Twenty seven recessive temperature sensitive mutants have been isolated in Schizosaccharomyces pombe which are unable to complete the cell division cycle at the restrictive temperature. These mutants define 14 unlinked genes which are involved in DNA synthesis, nuclear division and cell plate formation. The products from most of these genes complete their function just before the cell cycle event in which they are involved. Physiological characterisation of the mutants has shown that DNA synthesis and nuclear division form a cycle of mutually dependent events which can operate in the absence of cell plate formation. Cell plate formation itself is usually dependent upon the completion of nuclear division.

We present homologies between archaeal and eucaryal DNA-dependent RNA polymerase (RNAP) It is now accepted that the living world is divided into three domains: Bacteria, Archaea, and Eucarya (1, 2). There are a sufficient number of molecular features specifically shared between Archaea and Eucarya to suggest a common ancestry, apart from the Bacteria. This is most clearly documented in their transcription systems. Although the large components of the RNA polymerases (RNAPs) are homologous among all domains, a much higher similarity exists between the archaeal and eucaryal versions than between either of these and the (eu)bacterial version (3, 4). The canonical archaeal transcription promoter closely resembles the eucaryal TATA-boxcontaining [RNA] polymerase (pol) II promoters (5). Sequences of all but one subunit of the RNAP of the extremely thermophilic archaeon Sulfolobus acidocaldarius have now been determined, thus allowing a comprehensive comparison of RNAPs among the three domains.In the Bacteria, transcription involves a single RNAP with only four different basic subunits: B, 3', a, and o,. In cyanobacteria and chloroplasts, the (3' component is replaced by two fragments of about equal size (6, 7). In certain species, additional components have been reported, some of which, at least, effect specific initiation of transcription (8-10).In contrast, the nuclei of Eucarya harbor three specialized RNAPs-pol I (or A), pol II (or B), and pol III (or C)-which have been well characterized in Saccharomyces cerevisiae (11). The two largest subunits in each case are homologous to the bacterial components (.3 and (3', and there is some structuralThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact. and functional resemblance between the bacterial a subunit and the eucaryal AC40 and B44 subunits (12)(13)(14).Limited similarity also has been claimed between the bacterial a subunit and the eucaryal AC19 and B12.5 subunits (14-16). Claimed homologies between the bacterial transcription initiation factor o-and certain eucaryal RNAP subunits (17) or transcription factors (18) This overview reports the results of sequence comparison between all but one of the RNAP subunits from Sulfolobus acidocaldarius, including the sequences of subunits D, E, and Nt and their homologs in S. cerevisiae. In conclusion we show that, despite the Archaea being prokaryotic in cell type, their transcription system resembles that of Eucarya and is thus different from that of Bacteria. Homology of Eucaryal and Archaeal Small RNAP SubunitsLike their eucaryal counterparts, the archaeal RNAPs show a high complexity. The RNAP of Sulfolobus acidocaldarius comprises 13 different single-copy subunits. A semiquantitative immunoblotting approach (3,4) showed that the three (or four; see below) largest archaeal subunits are homologs of the two largest eucaryal subunits and, therefore, are also...

The structure of the yeast RNA polymerase (pol) III was investigated by exhaustive two-hybrid screening using a library of random genomic fragments fused to the Gal4 activation domain. This procedure allowed us to identify contacts between individual polypeptides, localize the contact domains, and deduce a protein-protein interaction map of the multisubunit enzyme. In all but one case, pol III subunits were able to interact in vivo with one or sometimes two partner subunits of the enzyme or with subunits of TFIIIC. Four subunits that are common to pol I, II, and III (ABC27, ABC14.5, ABC10␣, and ABC10␤), two that are common to pol I and III (AC40 and AC19), and one pol III-specific subunit (C11) can associate with defined regions of the two large subunits. These regions overlapped with highly conserved domains. C53, a pol III-specific subunit, interacted with a 37-kDa polypeptide that copurifies with the enzyme and therefore appears to be a unique pol III subunit (C37). Together with parallel interaction studies based on dosagedependent suppression of conditional mutants, our data suggest a model of the pol III preinitiation complex.Eukaryotic transcription is mediated by large multiprotein complexes in which each of the three nuclear RNA polymerases (pols) interact with their cognate preinitiation factors. The pols themselves have been well characterized in terms of subunit composition, especially in the case of the yeast Saccharomyces cerevisiae. However, the spatial organization of the enzyme subunits and the way they interact with preinitiation complexes or with other components of the yeast nucleus are still poorly understood. Electron microscopy so far has provided the most accurate structural description of the Escherichia coli enzyme (1) and of yeast pol I (2, 3) and II (refs. 4-6 and references therein), revealing a striking similarity in the overall shape of these enzymes. In the case of yeast pol I, six subunits (or domains thereof) were localized by immunoelectron microscopy of antibody-labeled enzymes (2, 7). Sitespecific protein-DNA crosslinking also shed light on the general architecture of pol II (8, 9) and III (10-12) transcription complexes.These studies are still far from providing a comprehensive picture of the structural organization of the eukaryotic pols. Alternatively, each subunit can be tested for its ability to selectively associate with other subunits of the same heteromultimeric complex. In the case of human pol II, an in vitro test based on glutathione S-transferase pull-down assays has suggested numerous contacts within the pol II complex (13). In Schizosaccharomyces pombe, studies based on Far Western blotting, which were in some cases supported by independent protein-protein crosslinking studies, suggested that the two large pol II subunits interact with all of the other smaller subunits (9, 14). The two-hybrid system is an alternative to biochemical methods that allows one to detect interactions between proteins in the cellular context of the yeast nucleus (ref. 15 and ...

Hmo1 is one of seven HMG-box proteins of Saccharo myces cerevisiae. Null mutants have a limited effect on growth. Hmo1 overexpression suppresses rpa49-Delta mutants lacking Rpa49, a non-essential but conserved subunit of RNA polymerase I corresponding to the animal RNA polymerase I factor PAF53. This overexpression strongly increases de novo rRNA synthesis. rpa49-Delta hmo1-Delta double mutants are lethal, and this lethality is bypassed when RNA polymerase II synthesizes rRNA. Hmo1 co-localizes with Fob1, a known rDNA-binding protein, defining a narrow territory adjacent to the nucleoplasm that could delineate the rDNA nucleolar domain. These data identify Hmo1 as a genuine RNA polymerase I factor acting synergistically with Rpa49. As an HMG-box protein, Hmo1 is remotely related to animal UBF factors. hmo1-Delta and rpa49-Delta are lethal with top3-Delta DNA topoisomerase (type I) mutants and are suppressed in mutants lacking the Sgs1 DNA helicase. They are not affected by top1-Delta defective in Top1, the other eukaryotic type I topoisomerase. Conversely, rpa34-Delta mutants lacking Rpa34, a non-essential subunit associated with Rpa49, are lethal in top1-Delta but not in top3-Delta.