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     

Neutral mutations and repetitive DNA

We have previously shown that computer simulations of processes that generate selectively advantageous changes together with random duplications and deletions give rise to genomes with many different genes embedded in a large amount of dispensable DNA sequence. We now explore the consequences of neutral changes on the evolution of genomes. We follow the consequences of sequence divergences that are neutral when they occur in dispensable sequences or extra copies of genes present in multigene families. We find that when divergence occurs at about the same frequency as duplication/deletion events, genomes carry repetitive sequences in proportion to their size. Inspection of the genomes as they evolved showed that multigene families were generated by relatively recent duplications of single genes and so would be expected to be highly homogeneous.     

Mutagenicity,sister chromatid exchange inducibility andin vitro cell transforming ability of particulates from Athens air

Airborne particulates were collected over a period of twelve months by the use of Hi-Vol samplers in the basin of Athens, Greece. N-Hexane extracts were tested in a battery ofin vitro tests for their ability to induce mutation in bacteria as well as mutation, sister chromatid exchange and morphological transformation in cultured mammalian cells. Positive results were found for mutagenicity withSalmonella strain TA98 in the Ames assay, for sister chromatid exchange induction in CHO cells and for transformation in BALB/c 3T3 cells in culture. They also showed weak non-doserelated induction of ouabain resistance in BALB/c 3T3 cells. The contribution of oxidizing and nitrating agents found in the Athens atmosphere, together with sunlight UV irradiation in the formation of direct acting mutagens and potential carcinogens from ambient polycyclic aromatic hydrocarbons, is suggested.Abbreviations FCS fetal calf serum - FPG fluorescent-plus-Giemsa technique - oua^R ouabain resistant - PAH polycyclic aromatic hydrocarbon - SCE sister chromatid exchange - TSP total suspended particulate     

A simple procedure for the isolation of streptomycin resistant plants in Solanaceae

A system has been developed for rapid selection of streptomycin resistant mutants, as adventitious shoots arising from explants of several Solanaceous species. Efficient mutagenesis was achieved by incubating shoot culture-derived leaf strips with 1 or 5 mM nitroso-methylurea, for 90 or 120 min. In Nicotiana tabacum and Lycopersicon peruvianum these treatments resulted in white or variegated adventitious shoots from up to 3.5% of explants placed on medium promoting shoot regeneration. Chlorophyll deficiencies were only observed very rarely in Solanum nigrum. Streptomycin resistant shoots were obtained from leaf explants placed on medium containing 500 mg l^-1 streptomycin sulphate, under which conditions explants are bleached and adventitious shoot development suppressed. Green adventitious s shoots appeared at a frequency dependent both on the mutagenic treatment and on the species. The best response was with S. nigrum where >70% of the explants produced streptomycin resistant shoots, most of which retained their resistance on subsequent testing. Maternal inheritance of streptomycin resistance has been confirmed for several N. tabacum and S. nigrum mutants, and there is also evidence for paternal transmission in the latter species. The procedure has been successfully extended to other species, including N. sylvestris and N. plumbaginifolia, and also to obtain spectinomycin resistant mutants.Communicated by R. Hagemann     

Isolation and characterization of Saccharomyces cerevisiae mutants supersensitive to G1 arrest by the mating hormone a-factor

Summary Nine independent mutants which are supersensitive (ssl ^–) to G1 arrest by the mating hormone a-factor were isolated by screening mutagenized Saccharomyces cerevisiae MAT cells on solid medium for increased growth inhibition with a-factor. These mutants carried lesions in two complementation groups, ssl1 and ssl2. Mutations at the ssl1 locus were mating type specific: MAT ssl1 ^– cells were supersensitive to -factor but MAT ssl1 ^– were not supersensitive to -factor. In contrast, mutations at the ssl2. locus conferred supersensitivity to the mating hormone of the opposite mating type on both MAT, and MATa cells. The -cell specific capacity to inactivate externally added a-factor was shown to be lacking in MAT ssl1 ^– mutants whereas MAT ssl2. cells were able to inactivate a-factor. Complementation analysis showed that ssl2 and sst2, a mutation originally isolated as conferring supersensitivity to -factor to MATa cells, are lesions in the same gene. The ssl1 gene was mapped 30.5 centi-Morgans distal to ilv5 on chromosome XII.     

A locus involved in kanamycin, chloramphenicol and L-serine resistance is located in the bglY-galU region of the Escherichia coli K12 chromosome

Summary Spontaneous mutants of Escherichia coli K12 displaying an increased level of the kanamycin resistance conferred by plasmid pGR71 were selected. Several mutants obtained in this way apparently carry large chromosomal deletions extending into galU and/or bglY (27 min). This positive selection of deletions allowed detection of a new locus located between galU and bglY. Deletions of this locus are responsible for increased resistance to kanamycin (Irk), decreased resistance to l-serine in minimal medium (Drs) and decreased resistance to chloramphenicol (Drc) when a cat gene is present in the bacteria.     

Ultraviolet photoproducts at the ochre suppressor mutation site in the glnU gene of Escherichia coli: relevance to "mutation frequency decline"

Summary Ochre suppressor mutations induced by UV in the Escherichia coli glnU tRNA gene are CG to TA transitions at the first letter of the anticodon-encoding triplet, CAA. Premutational UV photoproducts at this site have long been known to exhibit an excision repair anomaly (mutation frequency decline or MFD), whereby post-irradiation inhibition of protein synthesis enhances their excision and reduces suppressor mutation yields ten-fold. We sought to clarify the basis of this unique repair response by determining the spectrum of UV photoproducts on both strands of a 36 by region of glnU which includes the anticodon-encoding triplet. We found that four different photolesions are produced within the 3 by sequence corresponding to the tRNA anticodon: (i) on the transcribed strand, TC (6–4) photoproducts and TC cyclobutane dimers are formed in equal numbers at the site of the C to T transition, indicating that this site is a hotspot for the usually less frequent (6–4) photoproduct; (ii) on the nontranscribed strand, TT dimers are found opposite the second and third letters of the anticodon-encoding triplet, adjacent to the mutation site; and (iii) on the nontranscribed strand, an alkali-sensitive lesion other than a (6–4) photoproduct is formed, apparently at the G in the mutation site. We suggest that mutation frequency decline may reflect excision repair activity at closely spaced UV lesions on opposite strands, resulting in double-strand breaks and the death of potential mutants.     

Early-blocked sporulation mutations alter expression of enzymes under carbon control in Bacillus subtilis

Summary The physiological roles of the gene subset defined by early-blocked sporulation mutations (spo0) and their second-site suppressor alleles (rvtA11 and crsA47) remain cryptic for both vegetative and sporulating Bacillus subtilis cells. To test the hypothesis that spo0 gene products affect global regulation, we assayed the levels of carbon- and nitrogen-sensitive enzymes in wild-type and spo0 strains grown in a defined minimal medium containing various carbon and nitrogen sources. All the spo0 mutations (except spo0J) affected both histidase and arabinose isomerase levels in an unexpected way: levels of both carbon-sensitive enzymes were two- to six-fold higher in spo0 strains compared to wild type, when cells were grown on the derepressing carbon sources arabinose or maltose. There was no difference in enzyme levels with glucose-grown cells, nor was there a significant difference in levels of the carbonindependent enzymes glutamine synthetase and glucose-6-phosphate dehydrogenase. This effect was not due to a slower growth rate for the spo0 mutants on the poor carbon and nitrogen sources used. The levels of carbon-sensitive enzymes were not simply correlated with sporulation ability in genetically suppressed spo0 mutants, but the rvtA and crsA suppressors each had such marked effects on wild-type growth and enzyme levels that these results were difficult to interpret. We conclude that directly or indirectly the spo0 mutations, although blocking the sporulation process, increase levels of carbon-sensitive enzymes, possibly at the level of gene expression.     

Salmonella typhimurium LT2 metF operator mutations

Summary Using an Escherichia coli lac deletion strain lysogenized with lambda phage carrying a metF-lacZ gene fusion (Flac), in which -galactosidase levels are dependent on metF gene expression, cis-acting mutations were isolated that affect regulation of the Salmonella typhimurium metF gene. The mutations were located in a region previously defined as the metF operator by its similarity to the E. coli metF operator sequence. Regulation of the metF gene was examined by measuring -galactosidase levels in E. coli strains lysogenized with the wild-type Flac phage and mutant Flac phage. The results suggest that the mutations disrupt the methionine control system mediated by the metJ gene product, but not the vitamin B₁₂ control system mediated by the metH gene product. The results also demonstrate that negative control of the metF gene by the metH gene product and vitamin B₁₂ is dependent on a functional metJ gene product.Abbreviations Ap ampicillin - dNTP deoxyribonucleoside triphosphates - GM glucose minimal - Km kanamycin - L-agar Luria agar - LM lactose minimal - SAM s-adenosyl-L-methionine - TPEG phenylethyl -D-thiogalactoside - X-gal 5-bromo-4-chloro-3-indolyl -D-galactopyranoside - [] designates plasmid-carrier state - :: novel joint