Article

Identification of Genes That Promote or Antagonize Somatic Homolog Pairing Using a High-Throughput FISH-Based Screen

Department of Genetics, Harvard Medical School, Boston, Massachusetts, United States of America.
PLoS Genetics (Impact Factor: 8.17). 05/2012; 8(5):e1002667. DOI: 10.1371/journal.pgen.1002667
Source: PubMed

ABSTRACT The pairing of homologous chromosomes is a fundamental feature of the meiotic cell. In addition, a number of species exhibit homolog pairing in nonmeiotic, somatic cells as well, with evidence for its impact on both gene regulation and double-strand break (DSB) repair. An extreme example of somatic pairing can be observed in Drosophila melanogaster, where homologous chromosomes remain aligned throughout most of development. However, our understanding of the mechanism of somatic homolog pairing remains unclear, as only a few genes have been implicated in this process. In this study, we introduce a novel high-throughput fluorescent in situ hybridization (FISH) technology that enabled us to conduct a genome-wide RNAi screen for factors involved in the robust somatic pairing observed in Drosophila. We identified both candidate "pairing promoting genes" and candidate "anti-pairing genes," providing evidence that pairing is a dynamic process that can be both enhanced and antagonized. Many of the genes found to be important for promoting pairing are highly enriched for functions associated with mitotic cell division, suggesting a genetic framework for a long-standing link between chromosome dynamics during mitosis and nuclear organization during interphase. In contrast, several of the candidate anti-pairing genes have known interphase functions associated with S-phase progression, DNA replication, and chromatin compaction, including several components of the condensin II complex. In combination with a variety of secondary assays, these results provide insights into the mechanism and dynamics of somatic pairing.

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    • "Brain cell preparations from 72 to 96-hour third-instar larvae with empty guts showed mitotic pro-metaphases, metaphases and anaphases. Somatic pairing was observed in metaphase plates as reported for other dipterans (Stevens 1908, Metz 1916, Joyce et al. 2012). We distinguished two types of chromosome numbers within the studied samples: 2n=10 and 2n=11. "
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    • "In addition to its well-known functions in chromatin condensation along the chromosome axis during nuclear division and in interphase, condensin complexes inhibit associations between homologues (reviewed by Wood et al. 2010). Drosophila condensin II antagonizes the pairing of homologues (Joyce et al. 2012) and reduces transvection by limiting interactions between homologues during interphase. Furthermore, the condensin II subunits SMC4, Cap-H2 and Cap-D3 are necessary to disassemble aligned polytene chromosomes (Hartl et al. 2008a). "
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    • "However, our observations reported here and previous studies in polyploid , diploid, and male meiotic chromosomes suggesting that Cap-H2 is required for chromosome unpairing (Hartl et al. 2008a,b) support a model in which Cap-H2 and Mrg15 antagonize transvection by inhibiting trans-interactions . That Cap-H2 provides a homolog antipairing activity has recently been shown for euchromatic and heterochromatic sequences in cultured Drosophila cells (Joyce et al. 2012; Buster et al. 2013) also suggests a more direct function in antagonizing transvection. Interestingly, the Mrg15 homolog in Caenorhabditis elegans (Mrg-1) has also been suggested to antagonize ectopic pairing in meiosis (Dombecki et al. 2011). "
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