GEL SHIFT AND UV CROSS-LINKING ANALYSIS OF TETRAHYMENA TELOMERASE

Telomerase is an unusual ribonucleoprotein that synthesizes new telome res onto chromosome ends. The enzyme has been most extensively charact erized in ciliates, where the RNA component has been cloned from sever al species, and its elongation properties have been characterized in d etail. To understand the substrate specificity and protein composition of telomerase, we have used gel shift and UV cross-linking to charact erize the enzyme from the ciliate Tetrahymena thermophila. In a mobili ty shift assay, a complex was identified that contained telomerase RNA ,co-purified with telomerase activity, and was sensitive to nuclease t reatment. The mobility shift complexes specifically formed using sever al different single-stranded, telomeric sequences but not non-telomeri c primers. These results suggest that the specificity of telomerase fo r G-rich primer sequences occurs at least in part at the level of prim er binding. UV cross-linking analysis identified a 100-kDa cross-linke d protein that may be a telomerase component.