Ray Sachs, UCB. Computational Radiation Biology
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NSF Chromosome aberration grant

Sachs PI. 1996-2004. Abstract and Specific Aims. Abstract of creativity extension

Abstract

The proposal is to model chromosome aberrations in live mammalian cells, emphasizing implications for large-scale chromosome geometry and for the biology of ionizing radiation damage. Formation of chromosome aberrations after induction of DNA double strand breaks by radiation during the interphase part of the cell cycle will be analyzed, using probabilistic models for DNA geometry, motion, and reactions at the macromolecular level. For example, we will analyze illegitimate recombinations, involving the exchange of fragments between two different chromosomes. Polymer models will be used for large-scale chromosome structure and motion. Comparisons of the effects of densely ionizing radiation to those of sparsely ionizing radiation, relevant to chromosome geometry and to biological dosimetry, will be made. Statistical questions which arise when very detailed experimental information about specific individual chromosomes is made available by modern fluorescent in situ hybridization techniques will be answered by extending CAS (chromosome aberration simulator) Monte-Carlo computer software. The study will involve developing computer algorithms, explicitly solving stochastic process models, and collaborating with experimental groups.
The grant will study chromosome aberrations mathematically and biologically. When ionizing radiation damages cells, as occurs for example in tumor radiotherapy or radiation accidents, one gets breakage and large-scale reshuffling of DNA molecules. The resulting chromosome aberrations have been implicated as symptoms and/or causes of most major radiobiological effects, and are especially important in biodosimetry, the estimation of past exposure to radiation dose by looking at residual cellular damage. They are also a window on fundamental biology because they are influenced by, and symptomatic of, chromosome geometry and motion. The recent advent of chromosome painting has dramatically increased the amount of information obtained from aberration experiments. A very rich and colorful variety of aberrations can now be observed and trying to understand the number and kind of reshufflings one sees leads to mathematically nontrivial problems, amenable to computer simulations and/or "pen and paper" calculations. We will carry out the simulations and calculations. The results of the grant will facilitate systematic comparisons of radiation damage estimates made by different laboratories using different chromosome painting schemes. In addition, our mechanistic modeling of how chromosome aberrations develop in time may help attack the main mystery of radiobiology: how to extrapolate from the larger doses needed for statistically significant observed damage effects to the much smaller doses relevant to most human populations.

Specific Aims

Large-scale geometry of chromosomes in cell nuclei influences many molecular biology processes, including the formation of chromosome aberrations. We will analyze chromatin geometry, aberrations produced by ionizing radiation, and their interrelationships. The study will involve applying polymer theory, developing computer algorithms, and analytically solving stochastic process models. Close collaboration with various experimental groups will continue. The strongly interdisciplinary nature of the research will be maintained. The following detailed projects will be accomplished.
1.1. Chromosome Geometry. Polymer models of large scale chromatin geometry in mammalian cells during the interphase part of the cell cycle will be extended from those developed previously under the grant. The multivariate normal probability equations for an individual chromosome will be generalized, and the data analyzed in more detail for the statistics of distance distributions involving two defined points on a chromosome. Probabilistic models will be developed for recent, unpublished data which may indicate multiple backbones, or an excluded volume effect corresponding to a self-avoiding random walk, or systematic tethering to an extra-chromosomal structure. Models analyzing all 46 chromosomes within a nucleus will be developed, using recent data on the degree of overlap of two different chromosomes or the location of a single chromosome relative to the nuclear envelope.
1.2 Chromosome Aberrations. Formation of chromosome aberrations by ionizing radiation will be studied mathematically, emphasizing the underlying DNA structure and reaction kinetics at the molecular level. Experimental comparisons will emphasize the flood of recent data obtained by chromosome painting. Previously developed Markov models and Monte Carlo computer simulations will be extended. They will be used for more detailed predictions and comparisons with experiments of: (a) relative yields for many different aberration categories; (b) their per-cell statistical distributions; and (c) their dose-response curves.
Creativity Extension 2002-2003
Sachs PI, but the application was mainly written by my postdocs, Drs. J. Arsuaga and M. Vazquez

Summary

The PI's group in the UCB math department studies DNA radiation damage, especially chromosome aberrations. Continuing aberration studies, analyzing related microarray data, and analyzing DNA topology is proposed.

Broader impact. The main general scientific motivations for studying radiation-induced chromosome aberrations are their implications for DNA repair/misrepair pathways and their intimate relation to chromosome geometry in the interphase nucleus, which in turn has implications for gene regulation. The main practical applications, which are beyond the scope of the present proposal, are to risk estimation, diagnostics, and radiation biodosimetry.

Vertical integration in the group, from freshmen to senior faculty, is vigorous. The PI, Professor RK Sachs, is very active in mathematical and computational biology, having recent joint papers with many different experimental biologists worldwide. Drs. Mariel Vázquez and Javier Arsuaga, for whom the funds are being requested, are postdocs who joined the group in August 2000. They both hold recent mathematics PhD degrees with mathematical structural biology and computational biology emphasis. Their work is fully interdisciplinary and involves continuous interaction with experimental biologists. The group also strongly promotes interactions between research and teaching, emphasizing undergraduate research. Currently two graduate and five undergraduate students are being mentored. A number of peer-reviewed papers crediting the grant have graduate and/or undergraduate student coauthors. Diversity in the group is represented by two women, three members of Hispanic origin, and four of Asian origin.

Project. Chromosome aberrations involve reshuffling, based on DNA damage/repair/misrepair pathways, of broken chromosome pieces. Aberrations can be identified by recently developed combinatorial painting techniques (mFISH and SKY), in which each homologous chromosome pair is assigned its own color. The resulting patterns are so complicated that they call for mathematical and computational analysis. The group develops and uses analytic and Monte-Carlo computer methods to analyze aberration data, including mFISH data. In collaboration with two experimental biologists, Professor M. Cornforth (U Texas) and Dr. K. Greulich-Bode (Heidelberg), the distribution of radiation-induced breakpoints within the genome will be analyzed at different levels of resolution. mFISH data gives information at the Mb level. Statistical methods and previously developed computer software (CAS, Chromosome Aberration Simulator) will be used to analyze mFISH data, spatial associations for chromosome pairs participating in aberrations, and distribution of aberration breakpoints among individual chromosomes as well as within each chromosome. Currently there is a major effort to integrate cytogenetic data, such as that given by combinatorial painting and banding techniques, with the data arising from the human genome draft sequence. The project will seek correlations between chromatin breakpoints and large-scale genomic structure, applying information obtained in the human genome project to our own specialty. Being able to show systematic correlations would have major implications for theories of DNA repair and misrepair in mammalian cells.

There are two standard models of chromosome aberration production: breakage-and-reunion, and recombinational misrejoining. The group's recent work compared mFISH data to CAS results using these standard models and an additional instability model that better accounts for observed high levels of complex aberrations. The group's new mathematical method, which assigns a so-called "cycle structure" to each mFISH pattern, is now being used by experimentalists. A further analysis of cycle structure for additional tests of the three biophysical models is proposed.

Paralleling studies on radiation-induced chromosome aberrations, a complementary approach will be started in collaboration with Professor D. Brenner (Columbia University) and Dr. S. Amundson (NIH). cDNA microarray data obtained by Dr. Amundson will be analyzed to recover genetic regulatory networks that respond to ionizing radiation.

The tangle model (Sumners et al., 1995), based on mathematical knot theory and low-dimensional topology, can be successfully used in the topological analysis of site-specific DNA recombination. Topological analysis of Xer site-specific recombination will be continued. Computer algorithms which make the model more accessible to both experimental biologists and mathematicians will be extended.