Alkali sensitivity in plasmid-bearing strains of Escherichia coli

Introduction of the ColV,I-K94 plasmid into any of four strains of Escherichia coli gave derivatives which grew less well than the parent at alkaline pH. The alkali sensitivity of the 1829 derivative resulted from the presence of the plasmid rather than from its introduction into a less alkali tolerant variant. Of two other ColV plasmids tested, one (ColV-K30) conferred substantial alkali sensitivity whereas the other (ColV-41) had little effect. Of several other plasmids examined, R124-F2 (which confers derepressed transfer properties) resembled ColV,I-K94 in its effect on alkali sensitivity and ColV-K98 produced a marked effect. The other plasmids (F lac , R124 itself, R1 and R483ColIa) had only a small effect. For ColV plasmids, it appears to be the presence of transfer and colicin components together which leads to the reduced tolerance to alkaline pHs.     

Col V plasmids and the resistance of Escherichia coli to divalent copper ions

Free organisms of both plasmid-free and plasmid-carrying strains of Escherichia coli were killed by incubation in water containing low levels of cupric ions. Sensitivity was temperature-dependent with killing being more marked at 20° or 25°C than at 10° or 15°C. In contrast to the effects of other inhibitors from natural waters (which affect free Col V⁺organisms more than Col^-ones), free Col^-and Col V⁺organisms were equally sensitive to kill by Cu²⁺. Attachment to glass beads essentially abolished sensitivity to cupric ions with full survival after exposure to 15 μ g/ml. This applied to both p⁺and p^-strains but attachment would have more effect on the survival of p⁺organisms in natural waters because some plasmids markedly enhance attachment.     

Habituation to alkali and increased u.v.-resistance in DNA repair-proficient and -deficient strains of Escherichia coli grown at pH 9.0

Repair-deficient strains of Escherichia coli carrying polAI or recA mutations were more alkali-sensitive than was their repair-proficient parent but, like strain 1829 ColV, I-K94, they showed habituation to alkali (induction of increased resistance) when grown at pH 9.0. Occurrence of such increased alkali resistance in the recA mutant implies that habituation to alkali does not depend on induction of SOS-related repair mechanisms. Organisms of repair-proficient and repair-deficient strains also became more resistant to u.v.-irradiation after growth at pH 9.0; this increased u.v.-resistance also appeared to be RecA-independent.     

Suppression by the ColV,I-K94 plasmid of a growth lesion in ompA mutants of Escherichia coli

Organisms of three independently isolated ompA mutants of Escherichia coli failed to form colonies on glucose minimal agar (glucose MA) at 44 degrees C after growth in glucose minimal salts medium at 37 degrees C, although all three strains formed colonies on nutrient agar at 44 degrees C. Supplementation of the glucose MA with individual amino acids including L-methionine and/or L-cysteine did not allow colony formation at 44 degrees C, although addition of 0.1% Casamino acids was effective; replacement of glucose with other energy sources or ammonium ions with glutamate also did not allow growth at 44 degrees C. The failure to form colonies at 44 degrees C was not due to killing of the organisms, because colonies were formed if plates of the ompA mutant initially incubated at 44 degrees C were shifted to 30 degrees C after 16 h. Introduction of the ColV, I-K94 plasmid into P678-54 ompA, 1131 ompA or an ompC ompA mutant suppressed the 44 degrees C growth lesion, but other plasmids (F lac, R483ColIa, RI, ColB-K98, R124) tested in P678-54 ompA did not. Growth of the ColV, I-K94+ derivative at 44 degrees C was due to a suppressing effect of the plasmid rather than to introduction of the plasmid into a variant with normal or altered OmpA protein. An attempt was made to ascertain which component(s) encoded by ColV, I-K94 was (were) responsible for allowing growth at 44 degrees C. Transfer components appeared unlikely to be involved and plasmids which conferred individual colicins (plus the corresponding immunity component) did not suppress.(ABSTRACT TRUNCATED AT 250 WORDS)     

Trimethyloxonium modification of single batrachotoxin-activated sodium channels in planar bilayers. Changes in unit conductance and in block by saxitoxin and calcium

Single batrachotoxin-activated sodium channels from rat brain were modified by trimethyloxonium (TMO) after incorporation in planar lipid bilayers. TMO modification eliminated saxitoxin (STX) sensitivity, reduced the single channel conductance by 37%, and reduced calcium block of inward sodium currents. These effects always occurred concomitantly, in an all-or-none fashion. Calcium and STX protected sodium channels from TMO modification with potencies similar to their affinities for block. Calcium inhibited STX binding to rat brain membrane vesicles and relieved toxin block of channels in bilayers, apparently by competing with STX for the toxin binding site. These results suggest that toxins, permeant cations, and blocking cations can interact with a common site on the sodium channel near the extracellular surface. It is likely that permeant cations transiently bind to this superficial site, as the first of several steps in passing inward through the channel.     

Relative apparent synapomorphy analysis (RASA). I: The statistical measurement of phylogenetic signal

We have developed a new approach to the measurement of phylogenetic signal in character state matrices called relative apparent synapomorphy analysis (RASA). RASA provides a deterministic, statistical measure of natural cladistic hierarchy (phylogenetic signal) in character state matrices. The method works by determining whether a measure of the rate of increase of cladistic similarity among pairs of taxa as a function of phenetic similarity is greater than a null equiprobable rate of increase. Our investigation of the utility and limitations of RASA using simulated and bacteriophage T7 data sets indicates that the method has numerous advantages over existing measures of signal. A first advantage is computational efficiency. A second advantage is that RASA employs known methods of statistical inference, providing measurable sensitivity and power. The performance of RASA is examined under various conditions of branching evolution as the number of characters, character states per character, and mutations per branch length are varied. RASA appears to provide an unbiased and reliable measure of phylogenetic signal, and the general approach promises to be useful in the development of new techniques that should increase the rigor and reliability of phylogenetic estimates.      

Sodium chloride induces an NhaA/NhaR-independent acid sensitivity at neutral external pH in Escherichia coli.

Escherichia coli previously grown in low-salt broth, pH 7.0, produced organisms which were markedly more acid sensitive when subsequently cultured in the same broth with 200 mM or more salt (NaCl) added. Induction of acid sensitivity occurred rapidly at both 37 and 30 degrees C, with a substantial effect within 15 min. Sensitization was partially inhibited by chloramphenicol and tetracycline and may depend on both protein synthesis-dependent and -independent physiological changes in the NaCl-induced organisms; sensitization did not result from osmotic shocking on transfer to challenge medium. Induction of acid sensitivity was affected by neither the sodium ion pore inhibitor amiloride nor the DNA synthesis inhibitor nalidixic acid; rifampin had a small effect, similar to that of chloramphenicol. Chlorides of other monovalent cations, especially Li+ and NH4+, also produced sensitization to acid, although CsCl was ineffective but did not interfere with sensitization by NaCl. Other sodium salts were also active as sensitizers, as were chlorides of divalent cations, but although sucrose (but not glycerol) was a good inducer, the results were not fully in accord with triggering of induction solely by the NaCl-associated increase in osmotic pressure. Sensitization was not prevented by deletion of the nhaA, nhaR, or nhaB gene. Acid sensitivity of NaCl-induced cells was slightly reduced after 90 min of growth at 37 degrees C in low-salt broth but was completely lost after 240 min. For NaCl-induced cells, acid killing in challenge media was not inhibited by amiloride. The NaCl-induced sensitization is distinct from the phenomenon of acid sensitivity induction in E. coli at alkaline external pH.     

Variation in heat-shock proteins among species of desert fishes (Poeciliidae, Poeciliopsis)

Analysis of the heat-shock proteins (hsps) of six closely related species of Poeciliopsis demonstrated the existence of biochemical diversity in the hsp100, hsp70, hsp60, and hsp30 protein families among species. Each species expressed five to seven hsp70-related isoforms. Constitutive 70-kD isoforms were identical among species, but four different patterns of heat-inducible isoforms were seen in these six species. Members of the hsp70 family of molecular chaperones are included among the most highly conserved proteins known, and the possibility of variation in hsp70 among closely related species has rarely been addressed. The hsp30 family is known to be less conserved than the hsp70 family, and, as expected, the Poeciliopsis hsp30 patterns showed more variation. Most of the hsp30 isoforms characteristic of a particular species were unique to that species. Hsp100 and hsp60 were identical in five of the species, but alternate isoforms were found in P. monacha. The small size and limited geographical distribution of the P. monacha population have probably contributed to the uniqueness of the monacha pattern. Two of the species were shown to acquire thermotolerance, the ability to withstand normally lethal temperatures when subjected to a gradual temperature increase. Rapid-heating protocols commonly used to establish critical thermal maxima of organisms do not include this inducible component of thermoresistance and therefore do not adequately assess an organism's capacity to withstand thermal stress.