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Title: New tools for quantitative analysis of nuclear architecture
Author: Russell, Richard Anthony
ISNI:       0000 0004 2684 0171
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2010
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The cell nucleus houses a wide variety of macromolecular substructures including the cell’s genetic material. The spatial configuration of these substructures is thought to be fundamentally associated with nuclear function, yet the architectural organisation of the cell nucleus is only poorly understood. Advances in microscopy and associated fluorescence techniques have provided a wealth of nuclear image data. Such images offer the opportunity for both visualising nuclear substructures and quantitative investigation of the spatial configuration of these objects. In this thesis, we present new tools to study and explore the subtle principles behind nuclear architecture. We describe a novel method to segment fluorescent microscopy images of nuclear objects. The effectiveness of this segmentation algorithm is demonstrated using extensive simulation. Additionally, we show that the method performs as well as manual-thresholding, which is considered the gold standard. Next, randomisationbased tests from spatial point pattern analysis are employed to inspect spatial interactions of nuclear substructures. The results suggest new and interesting spatial relationships in the nucleus. However, this approach probes only relative nuclear organisation and cannot readily yield a description of absolute spatial preference, which may be a key component of nuclear architecture. To address this problem we have developed methodology based on techniques employed in statistical shape analysis and image registration. The approach proposes that the nuclear boundary can be used to align nuclei from replicate images into a common coordinate system. Each nucleus and its contents can therefore be registered to the sample mean shape using rigid and non-rigid deformations. This aggregated data allows inference regarding global nuclear spatial organisation. For example, the kernel smoothed intensity function is computed to return an estimate of the intensity function of the registered nuclear object. Simulation provides evidence that the registration procedure is sensible and the results accurate. Finally, we have investigated a large database of nuclear substructures using conventional methodology as well as our new tools. We have identified novel spatial relationships between nuclear objects that offer significant clues to their function. We have also examined the absolute spatial configuration of these substructures in registered data. The results reveal dramatic underlying spatial preferences and present new and clear insights into nuclear architecture.
Supervisor: Freemont, Paul Sponsor: Medical Research Council
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral