UCSF Study Reveals Mechanism Of Telomerase Enzyme That Could Lead To Target For Cancer Therapy, Cell Regeneration

The enzyme -- brought to popular fame two years ago by studies showing that itcould be manipulated in cell culture to increase the life span of cells - hasthe capacity to replenish the tips of chromosomes, known as telomeres, whichlose their final fragments with each cycle of cell division. When activated,telomerase replenishes telomeres by copying the RNA folded within it intotelomeric DNA and assembling it on the ends of the chromosomes.

Telomeres maintain the stability of chromosomes in numerous ways and, much likechemical bookends, prevent them from unwinding. But in a fine illustration ofnature's ingenuity, they also play a role in regulating the life span of acell, because the tips of telomeres drop off chromosomes with each cycle ofcell division.1 As the telomeres gradually shorten, the probability that theywill signal the cell to stop dividing gradually increases. As a result, aftermany cycles of cell division have occurred, all cells in a population havedied.

While the telomerase enzyme has the ability to replenish the telomeres on theends of chromosomes, it is inactive in many adult human tissues. However, it isactive when massive amounts of cell division are underway - as in self-renewingadult cells of the immune system, during the development of an embryo, and incancer.

In the UCSF study, reported in the May 5 issue of Science, the researchersdetermined that a small structure within the RNA molecule of yeast telomerasecontrols the precision with which the enzyme carries out its key function --spinning out the repeated sequences of telomeric DNA that bind the ends ofchromosomes.

When they disrupted this small region of RNA, the enzyme began to spin outtelomeres uncontrollably, until they stalled out like a car run into a ditch.This result occurred in culture and in the yeast cells themselves. Cell deathsoon ensued.

As the human version of telomerase appears to have a structural region that issimilar to that examined in the yeast enzyme, the region could prove to be atarget for killing cancer cells, says the senior author of the study, ElizabethBlackburn, PhD, UCSF professor of microbiology and immunology and biochemistryand biophysics, who co-discovered telomerase in 1985. (The function of thehuman version of the region has not been determined.) Moreover, she says, itcould prove a target for regenerating cells that have been damaged throughinjury or wear and tear.

Telomerase and other so-called reverse transcriptases, including HIV, bothcontain RNA enfolded in protein. As such, they are known as ribonucleoproteins.Most enzymes, by contrast, contain only protein.

Much of the excitement regarding the current finding stems from the fact that,to date, research on telomerase and HIV has focused only on the proteincomponents, as they make up a central part of the enzymes' active sites.

"This discovery represents the first time anybody has shown a mechanistic rolefor a structure of RNA in the action of telomerase, and we think this isprobably a universal kind of feature of telomerase," says Blackburn.

The finding also adds weight to recent evidence that the RNA component of HIV,traditionally disregarded in retroviral therapies as passive, plays an integralrole in HIV replication, and it therefore should be closely re-examined as apossible target for therapy, says Blackburn.

"This discovery should turn researchers' spotlights back to the RNA componentsof both telomerase and HIV," she says.

The discovery may also offer a glimpse into the evolutionary past. Someresearchers have believed that during an earlier period in evolutionary historythe RNA molecule in telomerase might have deferred functional power to theprotein component.

The discovery that the seemingly archaic RNA retains a key mechanistic role intelomerase function builds support for the theory that telomerase -- andreverse transcriptases such as HIV -- represent an intermediate step in theevolution of enzymes from strictly RNA sequences to strictly protein sequences,says Blackburn.

"Such a direct function for the RNA structure in the enzymatic action oftelomerase supports an evolutionary scheme in which RNA enzymes acquiredprotein components evolving into ribonucleoprotein enzymes," she says. "The RNAcomponents then gradually lost their functional roles in catalysis and weresubsequently dispensable."

Telomerase and the other reverse transcriptase enzymes function by synthesizingcopies of DNA from the RNA folded within their protein. Nearly all organismshave an enzyme that can stimulate the conversion of DNA, which contains anorganism's genetic code, or genes, into messenger RNA, the first step on theroad to developing protein. Only the reverse transcriptases do the opposite.

But telomerase and HIV diverge in a key aspect of their transcriptionmechanism. While HIV draws thousands of bases of RNA through its catalyticsite, spinning out long sequences of viral DNA, telomerase draws in only a verysmall portion of its long RNA sequence to the catalytic site, and copies thisone segment, known as the template, over and over into telomeric DNA, which itthen assembles and adds to the ends of chromosomes.

Until now, researchers have not understood what specifies the enzyme's templateboundaries, preventing unbridled spinning of telomeric DNA onto the ends ofchromosomes. In the current study, the researchers determined that thelimitation on DNA synthesis is controlled by a small segment of RNA nestled upadjacent to the downstream end of the RNA template. The region, made up of basepairs of RNA that are "zipped up" together within a larger segment of RNA, actsas a boundary for the replication process.

When the researchers altered this RNA region, the buffer "unzipped," providinga long strip of RNA - up one side of the zipper -- for the enzyme to continueconverting into DNA. With free reign, the enzyme drew more and more of theribonucleotides into its active site, until it synthesized so much telomericDNA that eventually, says Blackburn, the telomerase RNA may have bunched up,halting telomere synthesis and causing cell death.

Such abnormal, almost ceaseless, replication, says Blackburn, is reminiscent ofthe behavior of normal reverse transcriptases such as those found inretroviruses like HIV.

"Our finding in telomerase gives strength to the importance of looking at theRNA component of HIV," says Blackburn, "because by making just a simple changein telomerase RNA we can make it act more like HIV, and this suggests that HIVis actually like telomerase -- when it acts in cells to make new viruses, itreally is acting together with its RNAs. I think this is something one shouldbe thinking about for drug targets."

The finding adds weight to a recent UCSF study led by Tristram Parslow, MD,PhD, professor of pathology, and Thomas James, PhD, professor and chair ofpharmaceutical chemistry, who reported (Nature Structural Biology, vol. 5, p.432, 1998) that a different "zipped up" region of RNA base pairs is essentialfor replication of HIV. When the base pairs were mutated, HIV lost its abilityto infect a cell.

Co-authors of the study were Yehuda Tzfati, PhD, postdoctoral researcher, TracyB. Fulton, graduate student and Jagoree Roy, PhD, postdoctoral researcher, allof the UCSF Department of Microbiology and Immunology.

The study was funded by the National Institutes of Health.