Imaging of Repair Protein Dynamics on Radiation Tracks
The primary biological benefit of heavy ion radiotherapy (1) is the enhanced effectiveness of the applied carbon ions in the region of the tumor. At the cellular level, the underlying basis of this effect is the localized dose deposition leading to the production of clustered DNA lesions which are difficult to repair. These can be indirectly visualized in nuclei of mammalian cells by microscopy of immuno-stained repair proteins recruited to the damaged sites. Various DNA damage response proteins were shown to relocalize into foci directly at the sites of particle traversal (2 and ref. therein).
In this context the production of microscopically discernable regions of DNA damage using charged particles, combined with appropriate imaging approaches, provides a tool to study mechanistic aspects of ionizing radiation at the level of single cells. The analysis of the spatiotemporal dynamics of proteins at foci allows studying the hierarchy of protein recruitment and will help answering open questions related to the recognition and repair of DNA damage. The use of GFP-tagged repair proteins makes it possible to perform these studies in living cells. Real time measurements at a newly developed beamline microscope have shown that repair-related proteins are recruited to sites of damage within seconds (3).
On the other hand, the introduction of linear tracks of DNA damage within cell nuclei using particle beams parallel to the cell layer allows analyzing the spatial distribution of proteins along the ion trajectories of single traversing particles (4). Similar patterns of discrete protein clusters were generated irrespective of the ion beam applied or the immunodetected protein, suggesting the chromatin structure being responsible for the patterns observed. However, changes in track morphology with time and a constricted migration of foci were observed, probably in connection with lesion processing. Live imaging in connection with this type of studies will shed light into the movement of chromatin domains in the course of damage repair.
Summarizing, heavy ion radiotherapy has motivated basic research regarding the molecular events provoked in response to very localized but severe DNA damage. At the same time, in combination with newly adapted imaging techniques, the production of restricted regions of damage using heavy ions provides a valuable tool to study the organization of the DNA damage response.
Microscopic visualization of the extremely localized DNA damage induced in nuclei of mammalian cells following irradiation with accelerated ions. Immuno-fluorescence stained repair proteins accumulate at the lesions along the individual ion tracks traversing the nucleus of a human cell, appearing as parallel streaks. Red: DNA counterstain (Propidium Iodide). Green: Repair protein (Mre11).
(B.Jakob, M. Scholz and G. Taucher-Scholz, Radiat. Res. 2003)
1. D. Schultz-Ertner et al., Int J. Radiat. Oncol. Biol Phys 58, 631-40 (2004)
2. B. Jakob, M. Scholz and G. Taucher-Scholz, Int. J. Radiat. Biol. 78, 75-88 (2002)
3. B. Jakob, J. H. Rudolph, N. Gueven, M. Lavin and G. Taucher-Scholz, Radiat. Res. 163, 681-90 (2005)
4, B. Jakob, M. Scholz and G. Taucher-Scholz, Radiat. Res. 159, 676-84 (2003)
contact: g.taucher-scholz
gsi.de