ABSTRACT
The cleavage/polyadenylation factor (CPF) of Saccharomyces cerevisiae is thought to provide the catalytic activities of the mRNA 3'-end processing machinery, which include endonucleolytic cleavage at the poly(A) site, followed by synthesis of an adenosine polymer onto the new 3'-end by the CPF subunit Pap1. Because of similarity to other nucleases in the metallo-beta-lactamase family, the Brr5/Ysh1 subunit has been proposed to be the endonuclease. The C-terminal domain of Brr5 lies outside of beta-lactamase homology, and its function has not been elucidated. We show here that this region of Brr5 is necessary for cell viability and mRNA 3'-end processing. It is highly homologous to another CPF subunit, Syc1. Syc1 is not essential, but its removal improves the growth of other processing mutants at restrictive temperatures and restores in vitro processing activity to cleavage/ polyadenylation-defective brr5-1 extract. Our findings suggest that Syc1, by mimicking the essential Brr5 C-terminus, serves as a negative regulator of mRNA 3'-end formation.
Subject(s)
RNA, Fungal/metabolism , RNA, Messenger/metabolism , Saccharomyces cerevisiae Proteins/chemistry , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae/metabolism , mRNA Cleavage and Polyadenylation Factors/chemistry , mRNA Cleavage and Polyadenylation Factors/metabolism , Amino Acid Sequence , Gene Deletion , Genes, Fungal , Molecular Sequence Data , Mutagenesis , Pancreatitis-Associated Proteins , Protein Structure, Tertiary , RNA Processing, Post-Transcriptional , RNA, Fungal/genetics , RNA, Messenger/genetics , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae Proteins/genetics , mRNA Cleavage and Polyadenylation Factors/geneticsABSTRACT
Genomic libraries were constructed from three Drosophila species, namely Drosophila auraria, Drosophila serrata, and Drosophila kikkawai, belonging to the Drosophila montium subgroup of the Drosophila melanogaster species group. Clones containing beta-tubulin specific sequences were isolated, characterized by restriction endonuclease digestions and Southern hybridizations, and mapped by in situ hybridization on the polytene chromosomes of the species studied. The distribution of the beta-tubulin loci was found to be similar in D. montium species and D. melanogaster.
