Multimer formation reduces plasmid copy number and is an established cause of segregational instability. Nevertheless, it is difficult to rationalize observations that low levels of dimers can cause severe instability, if we assume they are distributed evenly in cell populations. We report here that dimer distribution is in fact heterogeneous in recombination-proficient strains. Most cells in the population contain only monomers; dimers are confined to a small subpopulation from which plasmid-free daughters arise at high frequency. In a rec+ culture where 4% of pBR322 molecules are dimers, more than half are in dimer-only cells. We show that this situation is inevitable because dimers replicate at twice the rate of monomers. Runaway multimerization is avoided because dimer-containing cells grow more slowly than their monomer-containing counterparts. A computer simulation is used to show how dimers proliferate after formation by homologous recombination. The equilibrium concentration of dimers is proportional to the inter-plasmid recombination rate and is essentially independent of the rate at which homologous recombination converts dimers to monomers.
The neurotransmitter norepinephrine (NE) stimulates the growth of low inocula of Escherichia coli in a minimal medium (SAPI) supplemented with serum (SAPI؉serum) and induces the production of an "autoinducer" (AI) which, in turn, promotes E. coli growth in the absence of NE. Given the importance of NE, epinephrine, and their corresponding adrenergic agonists and antagonists in clinical medicine, we sought to investigate the molecular basis for these observations. Using a variety of NE precursors, metabolites, and therapeutic agents, we demonstrated that their ability to stimulate E. coli growth in SAPI؉serum is dependent on the presence of a catechol (1,2-dihydroxybenzene) moiety with maximal activity requiring a two-carbon substituent incorporating a terminal primary amine. Serum contains the iron-binding glycoprotein, transferrin, and when SAPI؉serum was supplemented with sufficient Fe 3؉ to saturate transferrin, growth inhibition was relieved. Other metal cations, including Mg 2؉ , Ca 2؉ , and Zn 2؉ , had no effect. These data suggested that the stimulation of E. coli growth by NE in SAPI؉serum may involve the catecholate siderophore, enterobactin, a cyclic triester of 2,3-dihydroxybenzoylserine. Consistent with this hypothesis, E. coli strains with mutations in ferrienterobactin transport (fepA or tonB) or enterobactin biosynthesis (entA) did not respond to NE. Furthermore, NE induced expression of the ferrienterobactin receptor, FepA, during growth in SAPI؉serum. The enterobactin degradation product, 2,3-dihydroxybenzoylserine (DBS) was as effective as NE in stimulating the growth of E. coli and mutations in fepA or tonB abolished the DBS-dependent growth stimulation. In contrast to NE, however, DBS stimulated the growth of the entA mutant. Moreover, after growth in an ironlimited M9 medium in the absence of NE, ethyl acetate extracts of the E. coli entA ؉ parent but not of the entA mutant contained AI, i.e., stimulated the growth of E. coli in SAPI؉serum. Taken together, these data show that when low numbers of E. coli are inoculated into SAPI؉serum, NE, DBS, and related catecholamines induce the enterobactin iron uptake system. This, in turn, facilitates iron sequestration from transferrin and indicates that the AI present in NE-conditioned SAPI؉serum medium is enterobactin and its DBS breakdown products. Bacterial cells undergo a wide variety of physiological and morphological adaptations in response to chemical and physical changes in their environment. From the prokaryotic perspective, the successful interaction of bacterial cells with mammalian host tissues depends not only on a coordinated response to environmental cues such as nutrient availability, population density, temperature, osmolarity and pH (37, 56) but also on diverse host cell effector molecules. In general, the influence of host signaling molecules on bacteria has received relatively little attention (25). However, hormones such as epinephrine (12) and insulin (57), the neurotransmitter norepinephrine (NE [34]), and cytokines such as int...
Escherichia coli strains containing thermosensitivednaC alleles were studied by flow cytometry. Strains containing either the dnaC2 or dnaC28 allele were shifted between different temperatures, and DNA content distributions were gathered. Inhibition of initiation of chromosome replication at nonpermissive temperature, as well as reinitiation of replication at permissive temperature, were found to be affected by a number of parameters. These included the choice of permissive and nonpermissive temperatures, the length of the time of incubation at the nonpermissive temperature, the growth medium, the type of temperature shift used for reinitiation of replication (transient or nontransient), the genetic background of the host cell, and the cell concentration. Reinitiation of replication required neither transcription nor translation, whereas the elongation stage of replication was dependent upon ongoing protein synthesis in the mutants. Efficient use ofdnaC mutants for cell cycle studies is discussed.
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