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Transcriptional Traffic Control

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  2. Transcriptional Traffic Control
  3. Understanding Developmental Disruptions

Nucleosomes act as a roadblock to transcription, but there are a number of mechanisms by which RNA polymerase II can successfully bypass histone complexes. For example, chemical modifications to the ‘tails’ of histone subunits can enhance transcription, as can changes in DNA-protein interactions within a nucleosome. However, little is known about how these various factors collectively impact transcription.

A series of single-molecule experiments by Bintu et al. offer new insights into this process, yielding a multi-step model to explain how the polymerase complex negotiates its way past the nucleosome. The authors used an optical tweezers setup to monitor the pausing behavior of the RNA polymerase II elongation complex (EC) as it travels along DNA containing a single nucleosome positioning sequence (NPS), and identified three different NPS regions that they termed ‘entry’, ‘center’ and ‘exit’. Histone manipulation revealed that the absence of tail domains or introduction of conditions simulating acetylation of histone lysine residues specifically reduced pausing in the entry region, whereas point mutations to the H3 and H4 histones that loosen their association with DNA decreasedpausing at the NPS center. These effects were subsequently replicated in experiments that tested how different modifications modulate the ‘tightness’ of DNA wrapping around the nucleosome complex.

Previous work suggests that transcription is dependent on ongoing fluctuations in nucleosome structure, and that the EC will backtrack along the nascent RNA strand when forward progress is thwarted. The authors also observed this, and obtained evidence that the local DNA sequence can modulate this backtracking based on the extent to which newly-transcribed RNA forms stable secondary structure. The interplay between these various factors can thus collectively have a profound effect on the transcriptional dynamics within a given stretch of chromatin.1

Bintu, L. et al. Cell151, 738–749 (2012).

Understanding Developmental Disruptions

  1. Top of page
  2. Transcriptional Traffic Control
  3. Understanding Developmental Disruptions

Chromodomain helicase DNA-binding 7 (CHD7) is the vertebrate homolog of kismet, a fruit fly gene that plays a prominent role in body development via the regulation of homeotic gene expression. Accordingly, CHD7 mutations are linked with a host of developmental defects in humans, including a pattern of severe and potentially fatal abnormalities collectively described as CHARGE syndrome.

The CHD7 protein has proven challenging to purify and characterize due to its large size, but Bouazone and Kingston recently made important headway in clarifying its functional role. Previous evidence has indicated that CHD7 participates in chromatin remodeling, and the researchers confirmed that this protein can directly induce changes in nucleosome mobility via an ATP-dependent mechanism. Comparative assays with other remodeling factors indicated that CHD7 tends to act more like ISWI than hSWI/SNF, and exhibits a strong preference for acting on individual nucleosomes flanked by histone-free DNA and a partial dependence on histone tail interactions for efficient function.

Although CHARGE patients manifest a broad spectrum of CHD7 mutations, the majority of these are nonsense or frameshift mutations that result in a truncated protein. The researchers confirmed that such alterations would fatally impair this protein's capacity for histone remodeling, suggesting that CHD7 haploinsufficiency is responsible for the birth defects observed in these individuals. They also demonstrate varying functional impact for several different clinically-defined missense point mutations as well, highlighting the apparent importance of this protein's distinctive chromatin remodeling activity in regulating genes centrally associated with embryonic development.

Bouazone, K. & Kingston, R.E. Proc. Natl. Acad. Sci. USA, Published online 7 November 2012, doi: 10.1073/pnas1213825109.