Decision letter: The composition and organization of Drosophila heterochromatin are heterogeneous and dynamic

Abstract Heterochromatin is a ubiquitous yet poorly understood component of multicellular eukaryotic genomes. Major gaps exist in our knowledge of the nature and overall organization of DNA sequences present in heterochromatin. We have investigated the molecular structure of the 1 Mb of centric heterochromatin in the Drosophila minichromosome Dp1187. A genetic screen of irradiated minichromosomes yielded rearranged derivatives of Dp1187 whose structures were determined by pulsed-field Southern analysis and PCR. Three Dp1187 deletion derivatives and an inversion had one breakpoint in the euchromatin and one in the heterochromatin, providing direct molecular access to previously inaccessible parts of the heterochromatin. End-probed pulsed-field restriction mapping revealed the presence of at least three "islands" of complex DNA, Tahiti, Moorea, and Bora Bora, constituting approximately one half of the Dp1187 heterochromatin. Pulsed-field Southern analysis demonstrated that Drosophila heterochromatin in general is composed of alternating blocks of complex DNA and simple satellite DNA. Cloning and sequencing of a small part of one island, Tahiti, demonstrated the presence of a retroposon. The implications of these findings to heterochromatin structure and function are discussed.

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Chromatin organization in the homogeneously staining regions of a methotrexate-resistant mouse cell line: interspersion of inactive and active chromatin domains distinguished by acetylation of histone H4

Journal of Cell Science ◽

10.1242/jcs.109.9.2221 ◽

1996 ◽

Vol 109 (9) ◽

pp. 2221-2228 ◽

Cited By ~ 2

Author(s):

L. Nicol ◽

P. Jeppesen

Keyword(s):

Cell Line ◽

Dna Sequences ◽

Satellite Dna ◽

Mouse Cell ◽

Histone H4 ◽

Active Chromatin ◽

Chromatin Domains ◽

Resistant Mouse ◽

Major Satellite ◽

Drosophila Heterochromatin

We have analyzed the organization of the homogeneously staining regions (HSRs) in chromosomes from a methotrexate-resistant mouse melanoma cell line. Fluorescence in situ hybridization techniques were used to localize satellite DNA sequences and the amplified copies of the dihydrofolate reductase (DHFR) gene that confer drug-resistance, in combination with immunofluorescence using antibody probes to differentiate chromatin structure. We show that the major DNA species contained in the HSRs is mouse major satellite, confirming previous reports, and that this is interspersed with DHFR DNA in an alternating tandem array that can be resolved at the cytological level. Mouse minor satellite DNA, which is normally located at centromeres, is also distributed along the HSRs, but does not appear to interfere with centromere function. The blocks of major satellite DNA are coincident with chromatin domains that are labelled by an autoantibody that recognizes a mammalian homologue of Drosophila heterochromatin-associated protein 1, shown previously to be confined to centric heterochromatin in mouse. An antiserum that specifically recognizes acetylated histone H4, a marker for active chromatin, fails to bind to the satellite DNA domains, but labels the intervening segments containing DHFR DNA. We can find no evidence for the spreading of the inactive chromatin domains into adjacent active chromatin, even after extended passaging of cells in the absence of methotrexate selection.

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The composition and organization of Drosophila heterochromatin are heterogeneous and dynamic

eLife ◽

10.7554/elife.16096 ◽

2016 ◽

Vol 5 ◽

Cited By ~ 29

Author(s):

Joel M Swenson ◽

Serafin U Colmenares ◽

Amy R Strom ◽

Sylvain V Costes ◽

Gary H Karpen

Keyword(s):

Cell Cycle ◽

Image Analysis ◽

Drosophila Melanogaster ◽

Gene Silencing ◽

Epigenetic Factors ◽

Heterochromatin Protein ◽

Genome Wide ◽

A Genome ◽

Drosophila Heterochromatin ◽

Heterochromatin Domain

Heterochromatin is enriched for specific epigenetic factors including Heterochromatin Protein 1a (HP1a), and is essential for many organismal functions. To elucidate heterochromatin organization and regulation, we purified Drosophila melanogaster HP1a interactors, and performed a genome-wide RNAi screen to identify genes that impact HP1a levels or localization. The majority of the over four hundred putative HP1a interactors and regulators identified were previously unknown. We found that 13 of 16 tested candidates (83%) are required for gene silencing, providing a substantial increase in the number of identified components that impact heterochromatin properties. Surprisingly, image analysis revealed that although some HP1a interactors and regulators are broadly distributed within the heterochromatin domain, most localize to discrete subdomains that display dynamic localization patterns during the cell cycle. We conclude that heterochromatin composition and architecture is more spatially complex and dynamic than previously suggested, and propose that a network of subdomains regulates diverse heterochromatin functions.

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