A top-down analysis of Xa- and Xi-territories reveals differences of higher order structure at ≥ 20 Mb genomic length scales

• Published in: Nucleus (Austin, Tex.) Vol. 2; no. 5; pp. 465 - 477
• Main Authors: Teller, Kathrin, Illner, Doris, Thamm, Stefanie, Casas-Delucchi, Corella S., Versteeg, Rogier, Indemans, Mireille, Cremer, Thomas, Cremer, Marion
• Format: Journal Article
• Language: English
• Published: United States Taylor & Francis 01.09.2011
• Subjects: Binding; Biology; Bioscience; Calcium; Cancer; Cell; Cell Nucleus - ultrastructure; Cells, Cultured; chromatin; Chromosome Mapping; Chromosomes, Human, X - ultrastructure; Cycle; diploidy; DNA Probes - chemistry; Female; females; fibroblasts; fish; genes; genomics; histones; Humans; image analysis; Image Processing, Computer-Assisted; In Situ Hybridization, Fluorescence; Landes; Organogenesis; Proteins; RNA, Long Noncoding; RNA, Untranslated - analysis; RNA, Untranslated - ultrastructure; Sex Chromatin - ultrastructure; transcription (genetics)
• ISSN: 1949-1034; 1949-1042; 1949-1042
• DOI: 10.4161/nucl.2.5.17862

The active and inactive X (Xa;Xi) territory with its seemingly highly compacted Barr body in nuclei of female mammalian cells provide a key example for studies of structure/function relationships in homologous chromosomes with different functional properties. Here we used about 300 human X-specific large insert clones to generate probe sets, which target physically or functionally defined sub-chromosomal segments. We combined 3D multicolor FISH with quantitative 3D image analysis in order to compare the higher order organization in Xi-and Xa-territories in human diploid fibroblasts (HDFs) at various length scales ranging from about 50 Mb down to 1 Mb. Xi-territories were characterized by a rounder shape as compared to the flatter and more extended shape of Xa-territories. The overall compaction of the entire Xi-territory, including the Barr body, was only 1.2-fold higher than the Xa-territory. Significant differences, however, were noted between distinct subchromosomal segments: At 20 Mb length scales higher compaction in Xi-territories was restricted to specific segments, but higher compaction in these segments was not correlated  with gene density, transcriptional activity, LINE content or histone markers locally enriched in Xi-territories. Notably, higher compaction in Xi-territories observed for 20 Mb segments was not reflected accordingly by inclosed segments of 1-4 Mb. We conclude that compaction differences result mainly from a regrouping of ~1 Mb chromatin domains rather than from an increased condensation of individual domains. In contrast to a previous report, genes subject to inactivation as well as escaping from inactivation were not excluded from the interior of the Barr body.