S.pombe telomerase RNA identified

Blogging on Peer-Reviewed Research Webb, C.J., Zakian, V.A. (2008). Identification and characterization of the Schizosaccharomyces pombe TER1 telomerase RNA. Nature Structural & Molecular Biology, 15(1), 34-42. DOI: 10.1038/nsmb1354

Leonardi, J., Box, J.A., Bunch, J.T., Baumann, P. (2008). TER1, the RNA subunit of fission yeast telomerase. Nature Structural & Molecular Biology, 15(1), 26-33. DOI: 10.1038/nsmb1343

Two papers in Nature Structural & Molecular Biology identify the telomerase RNA in Schizosaccharomyces pombe. Telomerase is a multi-unit enzyme that has both protein and RNA components. While the protein subunit is highly conserved and identifiable through sequence comparisons of eukaryotes, the RNA subunit has a variable size and sequence making identification through comparative means more difficult. The S. pombe telomerase RNA subunit, or TER1, was discovered by two labs applying similar biochemical approaches to identify the locus.

The Webb et al paper used immunoprecipitation (IP) to enrich for molecules that interacted with the Trt1p catalytic subunit. This was followed by additional IP with antibodies that for the 2,2,7-trimethylguanosine (TMG) cap structure which is conserved in ncRNAs. The resulting enriched solution was subject to RT-PCR to then sequence and identify the transcripts. With some analyses they were able to determine some novel transcripts had been isolated and hypothesized these were the elusive RNA subunit. Genetic work confirmed this through disrupting the locus and observing teleomere shrinking and senescence in the modified strains.

The Leonardi et al paper described how even starting with bioinformatic analyses to identify potential candidate regions, more than 100 loci were predicted prompting a biochemical strategy to identify regions. myc-tagged Trt1 was IP-ed down with appropriate controls to insure that it was still an active telomerase, and then nucleic acid recovered and RT-PCRed to identify sequence associated with the complex. Genetic mutants of the locus were created and confirmation that the TER1 locus was required for telomerase activity.

Because of the similar size of several snRNAs have similar lengths in human and S. pombe but the S. cerevisiae is much shorter the authors hypothesize that the TER1 would be similarly length to human TERC (~450 nt). Instead it was found to be 1,200+ nt long similar to size of TLC1 in budding yeast species. Additional effort to characterize the TLC1 homologs in Saccharomyes spp showed that it was rapidly evolving suggesting. Similar analyses of TER1 in Schizosaccharomyces spp (like these I would guess) showed the gene to be rapidly evolving between moderately diverged species. This suggests that it will be difficult to identify the gene by homology alone, although perhaps sophisticated secondary-structure searching tools like XRate and Infernal can identify them through comparative analyses.

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