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.

The P-element-induced silencing effect of KP transposons is dose dependent in Drosophila melanogaster

Genome ◽  
2011 ◽  
Vol 54 (9) ◽  
pp. 752-762 ◽  
Author(s):  
Alireza Sameny ◽  
John Locke
Keyword(s):  
Drosophila Melanogaster ◽  
Critical Role ◽  
Mutagenic Effect ◽  
P Element ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
P Elements ◽  
Single Well ◽  
Dose Dependent

Transposable elements are found in the genomes of all eukaryotes and play a critical role in altering gene expression and genome organization. In Drosophila melanogaster, transposable P elements are responsible for the phenomenon of hybrid dysgenesis. KP elements, a deletion-derivative of the complete P element, can suppress this mutagenic effect. KP elements can also silence the expression of certain other P-element-mediated transgenes in a process called P-element-dependent silencing (PDS), which is thought to involve the recruitment of heterochromatin proteins. To explore the mechanism of this silencing, we have mobilized KP elements to create a series of strains that contain single, well-defined KP insertions that show PDS. To understand the quantitative role of KP elements in PDS, these single inserts were combined in a series of crosses to obtain genotypes with zero, one, or two KP elements, from which we could examine the effect of KP gene dose. The extent of PDS in these genotypes was shown to be dose dependent in a logarithmic rather than linear fashion. A logarithmic dose dependency is consistent with the KP products interacting with heterochromatic proteins in a concentration-dependent manner such that two molecules are needed to induce gene silencing.

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 Analysis of subtelomeric heterochromatin in the Drosophila minichromosome Dp1187 by single P element insertional mutagenesis.

Genetics ◽  
1992 ◽  
Vol 132 (3) ◽  
pp. 737-753 ◽  
Author(s):  
G H Karpen ◽  
A C Spradling
Keyword(s):  
Insertional Mutagenesis ◽  
Position Effect ◽  
P Element ◽  
Centromeric Heterochromatin ◽  
Small Subset ◽  
Position Effect Variegation ◽  
Subtelomeric Heterochromatin ◽  
P Elements ◽  
Salivary Gland Cells ◽  
And Function

Abstract We investigated whether single P element insertional mutagenesis could be used to analyze heterochromatin within the Drosophila minichromosome Dp1187. Forty-five insertions of the P[lacZ,rosy+] element onto Dp1187 (recovered among 7,825 transpositions) were highly clustered. None was recovered in centromeric heterochromatin, but 39 occurred about 40 kb from the distal telomere within a 4.7-kb hotspot containing tandem copies of a novel 1.8-kb repetitive DNA sequence. The DNA within and distal to this region lacked essential genes and displayed several other properties characteristic of heterochromatin. The rosy+ genes within the inserted transposons were inhibited by position-effect variegation, and the subtelomeric region was underrepresented in polytene salivary gland cells. These experiments demonstrated that P elements preferentially transpose into a small subset of heterochromatic sites, providing a versatile method for studying the structure and function of these chromosome regions. This approach revealed that a Drosophila chromosome contains a large region of subtelomeric heterochromatin with specific structural and genetic properties.

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.

Effect of the Suppressor of Underreplication (SuUR) Gene on Position-Effect Variegation Silencing in Drosophila melanogaster

Genetics ◽  
2003 ◽  
Vol 165 (3) ◽  
pp. 1209-1220
Author(s):  
E S Belyaeva ◽  
L V Boldyreva ◽  
E I Volkova ◽  
R A Nanayev ◽  
A A Alekseyenko ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Position Effect ◽  
Polytene Chromosomes ◽  
Morphological Characters ◽  
Position Effect Variegation ◽  
Dependent Manner ◽  
Dose Dependent ◽  
Suur Protein ◽  
Chromosome Regions ◽  
Suur Gene

Abstract It has been previously shown that the SuUR gene encodes a protein located in intercalary and pericentromeric heterochromatin in Drosophila melanogaster polytene chromosomes. The SuUR mutation suppresses the formation of ectopic contacts and DNA underreplication in polytene chromosomes; SuUR+ in extra doses enhances the expression of these characters. This study demonstrates that heterochromatin-dependent PEV silencing is also influenced by SuUR. The SuUR protein localizes to chromosome regions compacted as a result of PEV; the SuUR mutation suppresses DNA underreplication arising in regions of polytene chromosomes undergoing PEV. The SuUR mutation also suppresses variegation of both adult morphological characters and chromatin compaction observed in rearranged chromosomes. In contrast, SuUR+ in extra doses and its overexpression enhance variegation. Thus, SuUR affects PEV silencing in a dose-dependent manner. However, its effect is expressed weaker than that of the strong modifier Su(var)2-5.

scholarly journals Targeted transposition at the vestigial locus of Drosophila melanogaster.

Genetics ◽  
1994 ◽  
Vol 138 (4) ◽  
pp. 1127-1135
Author(s):  
T R Heslip ◽  
R B Hodgetts
Keyword(s):  
Drosophila Melanogaster ◽  
Position Effect ◽  
Regulatory Region ◽  
Current Method ◽  
P Element ◽  
Opposite Orientation ◽  
Position Effects ◽  
P Elements ◽  
In Trans ◽  
In Cis

Abstract Targeted transposition is the replacement of one P element with another. We are exploiting this unique property of P elements to study the complex regulatory domain of the Dopa decarboxylase (Ddc) gene in Drosophila melanogaster. P element constructs targeted to the same site in the genome will be subjected to the same position effect. This allows the subtle effects typical of most mutations in the Ddc regulatory region to be measured in the absence of the variable influences of position effects which are associated with the current method of germline transformation. We have investigated some of the parameters affecting targeted transposition of a Ddc transposon, P[Ddc], into a P element allele at the vestigial locus. These events were detected by an increased mutant vg phenotype. The location of the donor transposon in cis or in trans to the target had little effect on the frequency of targeting. Likewise, the mobility of different donor elements, as measured by their rate of transposition to a different chromosome, varied nearly 20-fold, while the rate of targeted transposition was very similar between them. All targeted alleles were precise replacements of the target P element by P[Ddc], but in several cases the donor was inserted in the opposite orientation. The targeted alleles could be described as the result of a replicative, conversion-like event.

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.

scholarly journals A P element containing suppressor of hairy-wing binding regions has novel properties for mutagenesis in Drosophila melanogaster.

Genetics ◽  
1995 ◽  
Vol 141 (3) ◽  
pp. 1061-1074 ◽  
Author(s):  
R R Roseman ◽  
E A Johnson ◽  
C K Rodesch ◽  
M Bjerke ◽  
R N Nagoshi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Drosophila Melanogaster ◽  
Insertional Mutagenesis ◽  
P Element ◽  
Gypsy Element ◽  
P Elements ◽  
Structural And Functional Properties ◽  
Binding Regions ◽  
Silencer Elements ◽  
P Transposon

Abstract P elements are widely used as insertional mutagens to tag genes, facilitating molecular cloning and analyses. We modified a P element so that it carried two copies of the suppressor of Hairy-wing [su(Hw)] binding regions isolated from the gypsy transposable element. This transposon was mobilized, and the genetic consequences of its insertion were analyzed. Gene expression can be altered by the su(Hw) protein as a result of blocking the interaction between enhancer/silencer elements and their promoter. These effects can occur over long distances and are general. Therefore, a composite transposon (SUPor-P for suppressor-P element) combines the mutagenic efficacy of the gypsy element with the controllable transposition of P elements. We show that, compared to standard P elements, this composite transposon causes an expanded repertoire of mutations and produces alleles that are suppressed by su(Hw) mutations. The large number of heterochromatic insertions obtained is unusual compared to other insertional mutagenesis procedures, indicating that the SUPor-P transposon may be useful for studying the structural and functional properties of heterochromatin.

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