Backbone structures of SUMO (green) superimposed on Rad60 (blue)
Human beings have tens of trillions of meters of DNA in their cells, and yet, in nearly every case, the genome is replicated only one time during cell division. Genetic defects can result in tumors, aging, and neurodegenerative diseases. Clearly, there is a great deal riding upon replication being absolutely accurate
To ensure the high-fidelity completion of replication, cells engage critical mechanisms that include cell cycle checkpoints and DNA repair. Cells also have a team at work to ensure stability during replication with SUMO, a small ubiquitin-like modifier, as the key player.
Now, in a recently published study, scientists at Scripps Research have found that the Rad60 DNA repair factor often inserts itself into the process as well. "By mimicking a particular surface feature of SUMO, Rad60 competes for binding to an essential enzyme of the SUMO machinery. Thus, Rad60 is a previously unidentified member of the SUMO team," senior author of the study and Scripps Research Associate Professor Michael "Nick" Boddy explains.
DNA repair protein Rad60 is a fission yeast (Schizosaccharomyce pombe), part of a unique protein family conserved from yeast to humans, and is essential for cell viability. Cells with a reduced level of Rad60 activity show sensitivity to a range of genotoxic stresses.
The new study, which combined structural, biochemical, and genetic analyses, showed that the two SUMO-like domains on Rad60 each bind to distinct components of the SUMO pathway.
"Even though the backbones in these two Rad60 domains are similar, their surfaces have been altered during evolution in such a way to maintain interaction with different parts of the SUMO pathway," says Boddy, who just received a prestigious Scholar Award from the Leukemia & Lymphoma Society. Boddy, with colleagues, first identified the Rad60 family in 2003 and was among the first to characterize SUMO in 1996.
In the study, the team used x-ray crystallography to determine the structure of a Rad60 SUMO-like domain at an ultra high-resolution, which is within the top half a percent of all published structures. "This resolution allowed us to develop and use a novel technique utilizing the Scripps Research supercomputer to solve the structure within a few weeks, which would have taken years if attempted on a desktop computer," said Andy Arvai, a scientific associate in the Tainer lab, which partnered with Boddy's lab on the study.
"The structural data allowed us to clearly understand how mimicry takes place and its importance in the SUMO pathway," said Scripps Research scientist Jeff Perry, one of the lead authors of the study. "In this case, we know that changing a single amino acid can break the binding. When you disrupt this interface, it creates instability and once that happens, the integrity of the genome can't be protected."
Scripps Research scientist John Prudden, another lead author of the study, added, "SUMO dysfunction is implicated in cancer and aging. Right now there is more to discover in terms of Rad60 mimicry - it's involved in other interactions within the SUMO pathway - so we want to know how it might affect those partners. And because these partners are implicated in disease, understanding the role of these interactions could be important for the clinical side of things."
As the teams bring their research to the next stage, its work will continue to rely on the state-of-the-art technology at Scripps Research. But running and maintaining this technology comes at a high cost.
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