Mutation in the MAF1 gene was identified in a screen for decreased efficiency of tRNA suppressor SUP11 in the yeast Saccharomyces cerevisiae (Sc). maf1-1 mutation exerts a dual phenotypic effect: antisuppression and temperature sensitive (ts) respiratory growth. MAF1, cloned by complementation of the ts phenotype of maf1-1, also alleviates the antisuppressor effect. The coding sequence of MAF1 is interrupted by an intron of 80 bp. The putative gene product, Maf1p, is a hydrophilic protein of 395 amino acids (aa) not showing significant similarity to known proteins which indicates that MAF1 encodes a novel protein. Maf1p may play a role in the tRNA biosynthetic pathway since a fragment of the RPO31/RPC160 gene encoding the largest subunit of RNA polymerase III was cloned as a multicopy suppressor of mafl-1.