Unknown,Transcriptomics,Genomics,Proteomics

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Global gene expression divergence among mutation-accumulation Y-chromosome substitution lines of Drosophila melanogaster

ABSTRACT: Gene expression profiling revealed that hundreds of genes were differentially expressed among these Y introgression lines, and that global gene expression pattern diverged between the low-PEV and high-PEV (defined in the accompanying publication) flies. The results further expand our understanding of the role of the Y chromosome in modulating global gene expression, and suggest a link with modifications of chromatin state. Keywords: position-effect variegation (PEV) The Y chromosomes of H15 and H23 have lower level of suppression on the expression of white gene, therefore show lower level of eye color variegation. In contrast, the Y chromosomes of H5 and H7 have higher level of suppression on the expression of white gene resulting higher level of eye variegation. A loop design was used with all the Y introgression lines. In addition to biological replicates, a Cy5-Cy3 dye swap was performed.

ORGANISM(S): Drosophila melanogaster

SUBMITTER: Jun Zhou

PROVIDER: E-GEOD-37068 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:H3K9 methylation (H3K9me) is a conserved marker of heterochromatin, a transcriptionally silent chromatin structure. Knowledge of the mechanisms for regulating heterochromatin distribution is limited. The fission yeast JmjC domain-containing protein Epe1 localizes to heterochromatin mainly through its interaction with Swi6, a homologue of heterochromatin protein 1 (HP1), and directs JmjC-mediated H3K9me demethylation in vivo. Here, we found that loss of epe1 (epe1∆) induced a red-white variegated phenotype in a red-pigment accumulation background that generated uniform red colonies. Analysis of isolated red and white colonies revealed that silencing of genes involved in pigment accumulation by stochastic ectopic heterochromatin formation led to white colony formation. In addition, genome-wide analysis of red- and white-isolated clones revealed that epe1∆ resulted in a heterogeneous heterochromatin distribution among clones. We found that Epe1 had an N-terminal domain distinct from its JmjC domain, which activated transcription in both fission and budding yeasts. The N-terminal transcriptional activation (NTA) domain was involved in suppression of ectopic heterochromatin-mediated red-white variegation. We introduced a single copy of Epe1 into epe1∆ clones harboring ectopic heterochromatin, and found that Epe1 could reduce H3K9me from ectopic heterochromatin but some of the heterochromatin persisted. This persistence was due to a latent H3K9me source embedded in ectopic heterochromatin. Epe1H297A, a canonical JmjC mutant, suppressed red-white variegation, but entirely failed to remove already-established ectopic heterochromatin, suggesting that Epe1 prevented stochastic de novo deposition of ectopic H3K9me in an NTA-dependent but JmjC-independent manner, while its JmjC domain mediated removal of H3K9me from established ectopic heterochromatin. Our results suggest that Epe1 not only limits the distribution of heterochromatin but also controls the balance between suppression and retention of heterochromatin-mediated epigenetic diversification.

Project description:Fly strains were obtained from the Bloomington Stock Center. The chromosomes containing the gene deletion mod(mdg4)L3101 {y[1] w[1118]; P{w[+mC]=lacW}mod(mdg4)[L3101]/TM3, Ser[1]}, HP1 {In(1)w[m4h]; Su(var)205[5]/In(2L)Cy,In(2R)Cy, Cy[1]}, mod(mdg4)G16853 {w[1118]; P{w[+mC]=EP}mod(mdg4)[G16853]/TM6C, Sb[1]}, and Jil-1Scim {y[1]; P{y[+mDint2] w[BR.E.BR]=SUPor-P}JIL-1[Scim] ry[506]} were introgressed into the genetic background y[1]; bw[1]; e[4]; ci[1] ey[R] to generate the strains Mod(mdg4)/+, Su(var)205/+ and JIL-1/+. All females used in the introgression were collected within 7 hours after eclosion. For gene expression analyses, flies were grown in incubators at 25°C, 65% of relative humidity, and constant light. Newly emerged male adults flies harboring the mutations and the Y chromosomes Yohio and Ycongo(Mod(mdg4)/+;Yohio and Mod(mdg4)/+;Ycongo, Su(var)205/+;Yohio and Su(var)205/+;Ycongo, and JIL-1/+;Yohio and JIL-1/+;Ycongo) were collected and aged for 2 days at the same rearing condition before they were flash-frozen in liquid nitrogen. Four replicas of each sample were collected and stored at -80°C. Total RNA was extracted from whole flies using TRIzol (Life Technologies). The synthesis of cDNA and its labeling with fluorescent dyes (Cy3 and Cy5) as well as hybridization reactions were carried out using 3DNA protocols and reagents (Genisphere). The genetic interaction of the Y chromosome and the mutation was studied by the following contrasts: 1) male flies Mod(mdg4)/+;Yohio versus male flies Mod(mdg4)/+;Ycongo; 2) male flies Su(var)205/+;Yohio versus male flies Su(var)205/+;Ycongo; 3) male flies JIL-1/+;Yohio versus male flies JIL-1/+;Ycongo). As reference we used the same genetic background without the mutations [+/+;Ycongo (background 2) and +/+;Yohio (background 2)]. Slides were scanned using Axon 400B scanner (Axon Instruments) and GenePix Pro 6.0 software. Foreground fluorescence of dye intensities was normalized by the Loess method in Bioconductor / Limma. Dye "swaps," loop design.

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Global gene expression divergence among mutation-accumulation Y-chromosome substitution lines of Drosophila melanogaster

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