Eukaryotic Cell doi:10.1128/EC.00192-07
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.
Tudor nuclease genes and programmed DNA rearrangements in Tetrahymena thermophila
Rachel A. Howard-Till
and
Meng-Chao Yao*
Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98109; Molecular and Cellular Biology Program, University of Washington, Seattle, WA 98195; and Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan 11529
* To whom correspondence should be addressed. Email:
mcyao{at}fhcrc.org.
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Abstract |
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Proteins containing a Tudor domain and domains homologous to staphylococcal nucleases are found in a number of eukaryotes. These "Tudor nucleases" have been found associated with the RNA induced silencing complex (6). We have identified two Tudor nuclease homologs, TTN1 and TTN2, in the ciliate Tetrahymena thermophila, which has two distinct small RNA pathways. Characterization of single and double knockouts of TTN1 and TTN2 shows that neither of these genes is essential for growth or sexual reproduction. Progeny of TTN2 knockouts and double knockouts occasionally show minor defects in the small RNA guided process of DNA deletion, but appear normal in hairpin RNA induced gene silencing, suggesting that Tudor nucleases play only a minor role in RNAi in Tetrahymena.
Previous studies of Tetrahymena have shown that inserted copies of the neo gene from E. coli are often deleted from the developing macronucleus during sexual reproduction (22, 38). This transgene deletion phenomenon is hypothesized to be a form of genome defense. Analysis of the Tudor nuclease mutants revealed exceptionally high rates of deletion of the neo transgene at the TTN2 locus but no deletion at the TTN1 locus. When present in the same genome, however, the neo gene becomes deleted at high rates even at the TTN1 locus, further supporting a role for trans-acting RNA in this process. This deletion is not affected by the presence of the same sequence in the macronucleus, thus providing a counter argument for the role of the macronuclear genome in specifying all sequences for deletion.
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