title: Chromatin decondensation is accompanied by a transient increase in transcriptional output.
abstract: BACKGROUND
  The levels of chromatin condensation usually correlate inversely with the levels of transcription. The mechanistic links between chromatin condensation and RNA polymerase II activity remain to be elucidated. In the present work, we sought to experimentally determine whether manipulation of chromatin condensation levels can have a direct effect on transcriptional activity.
  
  
  RESULTS
  We generated a U-2-OS cell line in which the nascent transcription of a reporter gene could be imaged alongside chromatin compaction levels in living cells. The transcripts were tagged at their 5' end with PP7 stem loops, which can be detected upon expression of a PP7 capsid protein fused to green fluorescent protein. Cycles of global chromatin hypercondensation and decondensation were performed by perfusing culture media of different osmolarities during imaging. We used the fluorescence recovery after photobleaching technique to analyse the transcriptional dynamics in both conditions. Surprisingly, we found that, despite a drop in signal intensity, nascent transcription appeared to continue at the same rate in hypercondensed chromatin. Furthermore, quantification of transcriptional profiles revealed that chromatin decondensation was accompanied by a brief and transient spike in transcriptional output.
  
  
  CONCLUSIONS
  We propose a model whereby the initiation of transcription is not impaired in condensed chromatin, but inefficient elongation in these conditions leads to the accumulation of RNA polymerase II at the transcription site. Upon chromatin decondensation, release of the RNA polymerase II halt triggers a wave of transcription, which we detect as a transient spike in activity.
  
  
  CONCLUSIONS
  The results presented here shed light on the activity of RNA polymerase II during chromatin condensation and decondensation. As such, they point to a new level of transcriptional regulation.
authors.affiliation: Institute of Cellular Biology and Pathology, First Faculty of Medicine, Charles University, Prague, Czech Republic.|BIOCEV and Department of Cell Biology, Faculty of Science, Charles University, Vestec u Prahy, 252 50, Czech Republic.
authors.forename: Viola
authors.initials: V
authors.lastname: Vaňková Hausnerová
authors.affiliation: Institute of Cellular Biology and Pathology, First Faculty of Medicine, Charles University, Prague, Czech Republic.|BIOCEV and Department of Cell Biology, Faculty of Science, Charles University, Vestec u Prahy, 252 50, Czech Republic.
authors.forename: Christian
authors.initials: C
authors.lastname: Lanctôt
mesh_terms: D002460:Cell Line; D002843:Chromatin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D036681:Fluorescence Recovery After Photobleaching; D006801:Humans; D012319:RNA Polymerase II / Q000378:metabolism; D015533:Transcriptional Activation; D019076:Transgenes
chemical_list: D002843:Chromatin; D012319:RNA Polymerase II
keywords: Chromatin; Decondensation; Hyperosmolarity; Live cell imaging; Transcription