BioWorld International Correspondent

LONDON - A new gene, which is related to a nematode gene involved in the biological clock, has been proved to also play a role in sensing and repairing DNA damage.

This surprising discovery could lead to new approaches for developing anticancer agents. It also may explain some odd observations that no one has been able to make sense of before - such as the finding that chemotherapeutic drugs may work better when given at certain times of day.

Simon Boulton, head of the Cancer Research UK DNA Damage Response laboratory in South Mimms, UK, told BioWorld International: "We have identified a new human gene that is important for preventing accumulation of damage in our cells. Understanding how this gene functions, and how it is regulated by other processes in an animal, such as the biological clock, is likely to shed new light on why this particular pathway is mutated in many cancers."

The research is reported in Nature Cell Biology in a paper titled: "HCLK2 is essential for the mammalian S-phase checkpoint and impacts on Chk1 stability."

In 2005, Boulton and his colleagues reported on the work they had done, characterizing a gene in the nematode worm Caenorhabditis elegans called CLK2. They showed that nematode worms lacking CLK2 had the same phenotype as worms deficient in ATR - a protein that detects problems during DNA replication and ensures that replication can continue.

Earlier studies by others had identified a mutation in CLK2 as responsible for a radiation-sensitive mutant of C. elegans, as well as suggesting that this gene played a role in determining lifespan.

"It was only in 2001, when the actual molecular mutation in the radiation-sensitive mutant was identified, that people realized that mutations in the same gene were responsible for both of these phenotypes," Boulton said.

In retrospect, he added, this was the point at which it first became clear that the DNA damage response machinery and the biological clock are, in fact, intimately linked. The paper in Nature Cell Biology takes this thinking further, with proof that the human homologue of CLK2, a gene called HCLK2, interacts with proteins known to play important roles in sensing and repairing damage to DNA.

Although HCLK2 showed some sequence similarity with CLK2, Boulton and his colleagues were by no means convinced that the two genes had similar functions. To find out the role of HCLK2, they raised antibodies to the gene, with the aim of identifying which proteins it associated with.

"When we identified the components that came out with HCLK2, it turned out that one of them was ATR," said Boulton. "This was a really important development - this was the first physical interaction with a protein that had been described for any CLK2 in any species. Together with the fact that we had shown in C. elegans that CLK2 and ATR mutants have the same phenotype, this finding suggested that HCLK2 functioned with ATR early in the signalling cascade that responds to DNA damage."

Further experiments allowed the team to show that HCLK2 also interacted with several other proteins known to play a role in sensing and repairing DNA damage, including Chk1, which has been shown to be mutated in some cancers.

Next, the team went on to analyze the consequences of eliminating HCLK2 from human cells, by knocking out the RNA message for the gene. Again, they found that these cells, as in the parallel experiment in C. elegans, showed many features of cells that lacked ATR.

Additional investigations pinpointed more abnormalities in the HCLK2-deficient cells. "We showed that these cells are defective for two major responses that are important for repair of DNA, Boulton said. "The first is recruitment of a protein called Rad51, which is critical for accurate repair via homologous recombination. The second is a repair pathway, which has been implicated in the disease syndrome called Fanconi anemia, and which has recently been shown to respond to DNA replication problems."

The team also worked out that, in HCLK2-deficient cells, the kinase Chk1, which functions downstream of ATR, becomes degraded prematurely. "We think that the role of HCLK2 in cells is to prevent the untimely degradation of Chk1, which is absolutely critical for many DNA damage repair responses," Boulton said.

Future investigations will probe whether HCLK2 is a biological clock gene in humans.

The study also throws new light on previously unexplained observations. For example, said Boulton, mice that lack biological clock genes (including genes called Clock 1, Period 1 and Period 2) develop cancer. Researchers have also observed that some of the Period proteins are aberrantly expressed in some colon cancers and breast cancers.

"We can now go back to findings such as these and investigate them from a different angle," Boulton said. "It will also be important to find out if the response to DNA damage is in some way linked to the daylight cycle. Sleep is critical to recovery, and maybe that is at the level of DNA repair as well."