Genetic analysis of sterile mutants in the dpy-5 unc-13 (I) genomic region of Caenorhabditis elegans

Essential genes were identified in the 1.5-map unit dpy-5 unc-13 region of chromosome I in the Caenorhabditis elegans genome by rescuing lethal mutations using the duplication sDp2. In this paper, we report the mapping and complementation testing of lethal mutations, 45 of which identify 18 new, essential genes. This analysis brings the number of essential genes defined by the sDp2 rescue of lethal mutants to 97; 64 of these map between dpy-5 and unc-13. 61% of these essential genes are identified by more than one allele. Positioning of the mutations was done using the breakpoints of six duplications. The mutant phenotypes of 14 loci essential for fertility were characterized by Nomarski microscopy and DAPI staining. None of the mutants were rescued by wild-type male sperm. The cytological data showed that four genes produced mutants with defects in gonadogenesis, let-395, let-603, let-605 and let-610. Mutations in seven genes, let-355, let-367, let-384, let-513, let-544, let-545 and let-606, affected germ cell proliferation or gametogenesis. Mutants for the remaining three genes, let-370, let-599 and let-604, produced eggs that failed to develop or hatch, thereby acting as maternal effect lethals. We observed a nonrandom distribution of arrest phenotypes with regard to map position. Received: 8 May 1996 / Accepted : 27 January 1997     

Electron paramagnetic resonance saturation studies of P-700+ reaction center chlorophyll in plant photosynthesis

Electron paramagnetic resonance (EPR) power saturation and saturation recovery methods have been used to determine the spin lattice, T1, and spin-spin, T2, relaxation times of P-700+ reaction-center chlorophyll in Photosystem I of plant chloroplasts for 10 K less than or equal to T less than or equal to 100 K. T1 was 200 mus at 100 K and increased to 900 mus at 10 K. T2 was 40 ns at 40 K and increased to 100 ns at 10 K. T1 for 40 K less than or equal to T less than or equal to 100 K is inversely proportional to temperature, which is evidence of a direct-lattice relaxation process. At T = 20 K, T1 deviates from the 1/T dependence, indicating a cross relaxation process with an unidentified paramagnetic species. The individual effects of ascorbate and ferricyanide on T1 of P-700+ were examined: T1 of P-700+ was not affected by adding 10 mM ascorbate to digitonin-treated chloroplast fragments (D144 fragments). The P-700+ relaxation time in broken chloroplasts treated with 10 mM ferricyanide was 4-times shorter than in the untreated control at 40 K. Ferricyanide appears to be relaxing the P-700+ indirectly to the lattice by a cross-relaxation process. The possibility of dipolar-spin broadening of P-700+ due to either the iron sulfur center A or plastocyanin was examined by determining the spin-packet linewidth for P-700+ when center A and plastocyanin were in either the reduced or oxidized states. Neither reduced center A nor oxidized plastocyanin was capable of broadening the spin-packet linewidth of P-700+ signal. The absence of dipolar broadening indicates that both center A and plastocyanin are located at a distance at least 3.0 nm from the P-700+ reaction center chlorophyll. This evidence supports previous hypotheses that the electron donor and acceptor to P-700 are situated on opposite sides of the chloroplast membrane. It is also shown that the ratio of photo-oxidized P-700 to photoreduced centers A and B at low temperature is 2 : 1 if P-700 is monitored at a nonsaturating microwave power.     

Nucleic acid relationships among Acholeplasma species.

3H-labeled Acholeplasma DNA probes were generated in vitro by the nick-translation method and used to determine the nucleotide sequence homology among the type strains of the eight currently recognized species of Acholeplasma. Very little nucleotide sequence homology (less than or equal to 18%) was found among the eight species, with heteroduplexes showing at least 12% or more mismatching as determined by thermal elution midpoints. The small amount of nucleotide sequence homology among the eight species indicates that these species are quite distinct and are not closely related to each other genomically.     

DIFFERENTIAL INCORPORATION OF [3H]LYSINE INTO VISUAL CORTEX PROTEIN FRACTIONS DURING FIRST EXPOSURE TO LIGHT

—A resolution of the enhancement of protein synthesis in the visual cortex of rats during first exposure to light (Richardson and Rose , 1972) was achieved by polyacrylamide gel electrophoresis using a double-labelling technique. Differential incorporation of lysine was established between exposed and control animals in two fractions of the soluble proteins and seven fractions of the insoluble proteins. This suggests that exposure to a new experience of this type involves a specific effect on protein synthesis, rather than a general stimulation across all fractions.     

β-Ar-ACETYL D-GALACTOSAMINIDASE IN BULK SEPARATED NEURONS AND NEUROPIL FROM RAT CEREBRAL CORTEX

Abstract— β- N -Acetyl D-galactosaminidase was studied in isolated neuronal and neuropil fractions from cerebral cortex and subcellular fractions derived from them. Although the enzyme activity evinced some latency properties, its subcellular distribution pattern was broader than that observed with other acid hydrolases. By contrast with nine other acid hydrolases, it was more active in neuropil than neuronal fractions (neuronal/neuropil activity ratio 0.63). This ratio was preserved in lysosomal subfractions derived from the isolated cell fractions. The data is taken as further evidence for the microheterogeneity of lysosomal particles from the brain.     

Improving Antibiotic Activity against Wound Pathogens with Manuka Honey In Vitro

Following the discovery of synergistic action between oxacillin and manuka honey against methicillin-resistant Staphylococcus aureus, this study was undertaken to search for further synergistic combinations of antibiotics and honey that might have potential in treating wounds. Fifteen antibiotics were tested with and without sublethal concentrations of manuka honey against each of MRSA and Pseudomonas aeruginosa using disc diffusion, broth dilution, E strip, chequerboard titration and growth curves. Five novel antibiotic and manuka honey combinations were found that improved antibacterial effectiveness in vitro and these offer a new avenue of future topical treatments for wound infections caused by these two important pathogens.     

Burn Injury Reduces Neutrophil Directional Migration Speed in Microfluidic Devices

Thermal injury triggers a fulminant inflammatory cascade that heralds shock, end-organ failure, and ultimately sepsis and death. Emerging evidence points to a critical role for the innate immune system, and several studies had documented concurrent impairment in neutrophil chemotaxis with these post-burn inflammatory changes. While a few studies suggest that a link between neutrophil motility and patient mortality might exist, so far, cumbersome assays have prohibited exploration of the prognostic and diagnostic significance of chemotaxis after burn injury. To address this need, we developed a microfluidic device that is simple to operate and allows for precise and robust measurements of chemotaxis speed and persistence characteristics at single-cell resolution. Using this assay, we established a reference set of migration speed values for neutrophils from healthy subjects. Comparisons with samples from burn patients revealed impaired directional migration speed starting as early as 24 hours after burn injury, reaching a minimum at 72–120 hours, correlated to the size of the burn injury and potentially serving as an early indicator for concurrent infections. Further characterization of neutrophil chemotaxis using this new assay may have important diagnostic implications not only for burn patients but also for patients afflicted by other diseases that compromise neutrophil functions.