Imaging chromatin transitions using high performance microscopy

Lead Research Organisation: University of Dundee


The DNA sequence that encodes the information required to specify many organisms, including humans is known. This information exists as functional units referred to as genes, and it is normally the case that in a specific context only a subset of genes are used. Understanding how genes are activated and switched off at the appropriate times represents a key challenge to understanding how healthy organisms develop normally and what goes wrong during disease.

A key step in the activation of genes is the generation of an RNA copy of the DNA gene by a protein called RNA polymerase. The ability of RNA polymerase to gain access to DNA is normally prevented by packaging of DNA with proteins to for a condensed structure termed chromatin. It is becoming apparent that the unfolding of chromatin during activation and the subsequent refolding represent important points of control. However, understanding what occurs during the regulation of individual genes is to some extent limited by the experimental techniques that are available.

In this proposal we propose to use very recent developments in high performance microscopy to directly track the motion of a single gene both when it is switched off and when it is on. This type of visual information is very informative, providing a measurement of the extent that genes are unfolded and for how long they are unfolded during the course of RNA synthesis.

We hope that the observations we make will help to fill gaps in our understanding of a process which is fundamental to most biological systems.

Technical Summary

The genomes of eukaryotes exist predominantly as chromatin. It's clear that the structure of chromatin is not static but subject to dynamic regulation. Major insight into chromatin organisation has been provided by biochemical and structural studies performed with isolated components. However, for complex rearrangements, such as those occurring during transcription, it is difficult to know whether effects observed in vitro truly reflect a process that occurs within cells. While it is possible to determine the positions of factors such as polymerases and nucleosomes along a DNA sequence using genomic approaches, they do not provide high resolution structural information in real time. Advances in the spatial and temporal resolution of fluorescent light microscopy make this approach far better suited for the study of chromatin rearrangements. Here we propose to adopt these approaches to study the changes in the conformation of a gene during the course of transcription. This will provide new insight into the folding of nucleosome arrays and the extent to which nucleosomes are removed from DNA during the course of transcription. As such it will provide a new route to addressing the long standing issue of how polymerases transit chromatin.

Planned Impact

It is envisioned that the major beneficiaries of this work will be academics. These will fall into those interested in genome organisation, transcriptional regulation and chromatin structure. In order to ensure that these groups of academics have the opportunity to engage with this research our work will be pretend at national and international research symposia. We will publish our work in open access publications and ensure a description of our research is kept up to date on our laboratory web site, both at a level understandable by experts and the general public.

There is also a technological aspect to our work in that it pushes in vivo 3D tracking to the limits of what is attainable. This involves iterative cycles of adjusting hardware and data analysis tools. The experience we gain is likely to be of interest to organisations developing subcellular imaging for a broad range of approaches. Our ethos will not be to restrict access to such developments in order to commercially exploit them, but to openly divulge what we learn. The involvement of the OMX technologist Markus Posche in the project provides one way of passing on this expertise to other users of the OMX in Dundee (which include users from other Scottish Universities). In addition, these technological developments will be passed on by publication and presentation at meetings. In this respect Jason Swedlow is well placed to pass on knowledge as an instructor at international workshops on microscopy.

The application of new systems for analysing high resolution images has relevance to community based projects such as the Open Microscopy Environment (Nature Methods 9:245) a collaborative effort to produce tools to support data management for light microscopy. We will interact with the developers of this project to enable new approaches to be incorporated. Open disclosure does preclude commercial development and this is illustrated by the activities of companies such as Glencoe Software, which has strong links to Dundee, specialising in the application of expertise in data management and analysis.

We have a track record of communicating our results to the public through participation in open days, visiting schools, producing movies that describe our research


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Description We developed a work flow to track the motion of individual genetic loci in live cells. This was used to track the motion of multiple chromosomal regions. Large variations in motion were detected over time together with differences in the way in which chromatin moves at different loci. Different behaviors at the same loci were however not inherited through cell divisions.
Exploitation Route Our work is published and has been cited. The raw data is accessible for analysis via the idr website described above. We have been contacted by David Holcman who is interested in analyzing the data.
Sectors Agriculture, Food and Drink,Healthcare,Pharmaceuticals and Medical Biotechnology

Description Structure and function of chromatin remodelling ATPase's and their dysfunction in in human disease.
Amount £1,232,837 (GBP)
Funding ID MR/S021647/1 
Organisation Medical Research Council (MRC) 
Sector Public
Country United Kingdom
Start 03/2019 
End 03/2024
Description Inner Space Outer Space at the Mills Observatory 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact The Mills Observatory in the heart of Dundee provides a unique location to develop, trial and offer public engagement activity that is immersive, uses multimodal-presentation techniques through a room-filling visual presentation screen, allowing visitors of all ages and abilities to engage with and explore the wonders of our micro and macro cosmos.

This ambitious project is funded by the Centre for Gene Regulation and Expression at the University of Dundee by a Strategic Grant from the Wellcome Trust in partnership with Leisure and Culture Dundee and Dundee City Council.

The project was initiated and is managed and run by the Dundee Imaging Facility the only multidisciplinary technology hub at the University which currently works with researchers from all of the STEMM subjects.

It is being developed in collaboration with the School of Computing Computing's Augmentative and Alternative Communication (AAC) Research Group with input from Life Scientists, Physicists, Mathematicians and Astronomers from across the University.

The new facilities allow accessible, interactive exploration of University research across all these disciplines. The technology used will also ensure that all audiences, especially people with impairments, can experience and interact with the presentations.
Year(s) Of Engagement Activity 2015,2016
Description Open Doors day 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? Yes
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Around 200 people attendend, members of the lab presented and interactive display describing protein puyrification, and participated in "coffeee with a scientist" session.

Hopefully, interactions with members of public had some impact, but little evidence of this yet.

Film crew attended gathering material for documentary describing research activity in Dundee
Year(s) Of Engagement Activity 2011,2012,2013,2014