Environmental Factor, November 2011, National Institute of Environmental Health Sciences
Crystallography reveals roots of DNA repair-linked neurodegenerative disease
By Robin Arnette
Percy Tumbale, Ph.D., is a postdoctoral fellow in Williams' lab and is lead author on the paper. She is a 2012 Fellows Award for Research Excellence winner. (Photo courtesy of Steve McCaw)
Williams is head of the Genome Stability Structural Biology Group within the Laboratory of Structural Biology at NIEHS. (Photo courtesy of Steve McCaw)
According to R. Scott Williams, Ph.D.(http://www.niehs.nih.gov/research/atniehs/labs/lsb/genome/index.cfm), the proteins that repair DNA are like the molecular sentinels that home in on damaged DNA and protect cells from environmental damage. He said oftentimes these same proteins are mutated in heritable neurodegenerative diseases and syndromes that predispose a person to cancer. When Williams came to NIEHS a year and a half ago, he focused on understanding these connections.
Williams and his research team determined how a protein known as aprataxin functions in yeast. Aprataxin participates in the repair of damaged DNA in humans, and mutations in the gene that encodes aprataxin (APTX) cause hereditary neurological disorders. Until recently, scientists didn't understand how aprataxin protected DNA or how the gene mutations shut down the protein. Williams and his colleagues uncovered how aprataxin recognizes and directly reverses DNA damage using X-ray crystallography. The results appeared online in the Oct. 9 issue of the journal Nature Structural and Molecular Biology. The findings have implications for human health.
Aprataxin reverses DNA damage
Williams explained that the final critical step in repairing damaged DNA is called ligation, which involves chemically joining broken strands of DNA together. Like many biological processes, ligation can fail when DNA ligases attempt to link normal DNA strands with strands that have been modified as a result of environmental DNA damage. Such failure results in the production of additional DNA damage called DNA adenylates. Aprataxin acts as a DNA ligase proofreader by repairing DNA adenylates, but mistakes in the protein can have deleterious effects in humans. Williams said mutations in aprataxin lead to deterioration in the cerebellum and a debilitating disorder known as ataxia with oculomotor apraxia type1 (AOA1).
“Although we have known that APTX mutations cause AOA1 for nearly a decade, researchers have not understood the molecular underpinning of how aprataxin works, or how mutations stop it from protecting our DNA,” he said. “Now, we have a hypothesis for how a number of different types of heritable mutations in aprataxin cause its inactivation and lead to AOA1.”
The Williams team took advantage of the high-throughput robotics infrastructure at NIEHS in the Laboratory of Structural Biology, to screen a large number of crystallization conditions for aprataxin bound to DNA. In a technical triumph, the group did something that's never been done before - crystallize a quaternary complex of the protein containing aprataxin, DNA, the DNA damage lesion 5'-adenosine monophosphate (AMP), and the metal cofactor zinc. Because Williams captured a molecular snapshot of aprataxin in action, he was able to rationalize how it recognizes and excises DNA damage.
“That's one of the beauties of structural biology,” Williams added. “By directly visualizing proteins and their complexes at work, you always gain critical insights into the functions of macromolecules that you'd never get using other methods.”
Cancer therapy and novel drug design
Figuring out how aprataxin fixes DNA ligation errors may also lead to potential advances in cancer research. Williams said that proteins involved in DNA repair are the Achilles' heel of rapidly dividing cancer cells. He believes that if scientists can target aprataxin with therapeutics, they may be able to augment standard cancer therapy regimens. The aprataxin work also provides a template that may be used for the development of small molecule compounds that zero in on and inhibit this enzyme class in humans.
Williams said that several different adducts, or cancer-causing molecules that chemically modify DNA, lead to ligation errors and adenylated DNA. He wants to find out which adduct is the most important one for aprataxin. In other words, are there certain types of damage that illicit a repair response from aprataxin? If so, how does that damage relate to the mechanism of neurodegenerative disease? Williams performed his experiments on the yeast version of aprataxin, but is currently extending his studies to work on the human form.
Citation: Tumbale P, Appel CD, Kraehenbuehl R, Roberson PD, Williams JS, Krahn J, Ahel I, Williams RS(http://www.ncbi.nlm.nih.gov/pubmed/21984210) . 2011. Structure of an aprataxin-DNA complex with insights into AOA1 neurodegenerative disease. Nat Struct Mol Biol; doi:10.1038/10.1038/nsmb.2146 [Online 9 October 2011].