Our Science – Bonner Website
William M. Bonner, Ph.D.
Each human cell contains about four billion base pairs (bp) of DNA divided into 23 pairs of chromosomes, the largest of which contains about 250 million bp. Maintaining the integrity of these long DNA double-helicies is essential to cell health, and considerable numbers of proteins are involved in various aspects of this task. One of the most serious threats to chromosomal integrity is the double-strand break (DSB) in which both DNA strands are broken in the same vicinity. If left unrepaired, such a break could lead to the loss of chromosomal fragments during mitosis. If erroneously repaired, fragments of different chromosomes might be joined together, leading to disruption of gene control and perhaps resulting in cancer. For example, chronic myelogenous leukemia is invariably associated with the inappropriate joining of DNA fragments from chromosomes 9 and 22. This translocation brings together parts of two genes to form the bcr-abl fusion gene, which encodes a disregulated protein tyrosine kinase, predisposing the cell to neoplasticity. Thus accurate rejoining of broken DNA fragments is essential for cellular health.
The accurate and efficient rejoining of broken DNA fragments is facilitated by histone H2AX. H2AX is a member of the histone H2A family, one of five histone families that package and organize the eucaryotic DNA into chromatin. The basic subunit of chromatin is the nucleosome, consisting of a core of eight proteins, two from each of the H2A, H2B, H3 and H4 families, and an outer wrapping of about 140 bp of DNA. In normal human fibroblasts, H2AX accounts for about 10% of the H2A complement, a ratio which places an H2AX molecule in every fifth nucleosome on average.
Within minutes of the occurence of a DSB, many hundreds of H2AX molecules in the chromatin adjacent to the break are phosphorylated on ser 139 (γ-H2AX). This γ-H2AX-containing region forms a focal point for the accumulation of DNA repair and chromatin remodeling factors. This local protein accumulation may facilitate the accurate repair of DNA DSBs. DNA rejoining is approximately six-fold more rapid in normal mice compared to H2AX-null mice. Mice lacking H2AX are substantially more sensitive to agents which cause DSBs and are more cancer prone. Thus the presence of γ-H2AX foci may enable more rapid rejoining, favoring more faithful repair.
Current Research Interests:
Our group looks at H2AX in two ways. The first involves the roles that it plays in DSB recognition, DNA rejoining and chromatin remodeling. The second rests on the fact that H2AX phosphorylation is more sensitive than previous methods of DSB detection, making is a uniquely useful means of uncovering novels roles for DSBs in cellular metabolism.
Our current program encompasses both basic and applied research projects and focuses on three areas. The first is a translational project investigating the parameters necessary to make H2AX a useful biodosimeter in humans and other animals. Two developments have given considerable impetus this project. The first is the finding by us and others that γ-H2AX is formed in tissue culture cells as a response not only to agents that directly cause DNA DSBs but also to those that indirectly cause DSBs by interfering with cellular metabolism. As many of the agents used for cancer treatment fall into this latter category, this finding greatly increases possible roles for γ-H2AX as a biomarker for drug responses. The second important development is the formation of a multi-disiplinary NCI team to expedite the clinical evaluation of new therapeutic and imaging agents, so-called phase zero trials. γ-H2AX formation is being examined as a possible biodosimeter in these studies. We are examining γ-H2AX formation in mouse models and are involved in several clinical protocols either approved or being approved in the phase zero trials. This work is important because it will permit clinicians to obtain immediate feedback from the cells of an individual patient, feedback which can then be used to tailor the treatment to that patient, improving their survival. The ultimate goal of this initiative is to develop γ-H2AX detection into a useful tool for human health.
The second project involves the use of γ-H2AX to probe for interactions between cells in animals. In vitro studies by us and others have shown that cells that have been exposed to ionizing radiation affect the viability and the presence of DNA damage products in unexposed cells that contact the exposed cells or media from the exposed cells, a controversial phenomenon known as the bystander effect. We have shown that a robust bystander effect is present in artificial human skin tissue. Working from the hypothesis that the bystander effect is an example of a larger phenomenon of communication among damaged and healthy cells, we have demonstrated that media from tumor cells induces responses in bystander cells indistinguishable from those induced by media from irradiated cells, and are currently demonstrating the ability of other stresses to induce bystander responses. In addition, this communication also takes place in the intact animal. We are finding that the presence of a human tumor implanted into nude mice leads to higher levels of DNA damage in certain other cells in the mouse. We are determining when and how this communication takes place in the animal and how this might be useful for human health. This work is important because it may increase the ways information can be obtained from individual patients. It is possible that by monitoring certain easily obtained tissues of a patient, information can be obtained about events in other less accessible tissues, perhaps leading to early detection of disease and cancer.
The third area focuses on the structure and dynamism of the γ-H2AX focus. We are developing tools that will permit us to study its substructure and how it changes with time. We have shown for example, that in yeast, Mre11 does not bind to the whole γ-H2AX domain, but is concentrated next to the break site. Studies such as these will complement other findings concerning the interactions of various proteins with γ-H2AX, leading to a greater understanding of DNA DSB repair and the maintenance of genome stability. Such understanding will also provide the basis for understanding the important parameters involved in utilizing γ-H2AX foci as a biodosimeter.
This page was last updated on 11/25/2013.