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The function of Sufl, a well-studied substrate of the TatABC translocase in Eseherichia coli, is not known. It was earlier implicated in cell division, based on the finding that multiple copies of sufI suppressed the phenotypes of cells with mutations in flsI (fts123), which encodes a divisomal transpeptidase. Recently, sufI was identified as both a multicopy suppressor gene and a synthetic lethal mutant of ftsEX, which codes for a division-specific putative ABC transporter. In this study, we show that sufI is essential for the viability of E. coli cells subjected to various forms of stress, including oxidative stress and DNA damage. The sufI mutant also exhibits sulA-independent filamentation, indicating a role in cell division. The phenotypes of the sufI mutant are suppressed by factors that stabilize FtsZ ring assembly, such as increased expression of cell division proteins FtsQAZ or FtsN or the presence of the gain-of-function ftsA* (FtsA R286W) mutation, suggesting that SufI is a divisomal protein required during stress conditions. In support of this, multicopy sufI suppressed the divisional defects of mutants carrying the ftsA12, ftsQ1, or ftsK44 allele but not those of mutants earrying ftsZ84. Most of the division-defective mutants, in particular those carrying a [DELTA] flsEX or flsI23 allele, exhibited sensitivity to oxidative stress or DNA damage, and this sensitivity was also abolished by multiple copies of SufI. All of these data suggest that SufI is a division component involved in protecting or stabilizing the divisomal assembly under conditions of stress. Since sufI fulfils the requirements to be designated an fts gene, we propose that it be renamedflsP.

In Escherichia coli, at least 12 proteins, FtsZ, ZipA, FtsA, FtsE/X, FtsK, FtsQ, FtsL, FtsB, FtsW, FtsI, FtsN, and AmiC, are known to localize to the septal ring in an interdependent and sequential pathway to coordinate the septum formation at the midcell. The FtsEX complex is the latest recruit of this pathway, and unlike other division proteins, it is shown to be essential only on low-salt media. In this study, it is shown that flsEX null mutations are not only salt remedial but also osmoremedial, which suggests that FtsEX may not be involved in salt transport as previously thought. Increased coexpression of cell division proteins FtsQ-FtsA-FtsZ or FtsN alone restored the growth defects of flsEX mutants. flsEX deletion exacerbated the defects of most of the mutants affected in Z ring localization and septal assembly; however, the flsZ84 allele was a weak suppressor of flsEX. The viability of flsEX mutants in high-osmolarity conditions was shown to be dependent on the presence of a periplasmic protein, SufI, a substrate of twin-arginine translocase. In addition, SufI in multiple copies could substitute for the functions of FtsEX. Taken together, these results suggest that FtsE and FtsX are absolutely required for the process of cell division in conditions of low osmotic strength for the stability of the septal ring assembly and that, during high-osmolarity conditions, the FtsEX and SufI functions are redundant for this essential process.

A conditional lethal galE(Ts)-based strategy was employed in Escherichia coli, first to eliminate all growth-associated chromosomal reversions in lacZ or forward mutations in lacI/lacO by incubation at the restrictive temperature and subsequently to recover (as papillae) spontaneous mutations that had arisen in the population of nondividing cells after shift to the permissive temperature. Data from lacZ reversion studies in mutator strains indicated that the products of all genes for mismatch repair (mutHLS, dam, uvrD), of some for oxidative damage repair (mutMT), and of that for polymerase proofreading (dnaQ) are required in dividing cells; some others for oxidative damage repair (mutY, nth nei) are required in both dividing and nondividing cells; and those for alkylation damage repair (ada ogt) are required in nondividing cells. The spectrum of lacI/lacO mutations in nondividing cells was distinguished both by lower frequencies of deletions and IS1 insertions and by the unique occurrence of GC-to-AT transitions at lacO +5. In the second approach to study mutations that had occurred in nondividing cells, lacI/lacO mutants were selected as late-arising papillae from the lawn of a [galE.sup.+] strain; once again, transitions at lacO +5 were detected among the mutants that had been obtained from populations initially grown on poor carbon sources such as acetate, palmitate, or succinate. Our results indicate that the lacO +5 site is mutable only in nondividing cells, one possible mechanism for which might be that random endogenous alkylation (or oxidative) damage to DNA in these cells is efficiently corrected by the Ada Ogt (or Nth Nei) repair enzymes at most sites but not at lacO +5. Furthermore, the late-arising papillae from the second approach were composed almost exclusively of dominant lacI/lacO mutants. This finding lends support to 'instantaneous gratification' models in which a spontaneous lesion, occurring at a random site in DNA of a nondividing cell, is most likely to be fixed as a mutation if it allows the cell to immediately exit the nondividing state.

Research reveals the consensus target site sequence for IS186 transposition/insertion by radiation in Escherichia coli B.

Research has been conducted on the uvrD gene in Escherichia coli. Results indicate that null mutations in uvrD are incompatible with lon.

Results show that uup locus is part of a complex operon located downstream of a conserved gene and mutations in this locus affect excisions and deletions of chromosomal tandem repeats. Data further reveal a differential response of mini-transposon 10 and transposon 10 to mutations.

A genetic strategy was designed to examine the occurrence of mutations in stationary-phase populations. In this strategy, a parental population of cells is able to survive under both permissive and restrictive conditions whereas mutants at a particular target locus exhibit a conditional-lethal phenotype. Thus, by growing the population to stationary phase under restrictive conditions and then shifting it to permissive conditions, mutations that had arisen in stationary phase can be studied without confounding effects caused by the occurrence of similar mutations during growth of the population. In two different applications of this strategy, we have studied the reversion to [Lac.sup.+] in stationary phase of several [Lac.sup.-] mutations in Escherichia coli. Our results indicate that a variety of spontaneous point mutations and deletions, particularly those that are sensitive to the mechanisms of replication slippage (for their generation) and methyl-directed mismatch repair (for their correction), can arise in nondividing populations of cells within a colony. The frequency of their occurrence was also elevated in mutS strains, which are defective in such mismatch repair. These data have relevance to the ongoing debate on adaptive or directed mutations in bacteria.