Several parameters of phage T4 adsorption to and growth in Escherichia coli B/r were determined. All changed monotonously with the bacterial growth rate (p), which was modified by nutritional conditions. Adsorption rate was faster at higher p values, positively correlated to cell size, and increased by pretreatment with low penicillin (Pn) concentrations; it was directly proportional to total cellular surface area, indicating a constant density of 14 receptors on cell envelopes irrespective of growth conditions. Parameters of phage development and cell lysis were pdependent. The rate of phage release and burst size increased, while the eclipse and latent periods decreased with increasing p. Differentiation between the contribution of several physiological parameters to the development of 14 was performed by manipulating the host cells. A competitive inhibitor of glucose uptake, methyl a-o-glucoside, was exploited to reduce the growth rate in the same effective carbon source. Synchronous cells were obtained by the 'baby-machine' and large cells were obtained by pretreatment with low Pn concentrations. Lysis was delayed by superinfection, and DNA content and concentration were modified by growing a thy mutant in limiting thymine concentrations. The results indicate that burst size is not limited by cell size or DNA composition, nor directly by the rate of metabolism, but rather by the rates of synthesis and assembly of phage components and by lysis time. The rates of synthesis and assembly of phage components seem to depend on the content of the protein-synthesizing system and lysis time seems to depend on cellular dimensions.

The entire 127,923-bp sequence of the toxin-encoding plasmid pBtoxis from Bacillus thuringiensis subsp. israelensis is presented and analyzed. In addition to the four known Cry and two known Cyt toxins, a third Cyt-type sequence was found with an additional C-terminal domain previously unseen in such proteins. Many plasmid-encoded genes could be involved in several functions other than toxin production. The most striking of these are several genes potentially affecting host sporulation and germination and a set of genes for the production and export of a peptide antibiotic.Isolates of Bacillus thuringiensis are the biological control agents most widely used to eradicate insect pests of crops or vectors of human disease. For the latter application, Bacillus thuringiensis subsp. israelensis is the bioinsecticide of choice in programs worldwide to control mosquitoes and blackfly vectors (29). The insect pathogenicity of this bacterium depends on the presence of the pBtoxis megaplasmid (13) that encodes all six of the previously described toxins in this isolate (Cry4Aa, Cry4Ba, Cry10Aa, Cry11Aa, Cyt1Aa, and Cyt2Ba) (7,18). In addition, the plasmid carries several insertion sequences and encodes two further proteins (P19 and P20) with roles in promoting crystal formation and enhancing cell viability, probably by acting as chaperones (12,27,50). The pBtoxis plasmid has been partially mapped (6, 7), but the nucleotide sequence is limited to toxin genes and their flanking regions. Since the toxicity of the B. thuringiensis subsp. israelensis crystal is greater than that of any combination of the known toxins derived from it (9), it seems that other toxins or virulence factors may play a role in the activity of wild-type crystals. One possible source of such additional factors is the approximately 80% of the pBtoxis sequence that has not previously been analyzed. In order to understand fully this highly important virulence plasmid, we have therefore determined its entire nucleotide sequence as presented here. MATERIALS AND METHODSPlasmid preparation. The pBtoxis plasmid was prepared from B. thuringiensis subsp. israelensis strain 4Q2-72 (also known as 4Q5) and purified on a CsClethidium bromide density gradient as previously described (6).Sequencing and analysis. Plasmid DNA was sonicated and size fractionated on agarose gels. Two libraries were generated in pUC18 using insert sizes of 1.4 to 2 and 2 to 4 kb. Each clone was sequenced once from each end using ABI Big-Dye terminator chemistry on ABI3700 capillary sequencing machines. The final sequence was generated from 1,467 sequencing reads, giving 6.4-fold total coverage. All repeats were bridged by clone end read pairs or end-sequenced PCR products to confirm the assembly.The finished sequence was annotated using Artemis software (41). Potential coding sequences were identified by codon usage (34) and positional base preference methods and compared to the nonredundant protein databases using BLAST (3) and FASTA (38) software. The entire DNA sequence was also compared ...

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Average cell mass is shown to be inversely related to the concentration of thymine in the growth medium of a thystrain of Escherichia coli. The kinetics of the transition from one steady-state average cell mass to another was followed in an attempt to determine the relationship between the chromosome replication time and the time between completion of a round of chromosome replication and the subsequent cell division. Differences in average cell mass are shown to be associated with similar differences in average cell volume. Changes in volume associated with changes in thymine concentration are shown to be due primarily to differences in the width of cells. It is proposed that extension in length of the cell envelope occurs at a linear rate which is proportional to the growth rate and which doubles at the time of termination of rounds of replication. Changes in volume not associated with a change in growth rate are therefore accommodated by a change in cell width. Conditions are described under which average cell mass can continue to increase in successive generations and no steady-state average cell mass is achieved.

The kinesin-5 Saccharomyces cerevisiae homologue Cin8 is shown here to be differentially phosphorylated during late anaphase at Cdk1-specific sites located in its motor domain. Wild-type Cin8 binds to the early-anaphase spindles and detaches from the spindles at late anaphase, whereas the phosphorylation-deficient Cin8-3A mutant protein remains attached to a larger region of the spindle and spindle poles for prolonged periods. This localization of Cin8-3A causes faster spindle elongation and longer anaphase spindles, which have aberrant morphology. By contrast, the phospho-mimic Cin8-3D mutant exhibits reduced binding to the spindles. In the absence of the kinesin-5 homologue Kip1, cells expressing Cin8-3D exhibit spindle assembly defects and are not viable at 37°C as a result of spindle collapse. We propose that dephosphorylation of Cin8 promotes its binding to the spindle microtubules before the onset of anaphase. In mid to late anaphase, phosphorylation of Cin8 causes its detachment from the spindles, which reduces the spindle elongation rate and aids in maintaining spindle morphology.

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It is crucial to the reproducibility of results and their proper interpretation that the conditions under which experiments are carried out be defined with rigour and consistency. In this review we attempt to clarify the differences and interrelationships among steady, balanced and exponential states of culture growth. Basic thermodynamic concepts are used to introduce the idea of steady-state growth in open, biological systems. The classical, sometimes conflicting, definitions of steady-state and balanced growth are presented, and a consistent terminology is proposed. The conditions under which a culture in balanced growth is also in exponential growth and in steady-state growth are indicated. It is pointed out that steady-state growth always implies both balanced and exponential growth, and examples in which the converse does not hold are described. More complex situations are then characterized and the terminology extended accordingly. This leads to the notion of normal growth and growth that can be synchronous or otherwise unbalanced but still reproducible, and to the condition of approximate steady state manifested by growth in batch culture and by asymmetrically dividing cells, which is analysed in some detail.

SummaryCell cycle events have been proposed to be triggered by the formation of membrane domains in the process of coupled transcription, translation and insertion ('transertion') of nascent membrane and exported proteins. Disruption of domain structure should lead to changes in membrane dynamics. Membrane viscosity of Escherichia coli and Bacillus subtilis decreased after inhibition of protein synthesis by chloramphenicol or puromycin, or of RNA initiation by rifampicin, but not after inhibition of RNA elongation by streptolydigin or amino acid starvation of a stringent strain. The decrease caused by inhibitors of protein synthesis was prevented by streptolydigin if added simultaneously, but was not reversed if added later. The drug-induced decrease in membrane viscosity is energy dependent: it did not happen in KCN-treated cells. All treatments decreasing membrane viscosity also induced nucleoid compaction and fusion. Inhibition of macromolecular synthesis without membrane perturbation caused nucleoids to expand. Changes in membrane dynamics were also displayed during a nutritional shift-down transition that causes imbalance in macromolecular syntheses. The results are consistent with the transertion model, predicting dissipation of membrane domains by termination of protein synthesis or detachment of polysomes from DNA; domain structure is conserved if the transertion process is 'frozen'.