NIH researchers link enzyme deficiency with faulty DNA replication and cancer

Asterisks highlight solid green PCNA protein that accumulates in cells with a lack of the enzyme ATAD5. The accumulation of protein occurs during stalled DNA replication.

As cells divide, they make a duplicate copy of their genetic blueprint through a process called DNA replication. NIH researchers have discovered a cancer-causing flaw that can occur when ATAD5, one of many enzymes in the replication process, is deficient. The defect causes DNA replication to stall and the normal precision of replication proteins to become chaotic. The study appeared in the Dec. 31, 2012, advance online issue of the Journal of Cell Biology.

The team of researchers from the National Human Genome Research Institute (NHGRI) and the National Cancer Institute (NCI) detected the enzyme malfunction in the biochemical machinery that processes one of the two strands of DNA during replication. The faulty machinery for copying that strand affects the progress of the cell through the DNA replication process.

In previous research, members of the team isolated ATAD5 by screening genes that code for a similar protein in yeast. They subsequently observed that mice deficient in the enzyme die or develop tumors. In the present study, performed with cells and with mice in the lab, they concluded that ATAD5, or its functionally similar enzyme in yeast, is essential for copying the lagging strand of all DNA, whether yeast, mouse or human.

The DNA molecule is a double helix, made of two bonded strands, aligned in opposite directions. One strand is called the leading strand, and the other is called the lagging strand. In DNA replication, various enzymes set to work — one enzyme breaks the bond of the two strands, while another enzyme prevents them from rebinding. Still others build new sequences of DNA that complement the originals. A slightly different copying process is required for the lagging strand, which relies on the enzyme ATAD5.

The ATAD5 enzyme acts on a protein called PCNA that anchors itself to the DNA lagging strand as it forks off from the leading strand during replication. The activity of many enzymes at work at the forked-off juncture is called its replication factory. This biochemical term suggests an orderly workflow, but ATAD5 deficiency stalls that workflow. Study researchers describe the harmful buildup of proteins when ATAD5 is deficient and fails to unseat the PCNA protein from its position along the DNA lagging strand.

"PCNA positions around a strand of DNA like a clamp," Kyungjae Myung, Ph.D., senior NHGRI investigator and senior author explained. "The clamp closes around the strand to guide the replication polymerase that goes forward and comes back." Polymerase is a protein that strings together the DNA bases making up the new strand of DNA.

When ATAD5 is deficient, it does not give a signal for release of the PCNA clamp that encircles the DNA strand as it is fed into the DNA replication factory. This PCNA enzyme is not only important in its role in clamping onto the strand of DNA, but also an orientation point for the many other enzymes that play a role in replication. Instead of doing their work in an orderly way, these enzymes bunch up where the PCNA stalls, bogging down the DNA replication process. They also associated this congestion of proteins with cancer in mice and humans.

"All the enzymes accumulate in the factory if you don't have ATAD5," said Dr. Myung. The researchers observed that cells lacking ATAD5 had slower phases of cell division. They noted that ATAD5-deficient cells had not accomplished a replication phase that cells normally completed within nine hours.

Kyoo-young Lee, Ph.D., a post-doctoral researcher in Dr. Myung's lab and lead author of the study, presented the study at the December 2012 Bioscience and Engineering Symposium of the NIH Korean Scientists Association. His presentation garnered the 2012 Johng S. Rhim Young Investigator Award.

For an animated depiction of the intricate process of DNA replication, play the movie clip at this site:
www.hhmi.org/biointeractive/dna/DNAi_replication_vo2.html
[hhmi.org]

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Posted: January 4, 2013