The structure of the telomere is fundamental to chromosomal integrity and, by extension, to the state of the cell. Chromosome ends are maintained by the ribonucleoprotein telomerase in a regulated manner. A complex of proteins at the telomere, anchored by TRF1, TRF2, and POT1, has been described for mammalian cells, and this complex regulates access of telomerase to the telomere and distinguishes the telomere from a broken chromosome end. The importance of this complex is indicated by the lethal nature of mouse knockouts for at least two components. Although humans stringently repress telomerase in most somatic tissues, the frog Xenopus does not, and maintains telomere length in all tissues. Xenopus TRF1, TRF2, and POT1 were identified and tested for telomere-binding function. This research seeks to identify the interaction partners of these Xenopus telomere binding proteins by database mining and molecular cloning procedures, and the new proteins will then be tested for functional interaction by co-immunoprecipitation from cells in which both partners are expressed. Lastly, the telomere complex will be perturbed in vivo in developing embryos by RNA interference-based knockdowns, or dominant-negative versions of proteins that interfere with function, and the embryos examined for effects on telomere length and for phenotypic abnormalities expected upon disruption of the cell cycle.
Chromosome ends, or telomeres, are critical to the cell, signalling whether the cell can continue to divide or should die. Though much is known about telomere function in cultured mammalian cells, this project will investigate the role of the telomere state in a whole organism, and one very different from mammals in telomere metabolism. Furthermore, as this project will be carried out at Reed College, a small liberal arts institution, it will provide the framework and resources for undergraduate students to pursue independent research as part of their educational experience, and prepare them more rigorously for their roles as educated citizens and future scientists.
