Extreme conditions of life are those that exceed conditions for growth and reproduction that are optimal for the majority of organisms. Organisms that thrive in or require such conditions are termed extremophiles.

There are many different classes of extremophiles, each corresponding to the way its environmental niche differs from those of the majority of terrestrial mesophile organisms. These classifications are not exclusive, thus many extremophiles fall under multiple categories. Desiccation and ionizing radiation are one of the most severe challenges to a cell [1]. Both cause double-strand breaks of DNA which is the most severe form of genomic damage [2].

Best known among extremophiles is the bacterium Deinococcus radiodurans. It is at the same time the most remarkable microbe due to the fact that it can survive extremely high exposures to desiccation and ionizing radiation which shatter its chromosome into hundreds of short DNA fragments [3, 4]. The bacterium owes its survival to a DNA repair process that accomplishes an efficient and precise reassembly of the DNA fragments. The mechanism of repair has been a mystery for decades.

However, it has recently been described as a two stage process: in the first stage most fragments reassemble in a process called ‘extended synthesis-dependent strand annealing’ while the second stage involves maturation of circular chromosomes by RecA-dependent crossovers [5]. Even though further details of the mechanism are still to be described, the general events taking place when DNA repair is concerned are now known and some others have been excluded. However, the exact course of events taking place in Deinococcus radiodurans after exposure to extreme conditions is far from understood and calls for further investigation. Namely, after shattering the chromosomal DNA into fragments due to irradiation, a latency period has been observed when neither DNA repair nor DNA synthesis are detected [5]. Following this is the period of massive DNA fragment reassembly and synthesis ending with cell division at 80-90 % survival [5]. All these events demand large amounts of energy and substrates that should be either stored within the cell before or during desiccation/irradiation or provided in the early stages of repair (i.e. latency period) by a mechanism so far unknown. Furthermore, macromolecular machinery of the D. radiodurans should be ready for the repair and synthesis events. The mechanism of survival of the crucial proteins is unknown as well.

Therefore, we in collaboration with Radman/Zahradka group intend to undertake original research to address the questions of:

1. Energy storage in the Deinococcus radiodurans before and/or during desiccation.
2. The complete description of biosynthetic events of both small molecules and macromolecules after periods of desiccation and ionizing radiation.
3. The mechanism of survival and identification of certain proteins that enable the cell to start the recovery process immediately after the end of the exposure to desiccation or ionizing radiation.
4. These questions will be addressed by various approaches of experimental biophysics in order to lead to better understanding of further details of the survival under extreme conditions.

Besides DNA, the other macromolecules in the cell also face the challenge of surviving the extreme conditions. The mechanisms by which certain proteins survive are, however, not understood, neither is which proteins do so. It is, therefore, our goal to study the adaptation of Deinococcus radiodurans proteins to survive extreme conditions such as desiccation or high temperatures also by using theoretical approaches. By extensive analysis of several extremophile proteomes, we have so far found that they are abundant in low-complexity regions. The next step is to study the structure and molecular dynamics of these proteins under both normal conditions of life and extreme conditions of desiccation and high temperature. The present project will make use of various software packages for protein structure prediction and analysis, as well as of molecular dynamics simulations to study the structural motifs and stability properties of extremophile proteins.

[1] Mattimore, V & Battista, J.R. Radioresistance of Deinococcus radiodurans: functions necessary to survive ionizing radiation are also necessary to survive prolonged desiccation. J. Bacteriol. 178, 633-637 (1996).

[2] Krasin, F. & Hutchinson, F. Repair of DNA double-strand breaks in Escherichia coli which requires recA function and the presence of duplicate genome. J. Mol Biol. 116, 81-98 (1977).

[3] Minton, K.W. DNA repair in the extremely radioresistant bacterium Deinococcus radiodurans. Mol. Microbiol. 13, 9-15 (1994).

[4] Battista, J.R., Earl, A.M. & Park, M.J. Why is Deinococcus radiodurans resistant to ionizing radiation? Trends Microbiol. 7, 362-365 (1999).