49. ADAPTIVE IMAGE ANALYSIS OF DNA REPAIR DYNAMICS IN INDIVIDUAL CELLS
Department: Computer Science & Engineering
Research Institute Affiliation: Graduate Program in Bioinformatics
Faculty Advisor(s):
Yoav Freund
Primary Student
Name: Boyko B Kakaradov
Email: bkakarad@ucsd.edu
Phone: 650-387-7631
Grad Year: 2012
Abstract
The cells in our bodies are constantly challenged with damage caused by intrinsic or environmental sources. DNA double strand breaks (DBSs) are one of the most dangerous types of cellular damage, as they can cause substantial rearrangements in the genome. We aim to understand how dynamic interactions in the DSB response network coordinate damage detection and the choice of repair mechanism1. To address this, we use time-lapse microscopy and measure the temporal and spatial dynamics of key components of the DNA damage response in individual, living cells. We have created a clonal cell line expressing the DSB marker 53BP1 fused to yellow fluorescent protein, and Rad52, a protein mediating DSB repair by homologous recombination, fused to the red fluorescent protein mCherry. Both proteins form complexes around double strand breaks, which can be observed by fluorescence microscopy in living cells 2, 3.
Manual analysis of these time-lapse images is very time-consuming. Instead, we are working on a tool that automates the process by segmenting and tracking cell nuclei. The standard approach to cell segmentation uses a dedicated fluorescence channel to image a DNA marker such as H2B-GFP. This simplifies the localization of nuclei, but reduces the number of channels available for concurrent imaging of network proteins. In contrast, we are developing a method that uses the existing phase field contrast and fluorescence channels. While localizing the nuclei in individual channels is challenging, our method achieves good segmentation results by combining visual information from several channels using adaptive filters trained with Adaboost.
References:
1. Su, T. T. Cellular responses to DNA damage: one signal, multiple choices. Annu Rev Genet 40, 187-208 (2006).
2. Pryde, F. et al. 53BP1 exchanges slowly at the sites of DNA damage and appears to require RNA for its association with chromatin. J Cell Sci 118, 2043-55 (2005).
3. Essers, J. et al. Nuclear dynamics of RAD52 group homologous recombination proteins in response to DNA damage. Embo J 21, 2030-7 (2002).
4. Y. Freund and R. E. Schapire. A short introduction to boosting. Journal of Japanese Society for Artificial Intelligence, 14(5):771--780, September, 1999.
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