◽

Position Effect ◽

Transcriptional Regulator ◽

Position Effect Variegation ◽

Missense Mutations ◽

Effect Variegation ◽

New Mutations

Abstract For many years it has been noted that there is a correlation between acetylation of histones and an increase in transcriptional activity. One prediction, based on this correlation, is that hypomorphic or null mutations in histone deacetylase genes should lead to increased levels of histone acetylation and result in increased levels of transcription. It was therefore surprising when it was reported, in both yeast and fruit flies, that mutations that reduced or eliminated a histone deacetylase resulted in transcriptional silencing of genes subject to telomeric and heterochromatic position effect variegation (PEV). Here we report the first mutational analysis of a histone deacetylase in a multicellular eukaryote by examining six new mutations in HDAC1 of Drosophila melanogaster. We observed a suite of phenotypes accompanying the mutations consistent with the notion that HDAC1 acts as a global transcriptional regulator. However, in contrast to recent findings, here we report that specific missense mutations in the structural gene of HDAC1 suppress the silencing of genes subject to PEV. We propose that the missense mutations reported here are acting as antimorphic mutations that “poison” the deacetylase complex and propose a model that accounts for the various phenotypes associated with lesions in the deacetylase locus.

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Competition Between Different Variegating Rearrangements for Limited Heterochromatic Factors in Drosophila melanogaster

Genetics ◽

10.1093/genetics/145.4.945 ◽

1997 ◽

Vol 145 (4) ◽

pp. 945-959

Author(s):

Vett K Lloyd ◽

Donald A Sinclair ◽

Thomas A Grigliatti

Keyword(s):

Drosophila Melanogaster ◽

Chromosome Rearrangement ◽

Position Effect ◽

Position Effect Variegation ◽

Euchromatic Region ◽

Single Chromosome ◽

Mosaic Expression ◽

Effect Variegation ◽

Protein Components

Position effect variegation (PEV) results from the juxtaposition of a euchromatic gene to heterochromatin. In its new position the gene is inactivated in some cells and not in others. This mosaic expression is consistent with variability in the spread of heterochromatin from cell to cell. As many components of heterochromatin are likely to be produced in limited amounts, the spread of heterochromatin into a normally euchromatic region should be accompanied by a concomitant loss or redistribution of the protein components from other heterochromatic regions. We have shown that this is the case by simultaneously monitoring variegation of a euchromatic and a heterochromatic gene associated with a single chromosome rearrangement. Secondly, if several heterochromatic regions of the genome share limited components of heterochromatin, then some variegating rearrangements should compete for these components. We have examined this hypothesis by testing flies with combinations of two or more different variegating rearrangements. Of the nine combinations of pairs of variegating rearrangements we studied, seven showed nonreciprocal interactions. These results imply that many components of heterochromatin are both shared and present in limited amounts and that they can transfer between chromosomal sites. Consequently, even nonvariegation portions of the genome will be disrupted by re-allocation of heterochromatic proteins associated with PEV. These results have implications for models of PEV.

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Phenotypic observations on modification of position-effect variegation in Drosophila melanogaster

Heredity ◽

10.1038/hdy.1974.98 ◽

1974 ◽

Vol 33 (3) ◽

pp. 317-326 ◽

Cited By ~ 4

Author(s):

J M Wargent ◽

I J Hartmann-Goldstein

Keyword(s):

Drosophila Melanogaster ◽

Position Effect ◽

Position Effect Variegation ◽

Effect Variegation

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The impact of genetic background and cell lineage on the level and pattern of gene expression in position effect variegation

Epigenetics & Chromatin ◽

10.1186/s13072-019-0314-5 ◽

2019 ◽

Vol 12 (1) ◽

Author(s):

Sidney H. Wang ◽

Sarah C. R. Elgin

Keyword(s):

Genetic Variants ◽

Genetic Background ◽

Position Effect ◽

Cell Lineage ◽

Inbred Lines ◽

Position Effect Variegation ◽

Fourth Chromosome ◽

Heterochromatin Formation ◽

Effect Variegation ◽

The Impact

Abstract Background Chromatin-based transcriptional silencing is often described as a stochastic process, largely because of the mosaic expression observed in position effect variegation (PEV), where a euchromatic reporter gene is silenced in some cells as a consequence of juxtaposition with heterochromatin. High levels of variation in PEV phenotypes are commonly observed in reporter stocks. To ascertain whether background mutations are the major contributors to this variation, we asked how much of the variation is determined by genetic variants segregating in the population, examining both the level and pattern of expression using the fruit fly, Drosophila melanogaster, as the model. Results Using selective breeding of a fourth chromosome PEV reporter line, 39C-12, we isolated two inbred lines exhibiting contrasting degrees of variegation (A1: low expression, D1: high expression). Within each inbred population, remarkable similarity is observed in the degree of variegation: 90% of the variation between the two inbred lines in the degree of silencing can be explained by genotype. Further analyses suggest that this result reflects the combined effect of multiple independent trans-acting loci. While the initial observations are based on a PEV phenotype scored in the fly eye (hsp70-white reporter), similar degrees of silencing were observed using a beta-gal reporter scored across the whole fly. Further, the pattern of variegation becomes almost identical within each inbred line; significant pigment enrichment in the same quadrant of the eye was found for both A1 and D1 lines despite different degrees of expression. Conclusions The results indicate that background genetic variants play the major role in determining the variable degrees of PEV commonly observed in laboratory stocks. Interestingly, not only does the degree of variegation become consistent in inbred lines, the patterns of variegation also appear similar. Combining these observations with the spreading model for local heterochromatin formation, we propose an augmented stochastic model to describe PEV in which the genetic background drives the overall level of silencing, working with the cell lineage-specific regulatory environment to determine the on/off probability at the reporter locus in each cell. This model acknowledges cell type-specific events in the context of broader genetic impacts on heterochromatin formation.

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