Protein composition of interband regions in polytene and cell line chromosomes of Drosophila melanogaster

<p>Abstract</p> <p>Background</p> <p>Despite many efforts, little is known about distribution and interactions of chromatin proteins which contribute to the specificity of chromomeric organization of interphase chromosomes. To address this issue, we used publicly available datasets from several recent Drosophila genome-wide mapping and annotation projects, in particular, those from modENCODE project, and compared molecular organization of 13 interband regions which were accurately mapped previously.</p> <p>Results</p> <p>Here we demonstrate that in interphase chromosomes of <it>Drosophila </it>cell lines, the interband regions are enriched for a specific set of proteins generally characteristic of the "open" chromatin (RNA polymerase II, CHRIZ (CHRO), BEAF-32, BRE1, dMI-2, GAF, NURF301, WDS and TRX). These regions also display reduced nucleosome density, histone H1 depletion and pronounced enrichment for ORC2, a pre-replication complex component. Within the 13 interband regions analyzed, most were around 3-4 kb long, particularly those where many of said protein features were present. We estimate there are about 3500 regions with similar properties in chromosomes of <it>D. melanogaster </it>cell lines, which fits quite well the number of cytologically observed interbands in salivary gland polytene chromosomes.</p> <p>Conclusions</p> <p>Our observations suggest strikingly similar organization of interband chromatin in polytene chromosomes and in chromosomes from cell lines thereby reflecting the existence of a universal principle of interphase chromosome organization.</p

Identical Functional Organization of Nonpolytene and Polytene Chromosomes in Drosophila melanogaster

Salivary gland polytene chromosomes demonstrate banding pattern, genetic meaning of which is an enigma for decades. Till now it is not known how to mark the band/interband borders on physical map of DNA and structures of polytene chromosomes are not characterized in molecular and genetic terms. It is not known either similar banding pattern exists in chromosomes of regular diploid mitotically dividing nonpolytene cells. Using the newly developed approach permitting to identify the interband material and localization data of interband-specific proteins from modENCODE and other genome-wide projects, we identify physical limits of bands and interbands in small cytological region 9F13-10B3 of the X chromosome in D. melanogaster, as well as characterize their general molecular features. Our results suggests that the polytene and interphase cell line chromosomes have practically the same patterns of bands and interbands reflecting, probably, the basic principle of interphase chromosome organization. Two types of bands have been described in chromosomes, early and late-replicating, which differ in many aspects of their protein and genetic content. As appeared, origin recognition complexes are located almost totally in the interbands of chromosomes

Accession numbers of chromosome proteins.

<p>*modENCODE data used (<a href="http://www.modencode.org/Genomes.shtml" target="_blank">http://www.modencode.org/Genomes.shtml</a>).</p><p>**GEO data used (<a href="http://www.ncbi.nlm.nih.gov/gds/" target="_blank">http://www.ncbi.nlm.nih.gov/gds/</a>).</p

Localization of proteins and DNA elements in 9F13 – 10B3 region of nonpolytene chromosomes (according to data of modENCODE).

<p>The positions of proteins were located as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#s4" target="_blank">Material and Methods</a>. <b>A</b> - physical map of DNA; positions of <i>v</i> and <i>sev</i> genes are taken from FlyBase, arrows 1–4 indicate position of probes for FISH on physical map. <b>B</b> - <i>P</i>-elements density in the region calculated as number of insertions per 1 kb in 10 kb interval (data on insertions are taken from FlyBase) <b>C</b> - interband specific and active chromatin specific proteins in S2 cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Kharchenko1" target="_blank">[48]</a>, <b>D</b> - DNase I hypersensitivity sites (DHS) in S2, BG3 and Kc cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Kharchenko1" target="_blank">[48] </a><b>E</b> - ORC2-binding sites in S2, BG3 and Kc cultural and salivary gland cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Eaton1" target="_blank">[59] </a><b>F</b> - histone H1 dips localization in Kc cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Braunschweig1" target="_blank">[45] </a><b>G</b> - histone H3.3 localization in S2 cells (modENCODE, Henikoff group) <b>H</b> - 30 chromatin states in BG3 and S2 cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Kharchenko1" target="_blank">[48]</a>, and 5 chromatin types in Kc cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Filion1" target="_blank">[47] </a><b>I</b> - nucleosome turnover dynamics in S2 cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Deal1" target="_blank">[60]</a>, <b>J</b> - D1 localization in Kc cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Filion1" target="_blank">[47]</a>, <b>K</b> - SUUR localization in Kc cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Filion1" target="_blank">[47]</a>, <b>L</b> - Lamin localization in Kc cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Filion1" target="_blank">[47] </a><b>M</b> - early (up) and late (down) replication in S2, Kc and BG3 cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Schwaiger1" target="_blank">[61] </a><b>N</b> - gene density (number of genes per 10 kb of DNA) <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Belyakin2" target="_blank">[54]</a>.</p

Comparison of extents of band/interbands in polytene chromosomes according to Flybase r5.25 (A, B) and to the data of this study (C).

<p>Physical DNA map is situated between 10792800 and 11220400 positions of the map of Flybase.</p

DNA compaction ratio of the bands in the 9F13 -10B3 region.

a<p>– estimated on electron microscope sections of 50 polytene chromosomes.</p><p>Notes: IB-interband; B-band.</p><p>Data of release FB2011_03 were used.</p

Relation of genetic map, and band/interband pattern in the region 9F13 – 10B3.

<p><b>A</b> - predicted bands <b>B</b> - FlyBase genes <b>C</b> – 30 chromatin states in BG3 and S2 cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Kharchenko1" target="_blank">[48] </a><b>D</b> - DNase I hypersensitivity sites (DHS) in S2, BG3 and Kc cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Kharchenko1" target="_blank">[48] </a><b>E</b> - ORC-binding sites in S2, BG3 and Kc cells <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Eaton1" target="_blank">[59] </a><b>F</b> – Nucleosome Density (modENCODE, Henikoff group) <b>G</b> - active chromatin specific <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Kharchenko1" target="_blank">[48]</a> and - interbands specific proteins. Predicted interbands (dotted vertical lines are according to peaks in distribution of corresponding elements, solid lines reflect the edges of distributions of different characteristics).</p

Comparison of several Electron Microscope sections of the region 9F11-12 – 10B (A, B) with revised Bridges map

<p><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0025960#pone.0025960-Bridges1" target="_blank">[<b>56</b>] </a><b>(C).</b> Vertical lines connect homologous bands. Scale represents 1 µm.</p

Coordinates and sizes of bands and interbands on physical map of the 9F13 – 10B3 region.

<p>Notes: IB-interband; B-band.</p><p>Data of release FB2011_03 were used.</p