UCLA biochemists have determined the three-dimensional structure of a major domain of telomerase, the enzyme that helps maintain telomeres - small pieces of DNA on the ends of chromosomes that act as protective caps - allowing DNA ends to be copied completely when cells are replicated.
This is the first major piece of telomerase for which the structure is known. Telomerase plays a key role in most cancers, and this work ultimately may lead to targets for drug intervention, the scientists said. The discovery is the cover story in the March 4 issue of the journal Molecular Cell.
"Knowledge of the structure should provide insights into how telomerase works," said Juli Feigon, professor of chemistry and biochemistry at UCLA, who led the research group. "Knowing the structure also will allow the pursuit of rational, structure-based drug design, and is a critical first step. The structure provides a potential target for drug intervention."
Feigon emphasized that her laboratory conducts basic research, and is not involved in cancer treatment.
Every time a cell divides, telomeres, which act like the plastic tips on the ends of shoelaces, get shorter. In the natural aging process, the telomeres eventually get so short that cells can no longer divide, and they die. While telomerase is turned off in most types of healthy cells in our bodies, it is active in the vast majority of cancer cells, Feigon said.
Because cancer cells divide rapidly, their telomeres should get shorter more quickly than normal cells. However, because cancer cells have high levels of telomerase activity, which rebuilds the telomeres, cancer cells can maintain the length of their telomeres indefinitely. Although it is not known whether telomerase activation is just a marker for cancer cells or involved in causing it, telomerase is an attractive target for development of anti-cancer drugs by pharmaceutical companies.
The research, which was federally funded by the National Science Foundation and the National Institutes of Health, could have applications for many kinds of cancers.
The domain of telomerase whose structure the biochemists have determined is essential for telomerase to add nucleotides to telomeres. Telomerase is composed of both RNA and proteins. The entire RNA domain is composed of 451 nucleotides, represented by the letters A, C, G and U. Feigon and co-authors UCLA postdoctoral scholar Carla Theimer and graduate student Craig Blois solved the structure of an essential piece of this RNA.
Telomerase has been extremely difficult to characterize structurally because of its size and complexity, and its low level in normal cells.
"This is a unique RNA structure, with distinctive features of RNA folding," said Feigon, who determined the structure with Theimer and Blois using nuclear magnetic resonance (NMR) spectroscopy.
Mutations in the RNA are associated with the inherited diseases aplastic anemia and dyskeratosis congenita, which frequently are manifested by progressive bone-marrow failure.
"When you look at the sequence on paper, it doesn't look like some of these mutations would have much effect on the overall three-dimensional structure," Feigon said. "However, it turns out, for instance, that changing a single 'C' nucleotide to a 'U' nucleotide has a dramatic effect on the stability of the three-dimensional fold of the RNA, which is essential for the function of the enzyme, and causes aplastic anemia in patients who have this mutation.
"There are five known disease mutations in this part of the RNA identified so far. For three of them, it was not clear why they would be a problem for telomerase, but by solving the structure, we now understand how they disrupt the folding and stability of the RNA and why they are disease mutations."
For telomerase to be active, it needs the telomerase RNA and a protein called human telomerase reverse transcriptase, which is related to the reverse transcriptase protein that is important for replicating the AIDS virus. Feigon's laboratory has been working on the RNA.
"It is a dream of mine to figure out what this RNA is doing with the protein," Feigon said. "Reverse transcriptases normally copy RNA to DNA, but do not contain RNA; in this enzyme, the protein requires the RNA component to function. The enzyme is unique because it has its own internal piece of RNA that is used to copy the DNA, but this 'template' is only approximately 10 of the 451 nucleotides. Nobody knows what the rest of the RNA is really doing as part of this enzyme; that is what we're trying to understand. We're getting closer to answering this question."
The structure reveals a "pseudoknot" that is required for telomerase activity, at whose core three strands of RNA come together to form a triple helix. All vertebrate animals investigated so far have nearly the identical sequence of nucleotides through the triple helix, Theimer said. "The triple helical structure of the pseudoknot must be significant since it is conserved from humans to marsupials, birds and marine animals," Theimer said.
Feigon's laboratory studies the three-dimensional structures of DNA and RNA, and how proteins and DNA and RNA recognize one another to switch genes on and off in cells. Her laboratory has worked on telomeres and telomerase for more than a decade.
A member of UCLA's faculty since 1985, Feigon was the first UCLA scientist to use NMR to determine DNA and RNA structures.