A temperature-sensitive lethal mutant nuc1-632 of Schizosaccharomyces pombe shows marked reduction in macromolecular synthesis and a defective nuclear phenotype with an aberrant nucleolus, indicating a structural role of the nuc1+ gene product in nucleolar organization. We cloned the nuc1+ gene by transformation and found that it appears to encode the largest subunit of RNA polymerase I. We raised antisera against nuc1+ fusion polypeptides and detected a polypeptide (approximately 190 kD and 2 x 10(4) copies/cell) in the S. pombe nuclear fraction. By immunofluorescence microscopy, anti-nuc1+ antibody revealed intense staining at a particular nuclear domain previously defined as the nucleolus. The nucleolar immunofluorescence by anti-nuc1+ was faded in nuc1-632 at restrictive temperature and dramatically diminished in the absence of DNA topoisomerases I and II. Thus active RNA polymerase I appears to be required for the formation of the nucleolus as its major component, and DNA topoisomerases appear to be required for the folding of rDNA and RNA polymerase I molecules into the functional organization of nucleolar genes.
Essential roles of the RNA polymerase I largest subunit and DNA topoisomerases in the formation of fission yeast nucleolus.
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T Hirano, G Konoha, T Toda, M Yanagida; Essential roles of the RNA polymerase I largest subunit and DNA topoisomerases in the formation of fission yeast nucleolus.. J Cell Biol 1 February 1989; 108 (2): 243–253. doi: https://doi.org/10.1083/jcb.108.2.243
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