Growth of Pseudomonas aeruginosa PAO1 at 15 to 45°C in tryptic soy broth resulted in changes in the lipids, lipopolysaccharides (LPSs), and outer membrane proteins of the cells. Cells grown at 15TC contained, relative to those cultivated at 45°C, increased levels of the phospholipid fatty acids hexadecenoate and octadecenoate and reduced levels of the corresponding saturated fatty acids. Furthermore, the lipid A fatty acids also showed thermoadaptation with decreases in dodecanoic and hexadecanoic acids and increases in the level of 3-hydroxydecanoate and 2-hydroxdodecanoate as the growth temperature decreased. In addition, LPS extracted from cells cultivated at the lower temperatures contained a higher content of long-chain S-form molecules than that isolated from cells grown at higher temperatures. On the other hand, the percentage of LPS cores substituted with side-chain material decreased from 37.6 mol% at 45°C to 19.3 mol% at 15°C. The outer membrane protein profiles indicated that at low growth temperatures there was an increase in a polypeptide with an apparent molecular weight of 43,000 and decreases in the content of 21,000 (protein Hl)-and 27,500-molecular-weight proteins.Since Pseudomonas aeruginosa is ubiquitous in nature and is a demonstrated pathogen for plants and insects as well as mamnmals we are interested in the changes that temperature may have on the cellular chemistry, particularly as it affects membrane lipids, lipopolysaccharides (LPSs), and outer membrane proteins. P. aeruginosa strains contain both ester-and amidebonded fatty acids. 3-Hydroxydodecanoic acid (3-OH C12:0) is found amide linked to C-2 of glucosamine residues in the lipid A component of LPS (9, 10). The lipid A also contains ester-bound 3-hydroxydecanoate (3-OH C10:0), dodecanoate (C12:0), 2-hydroxydodecanoate (2-OH C12:0), and a small amount of hexadecanoate (C16:0 141]). The chloroformmethanol-extractable cellular lipids (neutral and phospholipids) contain predominantly hexadecanoic (C16:0), hexadecenoic (C16:1), octadecanoic (C18:0), and octadecenoic (C18i1) acids (14,32,33). In addition, variable amounts of other fatty acids have been observed, including 17-and 19-cyclopropane acids and tetradecanoate (14, 32, 33). While the effects of temperature on the fatty acid profiles of a number of microorganisms, including psychrophilic Pseudomonas species, have been studied in detail (3,4,12,13, 20,21,26,29,31,35), only one study has been done on thermal adaptation of P. aeruginosa. The experiments of Gill and Suisted (13) with P. aeruginosa grown in a chemostat under conditions of nitrogen limitation showed that the extractable lipids of these cells possessed an increased content of unsaturated fatty acids when cultivated at suboptimal temperatures.Studies on the effect of growth temperature on LPS chemistry have been largely restricted to members of the family Enterobacteriaceae. Modifications to the composition of the lipid A component of LPS isolated from Salmonella species (43) We have demonstrated, on the basis of incr...
We have isolated, and characterized electrophoretically, two new lipopolysaccharide-defective (rough) mutants of Pseudomonas aeruginosa strain PAO. These strains, AK1401 and AK1414, together with two previously characterized isolates, AK1012 and AK1282, were used as recipients in transformation experiments with plasmid pR01614 DNA. The roughest mutant, AK1282, was not transformable, while the transformation efficiency of AK1012, and to a lesser extent the wild-type strain, was dependent upon the growth temperature. The two new isolates which are less rough than AK1012 were transformed at a frequency equivalent to that of the wild type-strain.
Nuclear factor I (NFI) is a site-specific DNA binding protein required for the replication of adenovirus DNA in vitro and in vivo. We have examined the effect of natural and synthetic binding sites for NFI (FIB sites) on RNA synthesis in HeLa whole cell extracts. The natural binding site used is the 26bp FIB-2 site previously isolated from the human genome. When present upstream of the TATA box of the adenovirus major late promoter, the FIB-2 site stimulates RNA synthesis 3 to 5-fold. This stimulation occurs with either orientation of the FIB-2 site. A point mutation in FIB-2 that decreases NFI binding at least 100-fold reduces, but does not completely abolish, the stimulation of transcription. A number of synthetic binding sites for NFI were tested for the ability to increase RNA synthesis. The strongest binding sites stimulated transcription the most, while the weakest sites had the least effect. These studies strongly suggest a role for NFI and cellular FIB sites in the control of RNA synthesis.
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