The work described in this thesis uses the fission yeast <i>Schizosaccharomyces pombe</i> as model system to analyse the molecular processes that control passage through the G<SUB>1</SUB> and S-phases of the cell-cycle. In particular, the work analyses the control of expression of <i>cdc22</i><SUP>+</SUP>, a gene whose transcript varies in abundance during the cell cycle with a maximum at the G<SUB>1</SUB>-S phase boundary. Chapter two describes the sequencing of <i>cdc22</i><SUP>+</SUP> and shows it encodes the large subunit of ribonucleotide reductase, an enzyme required for DNA precursor metabolism. In the next Chapter, <i>cis</i>-acting elements, resembling MCBs previously identified in budding yeast, are shown to be present 5' to the <i>cdc22</i><SUP>+</SUP> open reading frame. MCBs can confer cell cycle expression on a heterologous gene in fission yeast, implicating them in controlling periodic expression of <i>cdc22</i><SUP>+</SUP>. A <i>trans</i>-acting complex that specifically binds MCBs is identified, and called DSP1-for b DNA b synthesis control in <i>S.b pombe</i>. DSP1is related to DSC1, an MCB binding activity identified in budding yeast. Experiments in Chapter 3 demonstrate that the gene product of the <i>cdc10</i><SUP></SUP>+ START gene, p87<SUP>cdc10</SUP>, isa component of DSP1. Furthermore, <i>cdc22</i><SUP>+</SUP> is shown to be constitutively over-expressed in a <i>cdc10</i> mutant, <i>cdc10-C4</i>. Over-expression of <i>cdc22</i><SUP>+</SUP> in <i>cdc10-C4</i> is recessive to wild-type. It is proposed that DSP1 containing p87<SUP>cdc10-C4</SUP> is hyperactive and deregulated as a transcription complex. In summary, p87<SUP>cdc10</SUP> is part of a transcription complex that controls expression of <i>cdc22</i><SUP>+</SUP>, agene required for DNA synthesis. Thus this work demonstrates a molecular link between START and S-phase.

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The phenomenon of "epidemic spread" of a transmissible plasmid through a recipient population was re-examined. The phenomenon was studied quantitatively in the model system of F'lac in Escherichia coli K12. The rate of plasmid spread by retransfer was rather low because of the low frequency of continuing transfer by recipients that had received F'lac. The F'lac mating system was studied to answer the following questions: (i) What is the effect of parental concentration on mating efficiency? (ii) What proportion of parents are competent to mate? (iii) How many cells can competent parents mate with? (iv) How efficient is plasmid establishment in recipient cells after transfer? (v) How long are the lags between rounds of donor transfer and between a recipient receiving F'lac and becoming a competent donor? The variation in mating efficiency with parental concentration was incompatible with mating following bimolecular reaction kinetics and collision rate was not a limiting factor at concentrations above 5 x 107 cells/ml. The majority of both donors and recipients were competent to mate in 30', but whilst donors were fairly monogamous, recipients could mate with an average of 2-3 donors. Plasmid establishment after transfer was efficient and segregation had little effect on progeny growth rate. There was a lag of 30'-40' between rounds of donor competence and a lag of 90' before new progeny became competent donors. The rates of segregation due to plasmid incompatibility were calculated under several simple models. Experiments with two colEl derivatives were in reasonable agreement with the predictions of a model based on random pool replication. We studied the dependence on the recA function of the recombination needed for Hfr formation. We obtained Hfr clones from a recA F+ strain, but the rate of Hfr formation was only about 1% that in an isogenic rec+ strain, so Hfr formation in rec+ strains is probably mostly mediated by the host's normal recombination system. Our experiments suggested that "Type II" F+ strains were not defective in Hfr formation as had been previously reported, but had some secondary defect. The reA mutation reduced chromosome transfer in the same proportion as Hfr formation so the relationship between the two processes remains unclear.

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The morphogenes are genes required for cell growth and division in <I>Escherichia coli. </I>The morphogenes are often grouped as clusters in different regions of the chromosome. The <I>mra </I>region, located at 2 minutes on the <I>E. coli</I> chromosome is the largest cluster of morphogenes. It contains sixteen open reading frames from which fourteen genes have been identified as encoding for proteins involved in either cell division or the biosynthesis of the cell wall. The organisation of the cluster appears to be complex with the genes tightly packed and often overlapping. Many of the genes appear to share promoters for their expression. However, several of these genes have been incompletely characterized. The predicted proteins encoded by <I>ftsW, </I>a cell division gene, and <I>mraY</I>, a gene required for murein synthesis, have not yet been identified. One of the aims of this study was to identify the peptides produced from <I>ftsW </I>an <I>mraY. </I>The mutant phenotypes of these genes are poorly characterized. At the time of this study there was no temperature sensitive <I>ftsW </I>allele and there was no mutant allele of <I>mraY. </I>In this study null mutants were made for both <I>ftsW</I> and <I>mraY</I>. This allowed a more accurate characterization of their roles in cell division and cell growth, respectively. The isolation of the mutants allowed studies on their complementation. The promoter regions required for the independent expression of <I>ftsW </I>and <I>mraY</I> were then identified. The close proximity of genes in the <I>mra</I> region, the presence of overlaps and the sharing of promoters has provoked speculation that several of these genes may be translationally coupled. A previous study on three genes of the <I>mra</I> cluster revealed no translational coupling between the three genes but showed differences in the efficiency of translation of each gene product.

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The four essential cell division genes <i>ftsQ, ftsA, ftsZ</i>, and <i>envA</i> are arranged sequentially within a large cluster of genes required for cell envelope growth and form. The bacteriophage λJFL100 carries the 1.8 kilobase <i>Eco</i>R1-<i>Hin</i>dIII chromosomal restriction fragment which spans the <i>ftsQ</i> and part of <i>ftsA</i> coding sequences. This fragment contains at least two promoters; one within <i>ftsQ</i> is required for the expression of <i>ftsA</i> and one within <i>ftsA</i> is required for expression of <i>ftsZ</i>. The <i>fts</i> fragment is cloned upstream of a <i>lacZ</i> gene so that transcription originating from within the fragment leads to the production of β-galactosidase. Expression from the <i>fts</i> promoters in cells lysogenic for the phage is shown to be derepressed in the absence of FtsA protein. The results presented here suggest that transcription from these promoters is linked to the cell's periodic requirement for FtsA protein during septum formation. The exact positions of the promoters within the group are not yet known although their approximate locations have been determined by promoter assay and sequence analysis. An attempt has been made to define the position of the <i>ftsA</i> promoter more precisely by <i>in-vitro</i> transcription from short, defined templates and by the sequential deletion of <i>ftsQ</i>. Serial deletion through <i>ftsQ</i> has revealed what appears to be a complex upstream regulatory region, reminiscent of a eukaryotic enhancer element, which influences the expression of <i>ftsA</i>. A model is presented for the transcriptional control of <i>ftsA</i> expression.

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It has been shown (Devoret and George, 1967; Monk, 1969) that introduction of an ultraviolet-irradiated plasmid into a host bacterium can result in induction of prophage A in a lysogen, or suppression of cell division in a non-lysogen. In this thesis, the effect on cell division has been further studied. Irradiated P1 bacteriophages were used to infect calls of Escherichia coli X12 carrying the uvrA mutation. such cells are unable to excise pyrimidine dimers from DNA. The dimers induced in the phage genome by ultraviolet irradiation (Setlow, 1964) were measured by paper chromatography (Gunther and Prusoff, 1967). Inhibition of cell division was studied with respect to the effects of nutrients and host cell cycle on expression. Attempts were made to demonstrate replication of the damaged phage particle by differential radioactive labelling and density shift experiments. Autoradiography was used to determine the position of the damaged plasmid in the filament which it induced, and also the position of the cell chromosome strands which were present at the time of infection by the phage. The damaged phage was found to be located in the contra of the filament; this was not found when the phage was undamaged or subjected to phage repressor immediately on infection. The segregation pattern of host chromosomes differed from that found in penicillin-induced filaments. Cell membranes of P1-induced filaments, and control, uninfected, cells were examined by polyacrylamide gel electrophoresis (Laemmli, 1970; Lutkenhaus, 1977). The outer membrane of the filaments showed a changed profile, including a novel protein of molecular weight (Weber and Osborn, 1969) 53,000 daltons. The results are discussed in terms of two models, the SOS hypothesis (Radman, 1974) and the blocked replicon hypothesis (Donachie, 1974). The predictions of the blocked replicon hypothesis are fulfilled.