Genetic interaction between yeast Saccharomyces cerevisiae release factorsand the decoding region of 18S rRNA

Functional and structural similarities between tRNA and eukaryotic class 1 release factors (eRF1) described previously, provide evidence for the molec ular mimicry concept. This concept is supported here by the demonstration o f a genetic interaction between eRF1 and the decoding region of the ribosom al RNA, the site of tRNA-mRNA interaction. We show that the conditional let hality caused by a mutation in domain 1 of yeast eRF1 (P86A), that mimics t he tRNA anticodon stem-loop, is rescued by compensatory mutations A1491G (r dn15) and U1495C (hyg1) in helix 44 of the decoding region and by U912C (rd n4) and G886A (rdn8) mutations in helix 27 of the 18 S rRNA. The udn15 muta tion creates a C1409-G1491 base-pair in yeast rRNA that is analogous to tha t in prokaryotic rRNA known to be important for high-affinity paromomycin b inding to the ribosome. indeed, rdn15 makes yeast cells extremely sensitive to paromomycin, indicating that the natural high resistance of the yeast r ibosome to paromomycin is, in large part, due to the absence of the 1409-14 91 base-pair. The rdn15 and hyg1 mutations also partially compensate for in activation of the eukaryotic release factor 3 (eRF3) resulting from the for mation of the [PSI+] prion, a self-reproducible termination-deficient confo rmation of eRF3. However, rdn15, but not hyg1, rescues the conditional cell lethality caused by a GTPase domain mutation (R419G) in eRF3. Other antisu ppressor rRNA mutations, rdn2 (G517A), rdn1T(C1054T) and rdn12A(C526A), str ongly inhibit [PSI (+)]-mediated stop codon read-through but do not cure ce lls of the [PSI (+)] prion. Interestingly, cells bearing hyg1 seem to enabl e [PSI (+)] strains to accumulate larger Sup35p aggregates upon Sup35p over production, suggesting a lower toxicity of overproduced Sup35p when the ter mination defect, caused by [PSI (+)], is partly relieved. (C) 2003 Academic Press.