Termination by RNA polymerase III (Pol III) produces RNAs whose 3’ oligo(U) termini are bound by La protein, a chaperone that protects RNAs from 3’ exonucleases and promotes their maturation. Multiple reports indicate that yeasts use La-dependent and-independent pathways for tRNA maturation, with defective pre-tRNAs being most sensitive to decay and most dependent on La for maturation and function. The Rpc11p subunit of Pol III shows homology with the zinc ribbon of TFIIS and is known to mediate RNA 3’ cleavage and to be important for termination. We used a La-dependent opal suppressor, tRNASerUGAM, which suppresses ade6-704 and the accumulation of red pigment, to screen Schizosaccaromyces pombe for rpc11 mutants that increase tRNA-mediated suppression. Analyses of two zinc ribbon mutants indicate that they are deficient in Pol III RNA 3’ cleavage activity and produce pre-tRNASerUGAM transcripts with elongated 3_-oligo (U) tracts that are better substrates for La. A substantial fraction of pre-tRNASerUGAM contains too few 3’ Us for efficient La binding and appears to decay in wild-type cells but has elongated oligo (U) tracts and matures along the La-dependent pathway in the mutants. The data indicate that Rpc11p limits RNA 3’-U length and that this significantly restricts pre-tRNAs to a La-independent pathway of maturation in fission yeast.

In addition to directing transcription initiation, core promoters integrate input from distal regulatory elements. Except for rare exceptions, it has been generally found that eukaryotic tRNA and rRNA genes do not contain TATA promoter elements and instead use protein-protein interactions to bring the TATA-binding protein (TBP), to the core promoter. Genomewide analysis revealed TATA elements in the core promoters of tRNA and 5S rRNA (Pol III), U1 to U5 snRNA (Pol II), and 37S rRNA (Pol I) genes in Schizosaccharomyces pombe. Using tRNA-dependent suppression and other in vivo assays, as well as in vitro transcription, we demonstrated an obligatory requirement for upstream TATA elements for tRNA and 5S rRNA expression in S. pombe. The Pol III initiation factor Brf is found in complexes with TFIIIC and Pol III in S. pombe, while TBP is not, consistent with independent recruitment of TBP by TATA. Template commitment assays are consistent with this and confirm that the mechanisms of transcription complex assembly and initiation by Pol III in S. pombe differ substantially from those in other model organisms. The results were extended to large-rRNA synthesis, as mutation of the TATA element in the Pol I promoter also abolishes rRNA expression in fission yeast. A survey of other organisms’ genomes reveals that a substantial number of eukaryotes may use widespread TATAs for transcription. These results indicate the presence of TATA-unified transcription systems in contemporary eukaryotes and provide insight into the residual need for TBP by all three Pols in other eukaryotes despite a lack of TATA elements in their promoters.

Transcriptional silencing is a heritable form of gene inactivation that involves the assembly of large regions of DNA into a specialized chromatin structure that inhibits transcription. This phenomenon is responsible for inhibiting transcription at silent mating-type loci, telomeres and rDNA repeats in both budding yeast Saccharomyces cerevisiae and fission yeast Schizosaccharomyces pombe, as well as at centromeres in fission yeast. Although transcriptional silencing in both S.cerevisiae and S. pombe involves modification of chromatin, no apparent amino acid sequence similarities have been reported between the proteins involved in establishment and maintenance of silent chromatin in these two distantly related yeasts. Silencing in S. cerevisiae is mediated by Sir2p-containing complexes, whereas silencing in S. pombe is mediated primarily by Swi6-containing complexes. The Swi6 complexes of S.pombe contain proteins closely related to their counterparts in higher eukaryotes, but have no apparent orthologs in S. cerevisiae. Silencing proteins from both yeasts are also actively involved in other chromosome-related nuclear functions, including DNA repair and the regulation of chromatin