Meiotic mutants of rye Secale cereale L. II. The nonhomologous synapsis in desynaptic mutants sy7 and sy10

We studied the expression and inheritance of two spontaneous mutations found in different populations of rye Secale cereale L. that cause high univalent frequency in meiosis and low fertility. Both mutations were inherited as monogenic recessives. For each of the mutations the corresponding gene symbols (sy7 and sy10) were suggested although their allelism has not been studied. These mutants differ in chiasma frequency and in the number of univalents per meiocyte. Electron microscopy of the wholemount surface-spread synaptonemal complexes (SCs) from microsporocytes of both mutants revealed that during meiotic prophase I random synapsis began and progressed that involved not only homologous but also nonhomologous chromosomes. SCs were formed with frequent changes of pairing partners (switches) and intrachromosomal foldbacks of unpaired axial elements. As a result, incompletely synapsed, non-homologous and multivalent SCs were formed in mutants by the stage analogous to pachytene in normal plants. In sy7 a maximum in the number of switches and foldbacks were observed at zygotene, whereas in sy10 this occurred at pachytene. We suggest that it is the process of recognition of homology that is impaired in both mutants. This leads to indiscriminate synapsis and prevents chiasma formation. Both mutants may be classified as desynaptic.     

The genetic analysis of meiosis in female Drosophila melanogaster.

The three major features of meiosis are first synapsis, then exchange, and finally, disjunction of homologous chromosomes; these phenomena occur before pachytene, during pachytene, and after pachytene respectively. The effects of meiotic mutants, or other perturbations, either endogenous or exogenous, on the meiotic process may be assigned tentatively to one of these intervals, based on the earliest discernible abnormality. Thus mutants exhibiting abnormal disjunction and normal exchange affect post-pachytene functions; mutants exhibiting abnormal disjunction and exchange but with ultrastructurally normal appearing synaptonemal complex affect pachytene functions; and mutants with abnormal disjunction, exchange, and synaptonemal complex affect prepachytene functions. This rationale is applied to the temporal seriation of effects of meiotic mutants and chromosomal abnormalities on the meiotic programme.     

Mutants of TETRAHYMENA THERMOPHILA with Temperature-Sensitive Food Vacuole Formation. I. Isolation and Genetic Characterization

Germ-line mutants have been isolated in Tetrahymena thermophila that have recessive, temperature-sensitive defects in phagocytosis. Nitrosoguanidine-mutagenized cells were induced to undergo cytogamy, and clones were isolated that were unable to form food vacuoles after two days of growth at 39°. Most of the mutants belong to a single complementation group, designated vacA. They have defects in oral development—not in phagocytosis per se—that are undetectable under light microscopy. One fertile mutant, phenotypically indistinguishable from the vacA group, has its vac mutation(s) restricted to the macronucleus, and it is a heterokaryon for two other markers. This clone probably resulted from a failure of the two gametic nuclei to fuse after normal exchange. Two additional mutants were studied, but their sterility prevented a full genetic analysis. One of these clones has a rudimentary oral apparatus and defective contractile vacuole pores; both defects may be determined by the same mutation. The other clone has a structurally normal oral apparatus and may be defective in phagocytosis per se.—The induction and characterization of germ-line mutations that affect oral development open the way for the genetic dissection of the morphogenesis of a complex eukaryotic organelle, and make available additional useful mutants for the study of nutrition and transmembrane active transport.     

Sterile mutants in Caenorhabditis elegans: the synaptonemal complex as an indicator of the stage-specific effect of the mutation

Two sterile mutants of Caenorhabditis elegans hermaphrodites have been examined using the electron microscope and serial section analysis. The F4 and F80 mutants were described previously ( Mounier and Brun , 1980), and they were shown to be blocked at the start of oogenesis. In the F4 mutant, normal sperm are produced and the pachytene nuclei contain tripartite synaptonemal complexes (SC) between the homologously paired chromosomes. In the F80 asynaptic mutant, only a few sperm are produced and they have abnormal morphology. Whereas SCs and SC associated structures (termed 'SC knobs') are present in the F4, these structures are absent from the F80 . The mutation in F80 affects gametogenesis prior to the pachytene stage of meiosis and pairing of homologous chromosomes apparently does not occur. The SC knobs may influence the regulation of the disjunction of the chromosomes. For that reason, these structures are now termed 'disjunction regulator regions'.     

The maize desynaptic1 mutation disrupts meiotic chromosome synapsis

In most eukaryotes, homologous chromosomes undergo synapsis during the first meiotic prophase. A consequence of mutations that interfere with the fidelity or completeness of synapsis can be failure in the formation or maintenance of bivalents, resulting in univalent formation at diakinesis and production of unbalanced spores or gametes. Such mutations, termed desynaptic mutations, can result in complete or partial sterility. We have examined the effect of the maize desynaptic1-9101 mutation on synapsis, using the nuclear spread technique and electron microscopy to examine microsporocytes ranging from early pachytene until the diplotene stage of prophase I. Throughout the pachytene stage, there was an average of about 10 sites of lateral element divergence (indicating nonhomologous synapsis), and during middle and late pachytene, an average of two and three sites of foldback (intrachromosomal) synapsis, per mutant nucleus, respectively. By the diplotene stage, the number of sites of lateral element divergence had decreased to seven, and there was an average of one foldback synapsis site per nucleus. Lateral element divergence and foldback synapsis were not found in spread pachytene nuclei from normal plants. These results imply that the normal expression of the dsy1 gene is essential for the restriction of chromosome synapsis to homologues. The abundance of nonhomologous synapsis and the persistence of extended stretches of unsynapsed axial elements throughout the pachytene stage of dsy1–9101 meiocytes suggests that this mutation disrupts both the fidelity of homology search and the forward course of the synaptic process. This mutation may identify a maize mismatch repair gene. Dev. Genet. 21:146–159, 1997. © 1997 Wiley-Liss, Inc.     

Mutational analysis of morphogenesis of the maize embryo

Morphogenesis of the maize embryo is controlled by many genes. A group of 51 embryo-specific (emb) mutations representing at least 45 independent mutation events and many different gene loci have been isolated from active Robertson's Mutator stocks. The authors have reported previously that the embryo phenotype of 27 of these mutations, characterized by examining mature embryos in fresh dissection. The maximal development capacity of the 24 emb mutations are reported here which have not been reported previously. All result in retarded embryos that are morphologically abnormal. Three of the mutants are blocked during the first phase of morphogenesis, the period in which the basal-apical asymmetry is established and the embryo is regionalized into suspensor and embryo proper. Nineteen mutants are blocked during the second phase, the period in which radial asymmetry appears, the embryonic axis is established at a different angle than the original basal-apical axis of the zygote and the vegetative organ primordia of the adult plant make their first appearance. Two mutants are blocked or altered during the third phase, the period in which vegetative structures are elaborated. Some of the mutants affected in the first phase of morphogenesis may have defective mechanisms for establishing basal-apical asymmetry, including possibly the asymmetric distribution of morphogenic determinants. Similarly, some of the mutants affected in the second phase may be altered in the mechanisms establishing radial asymmetry and the origin of the meristems. Mutations of the first type may act as early as the first cell division when the zygote undergoes a transverse division, while mutations of the second type are likely to act during the proembryo and transition stages. Both types include mutations affecting embryo pattern formation. Mutations affecting the third phase of morphogenesis may identify genes regulating reiterative morphogenic processes of vegetative plant development and events of embryo maturation. This group of 24 mutations is like that reported previously in representing genes that are crucial to embryo morphogenesis.     

Sterol mutants of Neurospora crassa: their isolation, growth characteristics and resistance to polyene antibiotics

Summary A method is described for isolating sterol mutants of the filamentous fungus, Neurospora crassa. Most of the mutants carry gene mutations affecting the later stages of ergosterol biosynthesis and they accumulate other, as yet unidentified, sterol components but two mutants are blocked earlier in the pathway and respond to exogenous mevalonic acid. Altered sterol metabolism is associated with a reduced rate of growth, abnormal morphology, poor fertility and resistance to a variety of polyene antibiotics.     

Cytology of recessive sexual-phase mutants from wild strains of Neurospora crassa.

Wild-collected strains of Neurospora crassa harbor recessive mutations that are expressed in the sexual phase when homozygous. Thirty-two representative mutants that produced barren perithecia were examined cytologically. Six of these mutants failed to form asci. Of the remaining 26, chromosome pairing was disturbed in 12 and meiosis was disturbed at pachytene or diplotene in 5. Seven mutants showed normal meiosis I but then diverged from the normal sequence, and two showed perithecial beak abnormalities. In many mutants, ascus development and nuclear divisions continued after the initial defect, albeit abnormally. Nuclear divisions were often delayed, essentially uncoupling them from other ascus events such as the formation of enlarged spindle pole body plaques, ascospore wall membranes, and spore delimitation. All 32 mutants were recessive and none showed obvious morphological abnormalities during vegetative growth. This phenotype contrasts sharply with that of numerous laboratory-induced ascus mutants, which are frequently expressed pleiotropically in the vegetative phase and several are dominant in the sexual phase.     

Mutations affecting uridine monophosphate pyrophosphorylase or the argR gene in Escherichia coli. Effects on carbamoyl phosphate and pyrimidine biosynthesis and on uracil uptake

Summary In the course of experiments directed towards the isolation of mutants of Escherichia coli K12 with altered regulation of the synthesis of carbamoylphosphate synthetase, two types of mutations were found to affect the cumulative repression of this enzyme by arginine and uracil. Alteraction of the arginine pathway regulatory gene, argR, was shown to reduce the repressibility of the enzyme by both end products while mutations affecting uridine monophosphate pyrophosphorylase (upp) besides affecting uracil uptake preclude enzyme repression by uracil or cytosine in the biosynthesis of carbamoylphosphate and the pyrimidines. The upp mutations were located on the chromosome near the gua operon. Mutations previously designated as uraP are shown to belong to this class.The relation that could exist between the loss of uridine monophosphate pyrophosphorylase and the impairment of uracil uptake is discussed.A new method for isolating argR mutants in arginine-less strains is described.     

Respiratory-deficient Mutants in Saccharomyces lactis

Six independent ultraviolet-induced respiratory-deficient mutants (petites) of Saccharomyces lactis were isolated and characterized. Two possessed a normal cytochrome spectrum, another displayed an increased level of all the cytochromes, and three suffered from a partial or complete loss of one or more of the cytochromes a, b, c, and c₁. All of the mutants were segregational petites; none was vegetative. Determination of linkage relationships between mutants was restricted because matings between mutants, homozygous or heterozygous, for loci affecting cytochrome content were blocked at various stages in the mating-sporulating sequence. At least three of the petites were genetically nonidentical. Three of the mutations appeared to occupy loci within the same linkage group; two of the three mutations that mapped within this region were cytochrome-deficient. Growth at high or low temperatures, under increased osmotic pressure or in media supplemented with various fatty acids or sterols, did not relieve the physiological defects in these mutants. Reasons for the differences in survival of segregational and vegetative petites within this species are examined.     

Isolation and characterization of stable mutants ofStreptomyces peucetius defective in daunorubicin biosynthesis

Daunorubicin and its derivative doxorubicin are antitumour anthracycline antibiotics produced byStreptomyces peucetius. In this study we report isolation of stable mutants ofS. peucetius blocked in different steps of the daunorubicin biosynthesis pathway. Mutants were screened on the basis of colony colour since producer strains are distinctively coloured on agar plates. Different mutants showed accumulation of aklaviketone, ε-rhodomycinone, maggiemycin or 13-dihydrocarminomycin in their culture filtrates. These results indicate that the mutations in these isolates affect steps catalysed bydnrE (mutants SPAK and SPMAG),dnrS (SPFS and SPRHO) anddoxA (SPDHC) gene products.     

Interaction between avoidance of photon absorption,excess energy dissipation and zeaxanthin synthesis against photooxidative stress in Arabidopsis

Plants evolved photoprotective mechanisms in order to counteract the damaging effects of excess light in oxygenic environments. Among them, chloroplast avoidance and non‐photochemical quenching concur in reducing the concentration of chlorophyll excited states in the photosynthetic apparatus to avoid photooxidation. We evaluated their relative importance in regulating excitation pressure on photosystem II. To this aim, genotypes were constructed carrying mutations impairing the chloroplast avoidance response (phot2) as well as mutations affecting the biosynthesis of the photoprotective xanthophyll zeaxanthin (npq1) or the activation of non‐photochemical quenching (npq4), followed by evaluation of their photosensitivity in vivo. Suppression of avoidance response resulted in oxidative stress under excess light at low temperature, while removing either zeaxanthin or PsbS had a milder effect. The double mutants phot2 npq1 and phot2 npq4 showed the highest sensitivity to photooxidative stress, indicating that xanthophyll cycle and qE have additive effects over the avoidance response. The interactions between non‐photochemical quenching and avoidance responses were studied by analyzing the kinetics of fluorescence decay and recovery at different light intensities. phot2 fluorescence decay lacked a component, here named as qM. This kinetic component linearly correlated with the leaf transmittance changes due to chloroplast relocation induced by white light and was absent when red light was used as actinic source. On these basis we conclude that a decrease in leaf optical density affects the apparent non‐photochemical quenching (NPQ) rise kinetic. Thus, excess light‐induced fluorescence decrease is in part due to avoidance of photon absorption rather than to a genuine quenching process.     

The synaptonemal complexes of Caenorhabditis elegans: the dominant him mutant mnT6 and pachytene karyotype analysis of the X-autosome translocation

The dominant X-autosome heterozygous translocation mutant mnT6 of the nematode Caenorhabditis elegans has an X chromosome that has been reduced in size by 40%, yet the remainder of the bivalent pairs effectively at pachytene and has a synaptonemal complex (SC) that has a normal appearance. Six SCs are present in pachytene nuclei of this mutant which correspond to a haploid value of n=6. Nondisjunction of the X chromosome occurs at a rate of 37% and there are no Disjunction Regulator Regions (DRR) in this him (high incidence of males) mutant. This is consistent with the notion that DRRs either promote disjunction or inhibit nondisjunction of the X chromosome. Their occurrence in pachytene nuclei is independent of the mechanism responsible for nondisjunction, i.e. point mutations as in him-8 versus chromosomal aberrations as in mnT6. Although an SC is present along the entire length of the X chromosome, crossover suppression is observed in mnT6.     

Temperature-sensitive respiratory-deficient mitochondrial mutations: Isolation and genetic mapping

Summary In order to find new genetic loci and functions on the yeast mitochondrial DNA, especially mutations affecting the mitochondrial protein synthesis apparatus, temperature sensitive mutants have been isolated after MnCl₂ mutagenesis and mitochondrial and nuclear mutants classified according to their pattern of recombination with three rho^- tester strains.Eighteen cold- and heat-sensitive respiratory deficient mitochondrial mutants have been isolated and localized on the mitochondrial genome by deletion mapping using 113 rho^- strains. Eight of them appear to represent new loci, among which some are probably mutations of the tRNA and rRNA genes.     

Restoration of enzyme activity by recessive missense suppressors in the fungus Coprinus.

Summary The acu-1 locus in Coprinus is the structural gene for acetyl-CoA synthetase. Five suppressor gene mutations, which suppress the acu-1,34 missense allele, were induced by mutagen treatment. All five suppressors were shown to have properties expected for tRNA structural gene mutations: they are recessive, they show a gene dosage effect in any doubly heterozygous combination of two sup ⁺ mutations and they are allele specific in action.Crosses between suppressed mutants established that at least four suppressor loci were represented. Doubly suppressed mutants derived from these crosses were used to show that the gene dosage effect is maintained when two sup ⁺ mutations are in cis as well as trans combinations in the two nuclei of the basidiomycete dikaryon.Extracts of the unsuppressed acu-1.34 mutant contained less than 2% of wild type acetyl-CoA synthetase activity whereas extracts of four of the five suppressor strains showed activities ranging from 28 to 37% of wild type. Only a slight increase in activity was detected in the fifth suppressor strain but this was associated with a temperature sensitive sup ⁺ phenotype. All five sup ⁺ mutations restored the ability of the acu-1.34 mutant to induce isocitrate lyase, an enzyme which, under the conditions of growth used, can only be induced when acetyl-CoA synthetase activity is present. Thus all five suppressors act to restore normal acu-1 protein function.     

Genetic analysis of mutants of Aspergillus nidulans blocked at an early stage of sporulation.

Three mutants of Aspergillus nidulans, selected to have a block at an early stage of conidiation (asexual sporulation), exhibit similar pleiotropic phenotypes. Each of these mutants, termed preinduction mutants, also are blocked in sexual sporulation and secrete a set of phenolic metabolites at level much higher than wild type or mutants blocked at later stages of conidiation. Backcrosses of these mutants to wild type showed that the three phenotypes always cosegregated. Diploids containing the mutant alleles in all pairwise combinations were normal for all phenotypes, showing that the three mutations are nonallelic. This conclusion was confirmed by the finding that the mutations map at three unlinked or distantly linked loci. Ten revertants of the two least leaky preinduction mutants, selected for ability to conidiate, were found in each case to arise by a second-site suppressor mutation. All of the revertants still showed accumulation of some of the phenolic metabolites but differed from each other in certain components. Three of the revertants retained the block in sexual sporulation. In these cases the suppressor has thus uncoupled the block in asexual sporulation from the block in sexual sporulation. These results are understandable in terms of a model in which preinduction mutations and their suppressors affect steps in a single metabolic pathway whose intermediates include an effector specific for asexual sporulation and a second effector specific for sexual sporulation.     

Isolation and characterization of hydroxylamine-induced mutations in the Erwinia herbicola ice nucleation gene that selectively reduce warm temperature ice nucleation activity

Cells of ice nucleation active bacterial species catalyse ice formation over the temperature range of -2 to -12°C. Current models of ice nucleus structure associate the size of ice nucleation protein aggregates with the temperature at which they catalyse ice formation. To better define the structural features of ice nucleation proteins responsible for the functional heterogeneity of ice nuclei within a genetically homogeneous collection of cells we used in vitro chemical mutagenesis to isolate mutants with reduced ability to nucleate ice at warm assay temperatures but which retain normal or near normal nucleation activity at cold temperatures (WIND, i.e. w arm i ce n ucleus-d eficient mutants). Nearly half of the mutants obtained after hydroxylamine mutagenesis of the iceE gene from Erwinia herbicola had this phenotype. The phenotypes and location of lesions on the genetic map of iceE were determined for a number of mutants. All WIND mutations were restricted to the portion of iceE encoding the repetitive region of the poty peptide. DNA sequencing of two WIND mutants revealed single nucleotide substitutions changing a conserved serine or glycine residue to phenylalanine and serine, respectively. The implications of these findings in structure/function models for the ice nucleation protein are discussed.     

The Ph1 and Ph2 loci play different roles in the synaptic behaviour of hexaploid wheat Triticum aestivum

Triticum aestivum is an allohexaploid wheat (AABBDD) that shows diploid-like behaviour at metaphase-I. This behaviour is influenced by the action of several loci, Ph1 and Ph2 being the main loci involved. To study the effect of these two loci on chromosome pairing in T. aestivum we have analysed the synaptic pattern in fully traced spread nuclei at mid- and late-zygotene, and at pachytene, of three different genotypes of cv Chinese Spring: standard line, ph1b and ph2b mutants. The analysis of the synaptic progression showed that only a few nuclei accomplish synapsis in the ph2b genotype, whereas most nuclei completed synapsis in the standard and ph1b genotypes. This result indicates that the Ph2 locus affects synaptic progression. The number of synaptonemal complex (SC) bivalents and of the different SC multivalent associations were determined in each nucleus. The mean number of lateral elements involved in SC multivalent associations (LEm) at mid- zygotene was relatively high and showed similar values in the three genotypes. These values decreased progressively between mid-zygotene and pachytene in the genotypes with the Ph1 locus because of the transformation of multivalents into bivalents. In the ph1b genotype, this value only decreased between late-zygotene and pachytene. Therefore, multivalent correction was more efficient in the presence than in the absence of the Ph1 locus.It is concluded that the Ph1 and Ph2 loci bring about diploidization of allohexaploid wheat via a different mechanism. Received: 31 July 2000 / Accepted: 15 November 2000     

Mutations in a new Streptomyces coelicolor locus which globally block antibiotic biosynthesis but not sporulation

Streptomyces coelicolor produces four known antibiotics. To define genetic elements that regulate antibiotic synthesis, we screened for mutations that visibly blocked synthesis of the two pigmented antibiotics and found that the mutant strains which we recovered were of two classes--double mutants and mutants in which all four antibiotics were blocked. The mutations in these multiply blocked strains define a new locus of S. coelicolor which we have named absA. The genetic location of absA, at 10 o'clock, is distinct from the locations of the antibiotic gene clusters and from other known mutations that affect antibiotic synthesis. The phenotype of the absA mutants suggests that all S. coelicolor antibiotic synthesis genes are subject to a common global regulation that is at least in part distinct from sporulation and that absA is a genetic component of the regulatory mechanism.     

Genes involved in the control of nuclear fusion during the sexual cycle of Saccharomyces cerevisiae

Summary Mutants of Saccharomyces cerevisiae defective for nuclear fusion have been isolated. Their mutations have been characterized by meiotic analysis, dominance-recessivity and complementation. Twelve of the mutations are allelic to the previously described kar 1–1; five affect a second gene designated KAR 2 and three affect a third gene designated KAR 3. There is evidence suggesting that other two mutants are affected in a gene different from the three mentioned.Mutations in KAR 1 and KAR 2 genes are recessive and do not cause obvious effects other than the failure of the karyogamy. Mutations in KAR 3 are semidominant and do cause pleiotropic effects affecting both mitosis and meiosis.