BioWorld International Correspondent

LONDON - Investigations into how cells respond to DNA damage have trained a spotlight on a protein that plays a role in regulating the functions of the well-known tumor suppressor protein, p53.

The protein, until now not thought to be involved in cellular responses to DNA damage, has the potential to become an attractive target for cancer drugs. Researchers also believe that it might be possible to predict how particular tumors or individual patients will respond to radiotherapy or chemotherapy by measuring levels of that and other proteins in their cells.

Stephen Jackson, professor of biology at the Cancer Research UK Gurdon Institute in Cambridge, UK, told BioWorld International: "Our work provides new molecular insights into how cells respond to DNA damage. In particular, it takes our understanding of the regulation of the critical tumor-suppressor protein, p53, to a new level of sophistication."

A report of the study appears in the Dec. 15, 2005, issue of Cell in a paper titled "hnRNP K: An HDM2 Target and Transcriptional Coactivator of p53 in Response to DNA Damage." The first author of the paper is Abdeladim Moumen.

Understanding how cells respond to DNA damage is crucial to understanding how cancer arises in the first place. If normal cells are treated with an agent that causes DNA damage, they respond by stopping the cell cycle, gaining time to repair the damage before continuing cell division. Animals with defects in the pathways controlling repair of DNA damage often are at a higher risk of developing cancer. Variation in the ability to respond to and repair DNA damage also might be linked with differences in how cells respond to radiotherapy or chemotherapy that inflict DNA damage.

Jackson and his colleagues set out to investigate which proteins in the proteome become altered in level following DNA damage caused by irradiation, using a new method called 2-dimensional difference gel electrophoresis.

The team identified one of the proteins that increased in quantity as heterogeneous nuclear ribonucleoprotein K (hnRNP K). Experiments showed that the molecule was indeed part of the DNA damage-response pathway: If the researchers inhibited the protein kinases that activate that pathway, levels of hnRNP K failed to increase following irradiation.

Jackson said, "We also showed that if you deplete cells of hnRNP K, they lack the cell-cycle checkpoints that normally operate following DNA damage."

The team went on to determine the mechanism by which levels of hnRNP K are allowed to rise following DNA damage. They found that normal cells are making hnRNP K constantly, but that it is degraded quite rapidly. In cells that have suffered damage, however, degradation of hnRNP K stops, allowing levels to rise rapidly.

"This mechanism is called regulated proteolysis,'" Jackson said. "It allows very rapid changes to protein levels - we can see changes in hnRNP K levels within 15 minutes of irradiating the cells."

Further experiments established that a protein called HDM2 (also known as MDM2) plays a role in the proteolysis of hnRNP K levels.

HDM2 is a known oncogene; it is overexpressed in several human cancers. Previous work by others had shown that one of the prime roles of HDM2 is to regulate p53. Like hnRNP K, levels of p53 rise after the cell has suffered DNA damage and researchers have shown that this rise is needed for cells to implement cell-cycle checkpoints following DNA damage.

The final part of the study questioned how hnRNP K affects the cell-cycle checkpoints that depend on p53.

It is known that p53 is a transcription factor; once produced, it binds to its target genes and switches them on. Jackson's group established that hnRNP K is needed for that process to occur. Jackson said: "When we knocked down the expression of hnRNP K, we found that although p53 is still induced after DNA damage, it is not able to do its job as a transcription factor. It turns out that both hnRNP K and p53 co-assemble on the p53 target genes after DNA damage. hnRNP K provides an additional level of regulation. It is not just a simple one-switch mechanism."

Drugs already are available that target HDM2 and its interaction with p53, Jackson said. "It may be that we can arrive at a similar situation with respect to hnRNP K in some years' time, but this is very speculative," he concluded.