Telomerase is a telomere specific DNA polymerase required for the replication of chromosome ends. Conventional DNA polymerases do not complete the replication of the very end of the lagging strand of a linear DNA molecule. Telomerase compensates for this incomplete replication by adding telomere specific repeats onto chromosome ends de novo. Telomerase contains an essential RNA component which specifies the sequence added. Telomerase was first identified in the immortal single cellular protozoan Tetrehymena. Recent experiments suggest that human telomerase regulation may play a role in cancer progression. Telomerase is not active in most human somatic tissue, however in immortalized cells in culture and in cancer tissue telomerase is active. The reactivation of telomerase in tissue culture cells after immortalization suggests that telomerase may be required for the growth of immortalized cells. Thus telomerase has been proposed as a target for anti-cancer therapies. To determine the role of telomerase in cancer, and to help design potential inhibitors it is essential to have a thorough understanding of telomerase biochemistry. Tetrahymena remains the organism of choice from which to study the biochemical mechanism of telomerase. Large quantities of cells can be grown easily and there is 100 to 1000 times more activity than from an equivalent mass of human cells. The goal of my laboratory is to understand the mechanism and regulation of telomerase. The specific aim of this proposal is to identify and functionally analyze all of the components of the Tetrahymena telomerase. Telomerase proteins have not been cloned from any organism. We propose to clone and characterize the protein components of the enzyme, dissect the structural requirements for telomerase RNA, use gene replacement to study the requirement for telomerase in vivo and probe the biochemical reaction mechanism of the purified enzyme. The experiments proposed here will in effect work double time; they will establish the mechanism of a new class of DNA polymerase and also may shape the development of new cancer therapies.