Synthetic lethal interactions with conditional poly(A) polymerase alleles identify LCP5, a gene involved in 18S rRNA maturation

• Published in: RNA (Cambridge) Vol. 4; no. 11; pp. 1357 - 1372
• Main Authors: WIEDERKEHR, THOMAS, PRÉTÔT, RENÉ F., MINVIELLE-SEBASTIA, LIONEL
• Format: Journal Article
• Language: English
• Published: United States Cambridge University Press 01.11.1998; Cold Spring Harbor Laboratory Press
• Subjects: Alleles; Amino Acid Sequence; Basic-Leucine Zipper Transcription Factors; Biochemistry, Molecular Biology; DNA-Binding Proteins - genetics; Fungal Proteins - genetics; Fungal Proteins - metabolism; Genes, Essential - genetics; Genes, Fungal - genetics; Genes, Lethal - genetics; Genetics; Life Sciences; Molecular Sequence Data; Mutagenesis; Nuclear Proteins - genetics; Nuclear Proteins - metabolism; Pancreatitis-Associated Proteins; Phenotype; Polynucleotide Adenylyltransferase - genetics; Polyribosomes - metabolism; Protein Biosynthesis - genetics; Ribosomes - metabolism; RNA Processing, Post-Transcriptional; RNA, Fungal - genetics; RNA, Fungal - metabolism; RNA, Ribosomal, 18S - genetics; RNA, Ribosomal, 18S - metabolism; RNA, Small Nuclear - metabolism; RNA-Binding Proteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae Proteins; Schizosaccharomyces pombe Proteins; translation; U3 snoRNP; ribosome biogenesis
• ISSN: 1355-8382; 1469-9001
• DOI: 10.1017/S1355838298980955

To identify new genes involved in 3′-end formation of mRNAs in Saccharomyces cerevisiae, we carried out a screen for synthetic lethal mutants with the conditional poly(A) polymerase allele, pap1-7. Five independent temperature-sensitive mutations called lcp1 to lcp5 (for lethal with conditional pap1 allele) were isolated. Here, we describe the characterization of the essential gene LCP5 which codes for a protein with a calculated molecular mass of 40.8 kD. Unexpectedly, we found that mutations in LCP5 caused defects in pre-ribosomal RNA (pre-rRNA) processing, whereas mRNA 3′-end formation in vitro was comparable to wild-type. Early cleavage steps (denoted A0 to A2) that lead to the production of mature 18S rRNA were impaired. In vivo depletion of Lcp5p also inhibited pre-rRNA processing. As a consequence, mutant and depleted cells showed decreased levels of polysomes compared to wild-type cells. Indirect immunofluorescence indicated a predominant localization of Lcp5p in the nucleolus. In addition, antibodies directed against Lcp5p specifically immunoprecipitated the yeast U3 snoRNA snR17, suggesting that the protein is directly involved in pre-rRNA processing.