scholarly journals Genetics of P-Element Transposition Into Drosophila melanogaster Centric Heterochromatin

Genetics ◽  
2003 ◽  
Vol 165 (4) ◽  
pp. 2039-2053 ◽  
Author(s):  
Alexander Y Konev ◽  
Christopher M Yan ◽  
David Acevedo ◽  
Cameron Kennedy ◽  
Elaina Ward ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Position Effect ◽  
Molecular Genetic ◽  
Phenotypic Selection ◽  
Single Copy ◽  
P Element ◽  
Position Effect Variegation ◽  
Entry Points ◽  
Drosophila Heterochromatin ◽  
Genetic Entry

Abstract Heterochromatin is a major component of higher eukaryotic genomes, but progress in understanding the molecular structure and composition of heterochromatin has lagged behind the production of relatively complete euchromatic genome sequences. The introduction of single-copy molecular-genetic entry points can greatly facilitate structure and sequence analysis of heterochromatic regions that are rich in repeated DNA. In this study, we report the isolation of 502 new P-element insertions into Drosophila melanogaster centric heterochromatin, generated in nine different genetic screens that relied on mosaic silencing (position-effect variegation, or PEV) of the yellow gene present in the transposon. The highest frequencies of recovery of variegating insertions were observed when centric insertions were used as the source for mobilization. We propose that the increased recovery of variegating insertions from heterochromatic starting sites may result from the physical proximity of different heterochromatic regions in germline nuclei or from the association of mobilizing elements with heterochromatin proteins. High frequencies of variegating insertions were also recovered when a potent suppressor of PEV (an extra Y chromosome) was present in both the mobilization and selection generations, presumably due to the effects of chromatin structure on P-element mobilization, insertion, and phenotypic selection. Finally, fewer variegating insertions were recovered after mobilization in females, in comparison to males, which may reflect differences in heterochromatin structure in the female and male germlines. FISH localization of a subset of the insertions confirmed that 98% of the variegating lines contain heterochromatic insertions and that these schemes produce a broader distribution of insertion sites. The results of these schemes have identified the most efficient methods for generating centric heterochromatin P insertions. In addition, the large collection of insertions produced by these screens provides molecular-genetic entry points for mapping, sequencing, and functional analysis of Drosophila heterochromatin.

Mutations in ash1 and trx enhance P-element-dependent silencing in Drosophila melanogaster

Genome ◽  
2016 ◽  
Vol 59 (8) ◽  
pp. 527-540
Author(s):  
Allen McCracken ◽  
John Locke
Keyword(s):  
Drosophila Melanogaster ◽  
Spontaneous Mutation ◽  
Position Effect ◽  
P Element ◽  
Position Effect Variegation ◽  
Genetic Modifiers ◽  
Gypsy Element ◽  
P Elements ◽  
Opposite Action ◽  
Dose Dependent

In Drosophila melanogaster, the mini-w+ transgene in Pci is normally expressed throughout the adult eye; however, when other P or KP elements are present, a variegated-eye phenotype results, indicating random w+ silencing during development called P-element-dependent silencing (PDS). Mutant Su(var)205 and Su(var)3-7 alleles act as haplo-suppressors/triplo-enhancers of this variegated phenotype, indicating that these heterochromatic modifiers act dose dependently in PDS. Previously, we recovered a spontaneous mutation of P{lacW}ciDplac called P{lacW}ciDplacE1 (E1) that variegated in the absence of P elements, presumably due to the insertion of an adjacent gypsy element. From a screen for genetic modifiers of E1 variegation, we describe here the isolation of five mutations in ash1 and three in trx that enhance the E1 variegated phenotype in a dose-dependent and cumulative manner. These mutant alleles enhance PDS at E1, and in E1/P{lacW}ciDplac, but suppress position effect variegation (PEV) at In(1)wm4. This opposite action is consistent with a model where ASH1 and TRX mark transcriptionally active chromatin domains. If ASH1 or TRX function is lost or reduced, heterochromatin can spread into these domains creating a sink that diverts heterochromatic proteins from other variegating locations, which then may express a suppressed phenotype.

scholarly journals The dose of a putative ubiquitin-specific protease affects position-effect variegation in Drosophila melanogaster.

1996 ◽  
Vol 16 (10) ◽  
pp. 5717-5725 ◽  
Author(s):  
S Henchoz ◽  
F De Rubertis ◽  
D Pauli ◽  
P Spierer
Keyword(s):  
Drosophila Melanogaster ◽  
Position Effect ◽  
Transcript Level ◽  
Open Reading Frame ◽  
P Element ◽  
Position Effect Variegation ◽  
Reading Frame ◽  
Effect Variegation ◽  
Specific Protease ◽  
Ubiquitin Specific Protease

A dominant insertional P-element mutation enhances position-effect variegation in Drosophila melanogaster. The mutation is homozygous, viable, and fertile and maps at 64E on the third chromosome. The corresponding gene was cloned by transposon tagging. Insertion of the transposon upstream of the open reading frame correlates with a strong reduction of transcript level. A transgene was constructed with the cDNA and found to have the effect opposite from that of the mutation, namely, to suppress variegation. Sequencing of the cDNA reveals a large open reading frame encoding a putative ubiquitin-specific protease (Ubp). Ubiquitin marks various proteins, frequently for proteasome-dependent degradation. Ubps can cleave the ubiquitin part from these proteins. We discuss the link established here between a deubiquitinating enzyme and epigenetic silencing processes.

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

Genome ◽  
2001 ◽  
Vol 44 (4) ◽  
pp. 698-707
Author(s):  
Patrick Morcillo ◽  
Ross J MacIntyre
Keyword(s):  
Drosophila Melanogaster ◽  
Heat Shock ◽  
Fusion Gene ◽  
Position Effect ◽  
P Element ◽  
Lacz Fusion ◽  
Position Effect Variegation ◽  
Salivary Gland Cells ◽  
P Element Transformation

A hsp70–lacZ fusion gene introduced into Drosophila melanogaster at the euchromatic 31B region by P-element 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.Key words: Drosophila, euchromatic position effect, heat shock construct.

scholarly journals The AT-Hook Protein D1 Is Essential for Drosophila melanogaster Development and Is Implicated in Position-Effect Variegation

2002 ◽  
Vol 22 (4) ◽  
pp. 1218-1232 ◽  
Author(s):  
Nathalie Aulner ◽  
Caroline Monod ◽  
Guillaume Mandicourt ◽  
Denis Jullien ◽  
Olivier Cuvier ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Position Effect ◽  
D1 Protein ◽  
P Element ◽  
Centromeric Heterochromatin ◽  
Position Effect Variegation ◽  
Chromosomal Protein ◽  
Mrna Levels ◽  
Satellite Repeats ◽  
Effect Variegation

ABSTRACT We have analyzed the expression pattern of the D1 gene and the localization of its product, the AT hook-bearing nonhistone chromosomal protein D1, during Drosophila melanogaster development. D1 mRNAs and protein are maternally contributed, and the protein localizes to discrete foci on the chromosomes of early embryos. These foci correspond to 1.672- and 1.688-g/cm3 AT-rich satellite repeats found in the centromeric heterochromatin of the X and Y chromosomes and on chromosomes 3 and 4. D1 mRNA levels subsequently decrease throughout later development, followed by the accumulation of the D1 protein in adult gonads, where two distributions of D1 can be correlated to different states of gene activity. We show that the EP473 mutation, a P-element insertion upstream of D1 coding sequences, affects the expression of the D1 gene and results in an embryonic homozygous lethal phenotype correlated with the depletion of D1 protein during embryogenesis. Remarkably, decreased levels of D1 mRNA and protein in heterozygous flies lead to the suppression of position-effect variegation (PEV) of the white gene in the white-mottled (wm4h ) X-chromosome inversion. Our results identify D1 as a DNA-binding protein of known sequence specificity implicated in PEV. D1 is the primary factor that binds the centromeric 1.688-g/cm3 satellite repeats which are likely involved in white-mottled variegation. We propose that the AT-hook D1 protein nucleates heterochromatin assembly by recruiting specialized transcriptional repressors and/or proteins involved in chromosome condensation.

The Drosophila melanogaster sir2+ Gene Is Nonessential and Has Only Minor Effects on Position-Effect Variegation

Genetics ◽  
2003 ◽  
Vol 163 (3) ◽  
pp. 931-937 ◽  
Author(s):  
Stefan U Åström ◽  
Thomas W Cline ◽  
Jasper Rine
Keyword(s):  
Drosophila Melanogaster ◽  
Life Span ◽  
Transcriptional Start Site ◽  
Position Effect ◽  
Developmental Rate ◽  
P Element ◽  
Position Effect Variegation ◽  
Lethal Mutations ◽  
Effect Variegation ◽  
Dependent Protein

Abstract Five Drosophila melanogaster genes belong to the highly conserved sir2 family, which encodes NAD+-dependent protein deacetylases. Of these five, dsir2+ (CG5216) is most similar to the Saccharomyces cerevisiae SIR2 gene, which has profound effects on chromatin structure and life span. Four independent Drosophila strains were found with P-element insertions near the dsir2 transcriptional start site as well as extraneous linked recessive lethal mutations. Imprecise excision of one of these P elements (PlacW 07223) from a chromosome freed of extraneous lethal mutations produced dsir217, a null intragenic deletion allele that generates no DSIR2 protein. Contrary to expectations from the report by Rosenberg and Parkhurst on their P-mobilization allele dSir2ex10, homozygosity for dsir217 had no apparent deleterious effects on viability, developmental rate, or sex ratio, and it fully complemented sir2ex10. Moreover, through a genetic test, we ruled out the reported effect of dSir2ex10 on Sex-lethal expression. We did observe a modest, strictly recessive suppression of whitem4 position-effect variegation and a shortening of life span in dsir2 homozygous mutants, suggesting that dsir2 has some functions in common with yeast SIR2.

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

Genome ◽  
2000 ◽  
Vol 43 (2) ◽  
pp. 285-292 ◽  
Author(s):  
Bethany S Haller ◽  
R C Woodruff
Keyword(s):  
Drosophila Melanogaster ◽  
Y Chromosome ◽  
Genetic Background ◽  
Position Effect ◽  
P Element ◽  
Parental Origin ◽  
Position Effect Variegation ◽  
Reporter System ◽  
Heterochromatin Protein 1 ◽  
Parental Female

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. Key words: Drosophila melanogaster, heterochromatin, HP1, imprinting, P-element, Y chromosome.

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

Genetics ◽  
2000 ◽  
Vol 154 (2) ◽  
pp. 657-668 ◽  
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.

Enhancer of Polycomb Is a Suppressor of Position-Effect Variegation in Drosophila melanogaster

Genetics ◽  
1998 ◽  
Vol 148 (1) ◽  
pp. 211-220
Author(s):  
Donald A R Sinclair ◽  
Nigel J Clegg ◽  
Jennifer Antonchuk ◽  
Thomas A Milne ◽  
Kryn Stankunas ◽  
...  
Keyword(s):  
Sequence Similarity ◽  
Position Effect ◽  
Polycomb Group ◽  
P Element ◽  
Homeotic Gene ◽  
Position Effect Variegation ◽  
Negative Regulators ◽  
Recombination Mapping ◽  
Effect Variegation ◽  
Homeotic Gene Expression

Abstract Polycomb group (PcG) genes of Drosophila are negative regulators of homeotic gene expression required for maintenance of determination. Sequence similarity between Polycomb and Su(var)205 led to the suggestion that PcG genes and modifiers of position-effect variegation (PEV) might function analogously in the establishment of chromatin structure. If PcG proteins participate directly in the same process that leads to PEV, PcG mutations should suppress PEV. We show that mutations in E(Pc), an unusual member of the PcG, suppress PEV of four variegating rearrangements: In(l)wm4, BSV, T(2;3)SbV, and In(2R)bwVDe2. Using reversion of a P element insertion, deficiency mapping, and recombination mapping as criteria, homeotic effects and suppression of PEV associated with E(Pc) co-map. Asx is an enhancer of PEV, whereas nine other PcG loci do not affect PEV. These results support the conclusion that there are fewer similarities between PcG genes and modifiers of PEV than previously supposed. However, E(Pc) appears to be an important link between the two groups. We discuss why Asx might act as an enhancer of PEV.

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

Genetics ◽  
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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