Telomere
Telomere
Telomeres are structures found at the ends of chromosomes in the cells of eukaryotes . Telomeres function by protecting chromosome ends from recombination, fusion to other chromosomes, or degradation by nucleases . They permit cells to distinguish between random DNA breaks and chromosome ends. They also play a significant role in determining the number of times that a normal cell can divide. Unicellular forms whose cells have no true nuclei (prokaryotes) possess circular chromosomes that, therefore, have no ends. Thus, prokaryotes can have no telomeres.
Structure
Telomeres are extensions of the linear, double-stranded DNA molecules of which chromosomes are composed, and are found at each end of both of the chromosomal strands. Thus, one chromosome will have four telomeric tips. In humans, the forty-six chromosomes are tipped with ninety-two telomeric ends.
In most eukaryotic forms, telomeres consist of several thousand repeats of the specific nucleotide sequence TTAGGG and occur in organisms ranging from slime molds to humans. The entire length of repeated telomere sequences is known as the terminal restriction fragment (TRF). Sequences different from TTAGGG are found in more primitive eukaryotic forms, such as the ciliated protozoan Tetrahymena, in which Elizabeth Blackburn first characterized the repeated telomere sequence.
The polymerases that copy the chromosomes of DNA strands are unable to copy completely to the end. This became known as the "end-replication problem" when it was first recognized in the late 1960s. The TRF acts like a buffer that protects the information-containing genes, so that the loss of some telomeric nucleotide sequences at each round of DNA replication does not result in the loss of genetic information. The telomeres themselves end in large duplex loops, called T-loops.
A Simple Counting Mechanism
For the first half of the twentieth century it was believed that cells cultured in laboratory glassware could replicate indefinitely if the correct nutrient media and other conditions of growth could be found. Repeated initial failure at culturing indefinitely replicating cells was followed by success in the late 1940s, when the immortal L929 cancer cell population was developed from mouse tissue. Later, other immortal cell populations were found, including the first human cell line, HeLa, derived from a human cervical carcinoma.
It was originally, but erroneously, believed that normal cells also had the potential to divide and function indefinitely in culture, and so it was thought that aging could not be the result of events that occurred within normal cells. Instead, aging was thought to be the result of extracellular events such as radiation or of changes in the extracellular molecules that cement cells to each other.
In 1960, however, it was discovered that no culture conditions exist that will permit normal human cells to divide indefinitely. Rather, cells were found to have a built-in counting mechanism, called the Hayflick Limit, that limits their capacity to replicate. For example, human fibroblast cell populations, found in virtually all tissues, will double only about 50 times in culture when derived from fetal tissue. Fibroblast populations from older adults double fewer times, the exact number of doublings depending upon the age of the donor. Leonard Hayflick and P. S. Moorhead also suggested that only abnormal or cancer cells divide indefinitely. They theorized that the limited capacity for normal cells to divide is an expression of aging and that it determines the longevity of the organism.
In support of this theory, it was found that frozen normal fetal cells "remember" the doubling level at which they were frozen and, after thawing, will undergo additional doublings until the total of fifty is reached. These facts suggested to Hayflick that a replication-counting mechanism existed. Hayflick and coresearcher Woodring Wright later found that this mechanism was located in the nucleus of the cell.
The Discovery of Telomeres
The search for the molecular counting mechanism ended when Calvin Harley and Carol Greider discovered that the telomeres of cultured normal human fibroblasts become shorter each time the cells divide. When telomeres reach a specific short length, they signal the cell to stop dividing. Therefore, cellular aging, as marked by telomere shortening, is not based on the passage of time. Instead, telomere loss measures rounds of DNA replication. For this reason, Hayflick has coined the term "replicometer" for this mechanism.
An accumulation of evidence suggests that while telomere attrition explains the loss of replicative capacity in normal cells, the process may not be as simple as first believed. There are several essential DNA-binding proteins (for example, TRF1 and TRF2) associated with telomeres, and the role that they play in capping and uncapping the telomere ends undoubtedly will be found to complicate the oversimplified explanation given above.
Telomerase
Immortal cancer cells escape telomere loss by switching on a gene that expresses an enzyme called telomerase. This unusual enzyme is a reverse transcriptase that has an RNA template and a catalytic portion. At each round of DNA replication, telomerase adds onto the existing telomeres the nucleotides that would otherwise have been lost, thus maintaining a constant telomere length. In other words, telomerase acts as an "immortalizing" enzyme. In addition, it has several associated proteins whose roles are still under investigation.
Using what is called the TRAP assay (telomeric repeat amplification protocol), it has been found that about 90 percent of all human tumors produce telomerase, whereas the only normal adult somatic cells that produce telomerase are stem cell populations found, for example, in skin, the hematopoietic system, germ cells , and gut epithelia. In fact, the presence or absence of telomerase is the most specific property that distinguishes cancer cells from normal cells. This difference is currently under investigation as a diagnostic tool. If a chemical could be found to interfere with telomerase activity in cancer cells, an effective control of this disease might be found. Several candidate substances have been identified and are undergoing extensive studies in animals.
Telomerase is switched on in virtually all human cells at the moment of conception, but as the embryo matures the telomerase becomes repressed in all but the germ cells and stem cell populations. Further, the level of telomerase expressed in stem cells is much less than that expressed in cancer cells. Interestingly, telomerase expression has been found to occur in all the cells of animals that age slowly or not at all. These are animals, such as the American lobster and the rainbow trout, that do not stabilize at a fixed size in adulthood.
On the human genome, an enzyme known as human telomerase reverse transcriptase (hTERT) is found on the most distal gene on chromosome 5p. The transfection (introduction) of hTERT into cultured normal human fibroblasts has resulted in telomere elongation, telomerase expression, and the immortalization of these otherwise mortal cells. After several hundred population doublings, the transfected cells exhibit some drift from the diploid number of chromosomes but cancer cell properties do not occur. This experiment proves that telomerase is not a cancer enzyme but an immortalization enzyme. The ability to immortalize normal human cells via hTERT has important potential applications. Some immortalized cells could be cultured in the lab to produce therapeutically useful molecules. Others might be used directly within the body to repair tissue or replace lost or damaged cells.
see also Chromosome, Eukaryotic; DNA Polymerases; Replication; Reverse Transcriptase.
Leonard Hayflick
Bibliography
Bodnar, A. G., et al. "Extension of Life Span by Introduction of Telomerase intoNormal Human Cells." Science 279 (1998): 349-352.
Greider, Carol W. "Telomeres and Senescence: The History, the Experiment, the Future." Current Biology 8 (1998): 178-181.
Hayflick, Leonard. How and Why We Age. New York: Ballantine Books, 1996.
———. "The Illusion of Cell Immortality." British Journal of Cancer 83 (2000):841-846.