Project Summary/Abstract Telomeres of most species are simple repetitive sequences that are maintained by de novo telomere repeat addition by the ribonucleoprotein telomerase. Small RNAs composed of perfect telomere repeats have been observed in several organisms, and these have been linked to silencing marks at telomeres and to DNA damage at telomeres. We discovered an endogenous C. elegans small RNA pathway that promotes telomere stability in the absence of telomerase. C. elegans small RNAs that are perfectly complementary to telomeric DNA are very rare, measuring 1 read per 10 million small RNAs. However, two related Caenorhabditis species C. brennerei and C. remanei have abundant telomeric small RNAs whose levels were increased by several orders of magnitude. Perfect telomeric small RNAs in all three Caenorhabditis species were strongly depleted for 5' guanine nucleotides that characterize major C. elegans small RNA species like 26G and 22G RNAs. These results suggest that telomeric small RNAs have a distinct biogenesis mechanism and that a dramatic change in their role at telomeres has recently occurred in Caenorhabditis species. We propose to study telomere biology in C. brennerei and C. remanei, in an effort to understand why telomeric small RNAs are so abundant in these species, how telomeric small RNAs are created, and what their functions are. Potentially analogous telomeric small RNAs have been observed in ciliates and mammals, and we may elucidate their biological relevance.