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Neisseria gonorrhoeae and Neisseria meningitidis are exclusively human pathogens. A crucial property of the pathogenicity of neisserial infection is the ability to adhere to human epithelial cells. Pili mediate adherence of these bacteria to target cells and thereby promote colonization and infection of mucosal surfaces. In order to identify and to learn more about the initial event during infection, a cDNA clone from a human cervical epithelial cell line was identified in a panning experiment using purified gonococcal pili as probe. Upon transfection of the cloned cDNA into COS-7 cells, both gonococci and meningococci adhered to these otherwise non-binding cells. The deduced amino acid sequence of the cDNA clone showed homology to a recently reported human cDNA, called WWP2, that encodes an N-terminal C2-like domain. The C2 domain has been shown to bind membrane phospholipids in a calcium-dependent manner and is thought to function in the intracellular compartmentalization of proteins. Antiserum raised against the product encoded by the cDNA did not inhibit bacterial adherence, indicating that the cloned gene is most likely involved in up-regulation of a surface receptor for pathogenic Neisseria. Show less

The effect of caffeine on postreplication repair, as seen in alkaline sucrose gradients, conjugation, and ultraviolet light (UV) survival, was studied in excision deficient strains of Escherichia coli K12 and B. A caffeine concentration of 2 mg/ml was chosen for the study which did not inhibit colony formation. Both E. coli K12 AB2500 and E. coli B WWP2 were more sensitive to UV when plated on caffeine plates. Conjugation was not inhibited in the E. coli K12 strain; however, the same procedure confirmed caffeine inhibition in the E. coli B strain [17]. Caffeine did not inhibit postreplication repair in either strain, as determined by sedimentation profile studies of DNA on alkaline sucrose gradients. No strand breakage or degradation was observed in parental or post-UV replicated DNA for as long as 50 min incubation in caffeine. Thus caffeine concentrations that inhibited two recA gene product related phenomena did not cause immediate changes in size of DNA or inhibit the rate of a DNA gap generating postreplication type of DNA repair. Show less

Epidemiological studies and clinical observation suggesting potential hazards of arsenic compounds in increasing the incidence of cancer have been in complete contradiction with experimental findings in animals. Because of the predominance of skin cancers in the epidemiological reports, we decided to investigate the possibility that arsenic compounds might interfere with DNA repair. Using Escherichia coli as a test system, we show that this is indeed the case. Sodium arsenite, at concentrations of 0.1 mM and higher, decreases the survival of ultraviolet-irradiated E. coli WP2, a strain which possesses the full complement of repair genes. The effect of the arsenite increases with increasing ultraviolet dose. Similar results were obtained with the excision repair deficient strains WWP2 (uvrA) and WP6 (polA). Sodium arsenite had no effect on the survival of a recA mutant, WP10. Survival of ultraviolet-irradiated WP5 (exrA) was enhanced by sodium ardenite, the effect being greatest at low ultraviolet doses. It is postulated that arsenite inhibits a recA-dependent step in DNA repair. To account for the increased survival of the exrA mutant, we suggest that in the absence of the exr+ gene, the arsenite-sensitive recA-dependent function is deleterious. The ability of arsenite to inhibit DNA repair may account for the clinical and epidemiological reports linking arsenicals with an increased incidence of cancer. Show less

When the excision proficient strain E. coli WP2 Hcr+ trp- was grown to stationary phase by glucose starvation in M-9 minimal medium before UV -irradiation, the ability of nutrient broth enrichment of minimal medium to enhance trp- leads to Trp+ reversion was greatly reduced. Less than 50% of the Trp+ revertants were found to be ochre suppressors. However, in the WWP2 Hcr- strain, 75-86% of the tested revertants were ochre suppressors. This indicates that, under the cultural conditions employed, many potential suppressor mutations were removed by excision repair in the presence of broth enrichment. Broth enhancement of reversion also occurred in the Hcr- strain, which indicates that a less error-prone mode of recombination repair functions under minimal growth conditions. An Hcr+ strr derivative of WP2 Hcr+ was more resistant than its strs parent to the lethal effect of UV light and showed a lower UV-induced Trp+ reversion frequency. The percentage of Trp+ revertants that were due to ochre suppressors was markedly reduced in the strr strain. The Hcr- strr strain also had a lower UV-induced Trp+ reversion frequency than its strs parent. The excision repair inhibitor caffeine had little effect at lower UV doses on increasing Trp+ reversion in both Hcr+ strains. Acriflavine, however, was effective at lower UV doses in enhancing reversiin of the Hcr+ strains and the degree of enhancement increased with the dose. Acriflavine appeared to specifically enhance the number of ochre suppressing Trp+ revertants. In both Hcr- strains (strs and strr) caffeine (500 mug/ml) had no effect on survival but reduced the UV-induced Trp+ reversion frequency acting as an antimutagen. In contrast, acriflavine (2 mug/ml) decreased survival and increased the Trp+ reversion frequency of the Hcr- strains. The data on spontaneous Trp+ reversion frequencies show that the Hcr+ strs strain had a higher spontaneous reversion frequency than the Hcr- strs strain on all plating media. Further, caffeine was shown to reduce spontaneous Trp+ reversion in both Hcr+ and Hcr- strains while acriflavine increased the spontaneous reversion frequencies of both strains. Show less