Abstract
Giant lampbrush chromosomes (LBCs) typical for growing oocytes of various animal species are characterized by a specific chromomere-loop appearance and massive transcription. Chromomeres represent universal units of chromatin packaging at LBC stage. While quite good progress has been made in investigation of LBCs structure and function, chromomere organization still remains poorly understood. To extend our knowledge on chromomere organization, we applied microdissection to chicken LBCs. In particular, 31 and 5 individual chromomeres were dissected one by one along the macrochromosome 4 and one microchromosome, respectively. The data on genomic context of individual chromomeres was obtained by high-throughput sequencing of the corresponding chromomere DNA. Alignment of adjacent chromomeres to chicken genome assembly provided information on chromomeres size and genomic boarders, indicating that prominent marker chromomeres are about 4–5 Mb in size, while common chromomeres of 1.5–3.5 Mb. Analysis of genomic features showed that the majority of chromomere-loop complexes combine gene-dense and gene-poor regions, while massive loopless DAPI-positive chromomeres lack genes and are remarkably enriched with different repetitive elements. Finally, dissected LBC chromomeres were compared with chromatin domains (topologically associated domains [TADs] and A/B-compartments), earlier identified by Hi-C technique in interphase nucleus of chicken embryonic fibroblasts. Generally, the results obtained suggest that chromomeres of LBCs do not correspond unambiguously to any type of well-established spatial domains of interphase nucleus in chicken somatic cells.
Highlights
In highly extended chromosomes, such as polytene chromosomes, lampbrush chromosomes (LBCs), and pachytene chromosomes, a chromomere is defined as a universal unit of chromatin packaging (Vlad and Macgregor, 1975)
To analyze the genomic organization of LBC chromomeres, we applied mechanical microdissection to chicken lampbrush chromosomes followed by preparation of DNA-libraries of isolated chromomeres
All prominent chromomeres were dissected one by one along the chicken lampbrush macrochromosome 4 starting from the q-ter chromosome region: chromomeres ##1–31 (Supplementary Figure S1)
Summary
In highly extended chromosomes, such as polytene chromosomes, lampbrush chromosomes (LBCs), and pachytene chromosomes, a chromomere is defined as a universal unit of chromatin packaging (Vlad and Macgregor, 1975). An array of chromomeres constitutes an axis of each LBC, with neighboring chromomeres being connected by thin decondensed chromatin threads (interchromomeric fibers). Since chromomeres constantly appear in the same positions one can develop cytological maps reflecting the number, size, and general pattern of distribution of chromomeres along LBC's axes (Galkina et al, 2005; Galkina et al, 2006; Daks et al, 2010; Zlotina et al, 2012). LBC axes lack any linker histones H1 (Hock et al, 1993)
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