The finding, reported in the Dec. 17, 2004, issue of the journal Cell, provides the first link between a cell's DNA repair machinery and its DNA storage and retrieval machinery. The two processes have been studied independently, and each is essential for proper care and maintenance of the cell's genetic material, but until now there was little evidence of how the two might work together.
"We have brought together two fields that are essential for proper maintenance of DNA," said Xuetong "Snow" Shen, assistant professor in the Department of Carcinogenesis at M. D. Anderson. "It was generally understood there must be a connection between the two, but no direct connection had ever been seen. We have bridged that gap."
Many types of cancer, including human leukemias and lymphomas, have been linked to defects in DNA maintenance. Shen's lab studied a particular protein complex, called INO80, that regulates access to DNA. Inside cells, long strands of DNA are wound tightly around a series of proteins called histones. The combination of DNA and its associated proteins is called chromatin. The histone proteins help compact the DNA and help keep it organized within the chromosome, said Shen, but DNA tightly wound around histones is inaccessible. If DNA becomes damaged by radiation, reactive chemicals or ultraviolet light, for example, it must be repaired. But the bulky repair proteins need to gain access to the damaged areas of DNA. That's where INO80 chromatin remodeling might comes in. Its role, discovered by Shen and his colleagues, is likely to loosen the damaged DNA from the grip of histone proteins so the DNA repair machinery can access the damaged section. When INO80 is not working properly, damaged DNA can go unrepaired. Such damage can lead to unstable cells and eventually to cancer.
"We knew that at least one gene involved in the INO80 complex had been linked to cancer," said Shen. "This research helps provide a potential mechanism to account for those cancers."
The researchers, led by post-doctoral scientist Ashby Morrison, Ph.D., studied how yeast cells repair double-stranded DNA breaks.
"Double strand breaks are the most serious type of DNA damage," said Shen. "The two DNA strands are completely severed. It is a disaster for a cell. If it is not repaired, the chromosomes become unstable and can fuse to other chromosomes. Many types of cancer result from chromosome fusions."
The scientists created an experimental double strand break in the yeast DNA and monitored specially tagged INO80 molecules inside the cells. They found that INO80 proteins recognize a specific form of histone protein called gamma-H2AX that acts as a "flag" or "code" to direct DNA repair proteins to DNA breaks. Once attached to the histone protein, the INO80 proteins most likely loosen the histone grip on DNA so the repair machinery can gain access and repair the broken ends, the scientists report.
In particular, the scientists discovered one member of the INO80 complex, called Nhp10, is crucial to recognizing the histone code for damaged DNA.
Shen first discovered the INO80 complex in 2000 while studying yeast. Since then, he has revealed that this large protein complex plays an important role in making DNA available for copying into RNA. This latest discovery expands the importance of the INO80 complex, showing it is also crucial to helping repair broken DNA. The scientists discovered that if certain members of the INO80 complex are missing, the yeast becomes prone to the kind of serious damage to its genetic material that can lead to cancer in people.
"The INO80 complex is found in organisms from yeast to humans," said Shen. "Typically these kinds of universal proteins play important basic biological functions, and that is turning out to be the case here."
The scientists are now working out the precise role that INO80 plays in DNA repair and what the protein complex does to the chromosome structure at the double strand break.
"We have introduced a whole new player that has never been seen before in double-strand break repair," said Shen. "This is only the beginning."
In addition to Shen and Morrison, technician Jessica Highland from M. D. Anderson; Nevan Krogan and Jack Greenblatt, Ph.D., University of Toronto; and Ayelet Arbel-Eden and James Haber, Ph.D., Brandeis University, contributed to the research. The research was funded by grants from the National Institutes of Health and M. D. Anderson Cancer Center.