The nuclear RNA and DNA helicase Sen1 is essential in the yeast Saccharomyces cerevisiae and is required for efficient termination of RNA polymerase II transcription of many short non-coding RNA genes. However, the mechanism of Sen1 function is not understood. We created a plasmid-based genetic system to study yeast Sen1 in vivo. Using this system, we show that: 1) the minimal essential region of Sen1 corresponds to the helicase domain and one of two flanking nuclear localization sequences, 2) a previously isolated terminator read-through mutation in the Sen1 helicase domain, E1597K, is rescued by a second mutation designed to restore a salt bridge within the first RecA domain, and 3) the human ortholog of yeast Sen1, Senataxin, cannot functionally replace Sen1 in yeast. Guided by sequence homology between the conserved helicase domains of Sen1 and Senataxin, we tested the effects of 13 missense mutations that co-segregate with the inherited disorder ataxia with oculomotor apraxia type 2 on Sen1 function. Ten of the disease mutations resulted in transcription read-through of at least one of three Sen1-dependent termination elements tested. Our genetic system will facilitate the further investigation of structure-function relationships in yeast Sen1 and its orthologs.
- Received June 20, 2014.
- Accepted August 11, 2014.
- Copyright © 2014, The Genetics Society of America