INTERACTION BETWEEN NMD2P AND UPF1P IS REQUIRED FOR ACTIVITY BUT NOT FOR DOMINANT-NEGATIVE INHIBITION OF THE NONSENSE-MEDIATED MESSENGER-RNA DECAY PATHWAY IN YEAST

Rapid turnover of nonsense-containing mRNAs in the yeast Saccharomyces cerevisiae is dependent on the products of the UPF1 (Upf1p), NMD2/UPF 2 (Nmd2p) and UPF3 (Upf3p) genes. Mutations in each of these genes lea d to the selective stabilization of mRNAs containing early nonsense mu tations without affecting the decay rates of most other mRNAs. NMD2 wa s recently identified in a two-hybrid screen as a gene that encodes a Upf1p-interacting protein. To identify the amino acids essential to th is interaction, we used two-hybrid analysis as well as missense, nonse nse, and deletion mutants of NMD2, and mapped the Upf1p-interacting do main of Nmd2p to a 157-amino acid segment at its C-terminus. Mutations in this domain that disrupt interaction with Upf1p also disrupt nonse nse-mediated mRNA decay. A dominant-negative deletion allele of NMD2 i dentified previously includes the Upf1p-interacting domain. However, m utations in the Upf1p-interacting domain do not affect dominant-negati ve inhibition of mRNA decay caused by this allele, suggesting interact ion with yet another factor. These results, and the observation that d eletion of a putative nuclear localization signal and a putative trans membrane domain also inactivate nonsense-mediated mRNA decay, suggest that Nmd2p may contain as many as four important functional domains.