Facts and pictures have been taken from www.nobelprize.org.
The 2015 Nobel Prize was jointly awarded to Tomas Lindahl, Paul Modrich and Aziz Sancar for their independent studies on different mechanistic pathways of DNA repair.
It is a well-known fact that our DNA is prone to damage either due to external factors such as X-rays and UV radiation or during cell replication where copies of DNA is made. Sometimes, the reactive metabolites released during physiological processes can also cause lesions in the DNA. Despite all these factors, the DNA manages to maintain its fidelity and integrity. The question was then asked: are there processes within the cell which correct and repair the damage done to DNA? This year’s Nobel Prize was given to those three people who conducted pioneering research which gave answers to the said question.
The mutations occurring in DNA is actually a double-edged sword. On one hand these mutations are responsible for Darwinian evolution but on the other hand it can also cause deadly diseases such as cancers and neurodegenerative disorders. These lesions in DNA can stop cell proliferation and can also instigate programmed cell death, which, in a way can be used to treat cancer.
In early 1970s Tomas Lindahl demonstrated that the chemical stability of DNA is challenged when exposed to chemical reactions such as hydrolytic deamination, oxidation and non-enzymatic methylation. Most importantly, he discovered that as a consequence of being exposed to such conditions, deamination of cytosine to form uracil occurs. This mutation poses a high risk of depletion of genetic material as the cytosine-guanine base pair gets replaced with uracil-adenine base pair. Further work led to the discovery of base excision repair process.
He single-handedly identified the E. coli uracil-DNA glycosylase (UNG) as the first repair protein that can correct base lesions. We know today that UNG is the founding member of a large family of proteins that orchestrate base excision repair (BER). He also went on to show that this enzyme is specific to DNA and keeps the DNA backbone intact during the process. We know today that BER corrects different forms of lesions that affect the bases without causing gross structural perturbation in the DNA structure.
Base excision repair also occurs in human beings and, in 1996, Tomas Lindahl managed to recreate the human repair process in vitro.
The process which repairs the damage done to DNA by UV radiation was discovered by Aziz Sancar. The mechanism is known as nucleotide excision repair (NER) process.
For his doctoral dissertation in 1970s, Sancar worked on photolyase, an enzyme which is responsible for the repair of UV-damaged DNA. The observation of this enzymatic activity, first shown by Stanley Rupert, was of profound importance, since it demonstrated for the first time the existence of DNA repair enzymes that could rescue UV-irradiated DNA. At first, the photolyase was just an activity in an extract, but in 1978 Sancar, then Rupert’s student, could clone the E. coli photolyase gene and amplify the gene product in vivo.
Upon graduating, when his work was not well accepted to land a post-doctoral position, Sancar joined Yale University School of Medicine as a laboratory technician. During this time, a second process had been discovered: a light-independent process to repair UV-damaged DNA, which came to be known as dark repair.
His colleagues at Yale had studied this process in detail since 1960s, using three UV-sensitive strains of bacteria that carried three different genetic mutations: uvrA, uvrB and uvrC. Early pioneering work had indicated that there existed processes which corrected the damage done to DNA due to UV-radiation, but the exact mechanistic process remained vague. Previous bacterial work had identified uvrA, uvrB, and uvrC genes in a search for mutations that impaired NER and hindered growth resumption after UV irradiation, but could not be examined in detail due to the lack of purified proteins.
Sancar, now at Yale, studied this dark system and as in his previous work began investigating the molecular machinery behind the process. In a few years, he was able to identify, isolate and characterise the enzymes coded by the genes uvrA, uvrB and uvrC. In a series of ground-breaking in-vitro experiments Sancar started to map the repair process of the enzymes, meanwhile securing a position at University of North Carolina because of his findings. He then went on to extend his work to humans, although the molecular machinery in humans is more complex than that in bacteria but, in chemical terms, nucleotide excision repair functions similarly in all organisms.
Paul Modrich, early on in his career had worked on a series of enzymes that affect DNA such as DNA ligase and DNA polymerase. During the course of his research he came across the enzyme Dam methylase which couples methyl groups to DNA.
Matthew Meselson, a molecular biologist at Harvard University, had constructed a bacterial virus with several occurrences of mismatching bases in the DNA. For instance, A could be placed opposite C, instead of T. When he let these viruses infect bacteria, the bacteria corrected the mismatches. He speculated that there must be a repair mechanism in place which corrected such mismatched pairs of bases, which can mostly occur during DNA replication.
Meselson and Modrich then collaborated to create a virus with a number of mismatches in its DNA and the dam methylase enzyme was used to add methyl groups to one of the DNA strands. They allowed the virus to infect bacteria where they observed that the bacteria corrected those mismatched pairs which were present in the strand that was not methylated. It was then concluded that the DNA mismatch repair process is a natural process which corrects mismatched pairs of bases and recognises the defect strand by its unmethylated state.
Over the next decade, Modrich conducted meticulous research mapping the repair process and recreating the pathway in-vitro; his work was published in 1989. The extension of this process to the human system has still many unanswered questions and is a matter of scientific interest.
Besides the above three mechanisms, there are many other repair systems in place which protect DNA from damage caused by the sun, cigarette smoke or other genotoxic substances; they continuously counteract spontaneous alterations to DNA and, for each cell division, mismatch repair corrects some thousand mismatches.
Although the research work done is more than two decades old, they have captured the limelight now because of the increasing importance of elucidating processes related to DNA- may it be transcription, translation or repair of DNA, as they are crucial to finding cure for many devastating diseases, cancer being one of them.
In fact, in many forms of cancer, one or more of these repair systems have been entirely or partially switched off. This makes the cancer cells’ DNA unstable, which is one of the reasons why cancer cells often mutate and become resistant to chemotherapy. At the same time, these sick cells are even more dependent on the repair systems that are still functioning; without these, their DNA will become too damaged and the cells will die. Researchers are attempting to utilise this weakness in the development of new cancer drugs. Inhibiting a remaining repair system allows them to slow down or completely stop the growth of the cancer. One example of a pharmaceutical that inhibits a repair system in cancer cells is olaparib.
Digressing from the topic discussed, I think it is important in my line of work i.e., being a scientist or a researcher, to have faith in ones’ ability and more importantly patience to keep going despite the many hurdles one may come across, the most important being when other contemporaries and peers believe you may have taken the wrong path. Nevertheless, it should also be taken with a pinch of salt as venturing into the unknown, 99% of time you could be headed in the wrong direction.
Or, in the words of Paul Modrich: “That is why curiosity-based research is so important. You never know where it is going to lead… A little luck helps, too.”