Mutations in CG8878, a Novel Putative Protein Kinase, Enhance P Element Dependent Silencing (PDS) and Position Effect Variegation (PEV) in Drosophila melanogaster

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.

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

Molecular and Cellular Biology ◽

10.1128/mcb.22.4.1218-1232.2002 ◽

2002 ◽

Vol 22 (4) ◽

pp. 1218-1232 ◽

Cited By ~ 43

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.

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

Genetics ◽

10.1093/genetics/163.3.931 ◽

2003 ◽

Vol 163 (3) ◽

pp. 931-937 ◽

Cited By ~ 3

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.

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

Genetics ◽

10.1093/genetics/148.1.211 ◽

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.

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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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Mutation in P0, a Dual Function Ribosomal Protein/Apurinic/Apyrimidinic Endonuclease, Modifies Gene Expression and Position Effect Variegation in Drosophila

Genetics ◽

10.1093/genetics/150.4.1487 ◽

1998 ◽

Vol 150 (4) ◽

pp. 1487-1495

Author(s):

Maxim V Frolov ◽

James A Birchler

Keyword(s):

Gene Expression ◽

Ribosomal Protein ◽

Position Effect ◽

P Element ◽

Dual Function ◽

Insertion Mutant ◽

Position Effect Variegation ◽

Element Insertion ◽

Effect Variegation ◽

Ribosomal Protein P0

Abstract In a search for modifiers of gene expression with the white eye color gene as a target, a third chromosomal P-element insertion mutant l(3)01544 has been identified that exhibits a strong pigment increase in a white-apricot background. Molecular analysis shows that the P-element insertion is found in the first intron of the gene surrounding the insertion site. Sequencing both the cDNA and genomic fragments revealed that the identified gene is identical to one encoding ribosomal protein P0/apurinic/apyrimidinic endonuclease. The P-element-induced mutation, l(3)01544, affects the steady-state level of white transcripts and transcripts of some other genes. In addition, l(3)01544 suppresses the variegated phenotypes of In(1)wm4h and In(1)y3P, suggesting a potential involvement of the P0 protein in modifying position effect variegation. The revertant generated by the precise excision of the P element has lost all mutant phenotypes. Recent work revealed that Drosophila ribosomal protein P0 contains an apurinic/apyrimidinic endonuclease activity. Our results suggest that this multifunctional protein is also involved in regulation of gene expression in Drosophila.

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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 heterochromatin-associated protein HP-1 is an essential protein in Drosophila with dosage-dependent effects on position-effect variegation.

Genetics ◽

10.1093/genetics/131.2.345 ◽

1992 ◽

Vol 131 (2) ◽

pp. 345-352 ◽

Cited By ~ 17

Author(s):

J C Eissenberg ◽

G D Morris ◽

G Reuter ◽

T Hartnett

Keyword(s):

Germ Line ◽

Position Effect ◽

Inducible Promoter ◽

Specific Protein ◽

P Element ◽

Position Effect Variegation ◽

Chromosomal Protein ◽

Gene Encoding ◽

Position Effects ◽

Effect Variegation

Abstract Chromosome rearrangements which place euchromatic genes adjacent to a heterochromatic breakpoint frequently result in gene repression (position-effect variegation). This repression is thought to reflect the spreading of a heterochromatic structure into neighboring euchromatin. Two allelic dominant suppressors of position-effect variegation were found to contain mutations within the gene encoding the heterochromatin-specific chromosomal protein HP-1. The site of mutation for each allele is given: one converts Lys169 into a nonsense (ochre) codon, while the other is a frameshift after Ser10. In flies heterozygous for one of the mutant alleles (Su(var)2-504), a truncated HP-1 protein was detectable by Western blot analysis. An HP-1 minigene, consisting of HP-1 cDNA under the control of an Hsp70 heat-inducible promoter, was transduced into flies by P element-mediated germ line transformation. Heat-shock driven expression of this minigene results in elevated HP-1 protein level and enhancement of position-effect variegation. Levels of variegating gene expression thus appear to depend upon the level of expression of a heterochromatin-specific protein. The implications of these observations for mechanism of heterochromatic position effects and heterochromatin function are discussed.

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