Phenotypes Of Double Mutants Research Articles

Phenotypic and Genetic Analysis of det2, a New Mutant That Affects Light-Regulated Seedling Development in Arabidopsis.

The greening phenotypes produced by recessive mutations in a gene designated de-etiolated-2 (DET2) are described. Recessive mutations in the DET2 gene uncouple light signals from a number of light-dependent processes. det2 mutations result in dark-grown Arabidopsis thaliana seedlings with many characteristics of light-grown plants, including hypocotyl growth inhibition, cotyledon expansion, primary leaf initiation, anthocyanin accumulation, and derepression of light-regulated gene expression. In contrast to these morphological and gene expression changes, however, the chloroplast development program is not initiated in the dark in det2 mutants, suggesting that light-regulated gene expression precedes the differentiation of etioplasts to chloroplasts. det2 mutations thus reveal at least two classes of downstream light-regulated responses that differ in their timing and control mechanisms. Homozygous det2 mutations also affect photoperiodic responses in light-grown plants, including timing of flowering, dark adaptation of gene expression, and onset of leaf senescence. The phenotype of det1 det2 double mutants is additive, implying that DET1 and DET2 function in distinct pathways that affect downstream light-regulated genes. Furthermore, these pathways are not utilized solely during early seedling development but must also be required to regulate different aspects of the light developmental program during later stages of vegetative growth.

Implications of Double Mutant Phenotypesa

Annals of the New York Academy of SciencesVolume 605, Issue 1 p. 215-229 Implications of Double Mutant Phenotypesa SUSAN BILLINGS-GAGLIARDI, SUSAN BILLINGS-GAGLIARDI Department of Cell Biology University of Massachusetts Medical School Worcester, Massachusetts 01655Search for more papers by this authorMERRILL K. WOLF, MERRILL K. WOLF Department of Cell Biology University of Massachusetts Medical School Worcester, Massachusetts 01655Search for more papers by this author SUSAN BILLINGS-GAGLIARDI, SUSAN BILLINGS-GAGLIARDI Department of Cell Biology University of Massachusetts Medical School Worcester, Massachusetts 01655Search for more papers by this authorMERRILL K. WOLF, MERRILL K. WOLF Department of Cell Biology University of Massachusetts Medical School Worcester, Massachusetts 01655Search for more papers by this author First published: November 1990 https://doi.org/10.1111/j.1749-6632.1990.tb42394.xCitations: 3 a This work was supported by NIH Grant #NS 11425 (Javits Award). b In the interest of legibility we have simplified formal genetic notation as follows: shimld to mld and jpmsd to msd. AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume605, Issue1Myelination and DysmyelinationNovember 1990Pages 215-229 RelatedInformation

Induction of levansucrase in Bacillus subtilis: an antitermination mechanism negatively controlled by the phosphotransferase system

The target of the induction by sucrose of the levansucrase gene is a transcription terminator (sacRt) located upstream from the coding sequence, sacB. The two-gene locus sacX-sacY (formerly sacS) and the ptsI gene were previously shown to be involved in this induction. ptsI encodes enzyme I of the phosphoenolpyruvate-dependent phosphotransferase system. SacX is strongly homologous to sucrose-specific phosphotransferase system-dependent permeases. SacY is a positive regulator of sacB. Here we show that SacY is probably an antiterminator interacting directly with sacRt, since in Escherichia coli the presence of the sacY gene stimulates the expression of a reporter gene fused downstream from sacRt. Missense mutations affecting sacY were sequenced, and the sacB regulation was studied in isogenic strains carrying these mutations or in vitro-generated mutations affecting sacX, sacY, or ptsI. The phenotype of double mutants suggests a model in which SacX might be a sucrose sensor that would be phosphorylated by the phosphotransferase system and, in this state, could inhibit the SacY antiterminator. Exogenous sucrose, or a mutation inactivating the phosphotransferase system, would dephosphorylate SacX and allow antitermination at sacRt.

Construction and characterization of an Azotobacter vinelandii strain with mutations in the genes encoding flavodoxin and ferredoxin I.

Flavodoxin and ferredoxin I have both been implicated as components of the electron transport chain to nitrogenase in the aerobic bacterium Azotobacter vinelandii. Recently, the genes encoding flavodoxin (nifF) and ferredoxin I (fdxA) were cloned and sequenced and mutants were constructed which are unable to synthesize either flavodoxin (DJ130) or ferredoxin I (LM100). Both single mutants grow at wild-type rates under N2-fixing conditions. Here we report the construction of a double mutant (DJ138) which does not synthesize either flavodoxin or ferredoxin I. When plated on ammonium-containing medium, this mutant had a very small colony size when compared with the wild type, and in liquid culture with ammonium, this double mutant grew three times slower than the wild type or single mutant strains. This demonstrated that there is an important metabolic function unrelated to nitrogen fixation that is normally carried out by either flavodoxin or ferredoxin. If either one of these proteins is missing, the other can substitute for it. The double mutant phenotype can now be used to screen site-directed mutant versions of ferredoxin I for functionality in vivo even though the specific function of ferredoxin I is still unknown. The double mutant grew at the same slow rate under N2-fixing conditions. Thus, A. vinelandii continues to fix N2 even when both flavodoxin and ferredoxin I are missing, which suggests that a third as yet unidentified protein also serves as an electron donor to nitrogenase.

Terminal versus segmental development in the Drosophila embryo: the role of the homeotic gene fork head.

Mutations of the homeotic gene fork head (fkh) of Drosophila transform the non-segmented terminal regions of the embryonic ectoderm into segmental derivatives: Pre-oral head structures and the foregut are replaced by post-oral head structures which are occasionally associated with thoracic structures. Posterior tail structures including the hindgut and the Malpighian tubules are replaced by post-oral head structures associated with anterior tail structures. The fkh gene shows no maternal effect and is required only during embryogenesis. The phenotypes of double mutants indicate that fkh acts independently of other homeotic genes (ANT-C, BX-C, spalt) and caudal. In addition, the fkh domains are not expanded in Polycomb (Pc) group mutant embryos. Ectopic expression of the homeotic selector genes of the ANT-C and BX-C in Pc group mutant embryos causes segmental transformations in terminal regions of the embryo only in the absence of fkh gene activity. Thus, fkh is a region-specific homeotic rather than a selector gene, which promotes terminal as opposed to segmental development.

Head and tail development of the Drosophila embryo involves spalt, a novel homeotic gene

Mutations in spalt (sal), a novel homeotic gene on the second chromosome of Drosophila, cause opposite transformations in two subterminal regions of the embryo: posterior head segments are transformed into anterior thoracic structures and anterior tail segments are transformed into posterior abdominal structures. The embryonic phenotypes of double mutants for sal and various Antennapedia (ANT-C) or bithorax (BX-C) genes indicate that sal acts independently of the hierarchical order of the latter gene complexes. Trans-regulatory gene mutations causing ectopic expression of ANT-C and BX-C genes do not change the realms of sal action. It is proposed that the region-specific action of the sal gene primarily promotes head as opposed to trunk development, while the BX-C gene AbdB distinguishes tail from head.

Interactions between janus and bcd cortical pattern mutants in Tetrahymena thermophila

An analysis of bcd, janA; bcd, janB; and bcd, janC double-mutant phenotypes in Tetrahymena thermophila has allowed us to examine patterning processes affected by two different classes of mutations. bcd brings about a broadening of the oral and contractile vacuole pore domains in the ciliate cortex, while the janus mutations generate a mirror-image duplication of the ventral cortical pattern. We observed both bcd and janus characteristics expressed in the double mutants, as well as features unique to the double-mutant. Temperature-shift experiments employing the temperature-sensitive janB mutation in a double-mutant (bcd, janB) combination allowed us to observe the changes in pattern as a mirror-image geometry was brought into expression and subsequently removed within the bcd, janB double homozygote. These experiments suggest that there are multiple pattern-mechanisms at work with differing kinetics of expression in the ciliate cortex. We discuss how the bcd mutation could influance expression of the janus mutations in light of a model previously proposed to account for the janus phenotype.

Genetic studies of mutations at two loci of Drosophila melanogaster which cause a wide variety of homeotic transformations.

The ash-1 locus is in the proximal region of the left arm of the third chromosome of Drosophila melanogaster and the ash-2 locus is in the distal region of the right arm of the third chromosome. Mutations at either locus can cause homeotic transformations of the antenna to leg, proboscis to leg and/or antenna, dorsal prothorax to wing, first and third leg to second leg, haltere to wing, and genitalia to leg and/or antenna. Mutations at the ash-1 locus cause, in addition, transformations of the posterior wing and second leg to anterior wing and second leg, respectively. A similar spectrum of transformations is caused by mutations at yet another third chromosome locus, trithorax. One extraordinary aspect of mutations at all three of these loci is that they cause such a wide variety of transformations. For mutations at both of the loci that we have studied the expression of the homeotic phenotype is both disc-autonomous (as shown by injecting mutant discs into metamorphosing larvae) and cell autonomous (as shown by somatic recombination analysis). The original mutations which identified these two loci, although lethal, manifest variable expressivity and incomplete penetrance of the homeotic phenotype suggesting that they are hypomorphic. The phenotype of double mutants which were synthesized by combining different pairs of those original mutations manifest for two of the four pairs a greater degree of expressivity and slightly more penetrance of the homeotic transformations. This mutual enhancement suggests that the products of both loci interact in the same process. A third double mutant expresses a discless phenotype.Additional alleles have been recovered at both the ash-1 and the ash-2 loci. Some of these alleles as homozygotes or transheterozygotes express the wide range of transformations revealed first by double mutants. One of the alleles at the ash-1 locus when homozygous and several transheterozygous pairs can cause either the homeotic transformation of discs or the absence of those discs. The fact that these two defects, absence of specific discs and homeotic transformations of those same discs can be caused by mutations within a single gene suggests that the activity of the product of this gene is essential for normal imaginal disc cell proliferation. Loss of that activity leads to the absence of discs, whereas, reduction of that activity leads to homeotic transformations.

Isolation and characterization of Escherichia coli pantothenate permease (panF) mutants.

Mutants of Escherichia coli K-12 defective in the pantothenate permease (panF) were isolated and characterized. The panF mutation resulted in the complete loss of pantothenate uptake and of the ability to use extracellular vitamin for growth. The growth phenotypes of panF panD, panF panB, and panF panC double mutants showed that the cytoplasmic membrane was impermeable to external pantothenate. Analysis of the intracellular and extracellular metabolites from strain DV1 (panF panD) labeled with beta-[3-3H]alanine demonstrated that a carrier-mediated mechanism for efficient pantothenate efflux remained in the panF mutant. Genetic mapping of this nonselectable allele was facilitated by the isolation of three independent Tn10 insertions close to panF. Two- and three-factor crosses located panF at minute 72 of the E. coli chromosome and established the gene order fabE panF aroE.

Establishment of dorsal-ventral polarity in the Drosophila embryo: Genetic studies on the role of the Toll gene product

Within the group of maternal effect genes necessary for the establishment of the dorsal-ventral pattern of the Drosophila embryo, the Toll gene mutates to give a singular variety of embryonic phenotypes. Lack of function alleles produce dorsalized embryos as a recessive maternal effect. Dominant gain of function alleles result in ventralized embryos. Other recessive alleles cause partial dorsalization or lateralization of the embryonic pattern. Gene dosage studies indicate that the dominant ventralized phenotype results from an altered activity of the Toll product. Complementation studies show specific trans interactions between copies of the Toll product. Double mutant phenotypes suggest that the products of several other dorsal-group genes regulate the activity of Toll.

Genetic Recombination in the Lignin-Degrading Basidiomycete Phanerochaete chrysosporium

Heterokaryons made from auxotrophic strains of the lignin-degrading basidiomycete Phanerochaete chrysosporium were induced to fruit. The isolation of wild-type and double-mutant phenotypes from these crosses indicated that genetic recombination had occurred. Cytological studies demonstrated that more than 90% of the basidiospores from the wild-type and auxotrophic strains and from forced heterokaryons were binucleate. Colonies of the wild-type strain of P. chrysosporium arising from single, predominantly uninucleate conidia were all capable of producing fruit bodies and basidiospores.

Dependency relations between events in mitosis in Schizosaccharomyces pombe.

ABSTRACT The dependency relationships between various events in mitosis in the fission yeast Schizo-eaccharomycee pombe have been investigated using a combination of approaches. The events concerned are those controlled by specific genes, a step or steps in mitosis sensitive to benomyl, and protein synthesis. The anti-microtubule agent benomyl was shown to inhibit mitosis specifically, while not significantly affecting the accumulation of RNA or protein. DNA synthesis was not directly affected, though accumulation ceased after mitosis was inhibited. The inhibition of mitosis was readily reversible and was followed by a synchronous cell division. Reciprocal shift experiments were carried out using benomyl in conjunction with temperature-sensitive mutants defective in mitosis. Consideration of these experiments and the transition points of the mutants allowed the mutant and the steps controlled by the respective genes to be ordered functionally. Group (i) consisted of cdc6 23, which showed an early transition point: the benomyl-sensitive step was dependent on completion of the cdc6 step. Group (ii) consisted of cdc2·33 and cdc2·33 K3, which had transition points close to the time of mitosis itself. The benomyl-sensitive step was dependent on the cdc2 and cdc27 steps, but the relationship between these gene-controlled steps was uncertain. Group (ii) processes may be dependent on those in group (i), or they may act independently of one another on separate pathways. Group (iii) mutants consist of 1·7 cdc13·117 and cdc25·22. These have transition points close to mitosis, and act interdependently with benomyl. It is likely, though not directly proven, that all group (iii) processes are dependent on those in group (ii). The strikingly different terminal phenotype of cdc13·17 compared with other mutants of mitosis allowed further analysis of the group (iii) mutants according to the terminal phenotypes of double mutants of cd13·117 with other mutants. Double mutants with cdc1·7, cdc2·33, 25·22 and cdc27·1<3 all showed a terminal phenotype unlike cdc13·117, and similar to the other single mutant parent. It is therefore likely that the cdc13 step is dependent on cdci, 2, 25 and 27; that is, later in mitosis, consistent with its unusual terminal phenotype. Two mutants defective in stages of the cell cycle other than mitosis were subjected to reciprocal shifts with benomyl. The process controlled by cdc15·140, an event in septum formation, was dependent on the benomyl-sensitive step. The other mutant, cdc10·129, with a defect very early in the cycle, has been proposed to arrest at a stage analogous to ‘start’ in budding yeast. The completion of the cdcio function was shown to be dependent on the occurrence of mitosis in the previous cycle. Cells arrested early in mitosis by cdc 2·33 were able to undergo mitosis when returned to the permissive temperature in the presence of cycloheximide. This supports previous observations that protein synthesis is not required for mitosis once the process has started. Extrapolation of this result to the normal cycle suggests that sequential switching on of cdc genes is not the basis of the dependency relations determined in this study, and that periodic synthesis of proteins coded by cdc genes may not be an important part of the mechanism that controls the sequence of events in mitosis.

Determining the phoM map location in Escherichia coli K-12 by using a nearby transposon Tn10 insertion.

A phoR strain was constructed with transposon Tn10 inserted near the phoM+ locus. This was done without any prior knowledge of the phoM map location. Subsequently, we defined the phoM map position by screening tetracycline-sensitive (Tcs) derivatives for mutants which were both alkaline phosphatase negative (ther phoR phoM double mutant phenotype) and auxotrophic simultaneously. Some of these mutants were Thr-. Bacteriophage P1-mediated transductions were used to confirm that phoM and its nearby Tn10 insertion were closely linked to thr. Unexpectedly, 7 of 10 mutants analyzed also had mutations unlinked to the phoM-thr-Tn10 region. These may represent a new type of Tn10-promoted molecular event which is caused by transposition of a Tn10 end (IS10).

Pigmentation and lysosome function in mice homozygous for both pale ear and beige-J pigment genes.

SUMMARYWe have examined mice doubly homozygous for both pale ear (ep/ep) and beige (bgJ/bgJ) mutations in order to detect genetic interactions between these 2 loci affecting pigmentation and lysosome physiology. The doubly homozygous mouse has a new pigmentation phenotype consistent with independent effects ofepandbg. The beige (Brandt, Elliott & Swank, 1975) and pale ear (Novak & Swank, 1979)genes have abnormal kidney lysosomal enzyme accumulation caused by defective secretion into urine. No cumulative effect on these functions was observed in the new double mutant phenotype. The new phenotype has giant lysosomes typical of the beige mutation. Unexpectedly, the beige gene corrects the effect of the pale ear on serum lysosomal enzyme concentration. There is also a gene dosage effect of the beige gene on this serum lysosomal enzyme phenotype. The results suggest that the beige and pale ear genes affect the same pathway(s) of lysosome biosynthesis and/or processing. The action of the beige gene may precede that of the pale ear gene in lysosome physiology.

Phenotypes of Double Conidiation Mutants of Aspergillus nidulans

A series of strains, doubly mutant at conidiation loci, have been made. The phenotypes of these strains reflected the epistasy of earlier blocking mutants over later ones and confirmed the order of gene sequence predicted from the phenotypes of single mutants. Oligosporogenous mutants gave complex interactions, especially between brl and med mutants. These results indicated that (i) gene action overlapped in time, (ii) several parts of the conidial apparatus were interchangeable and (iii) nuclei leaving the vesicle were not irreversibly programmed. Structures produced by mutants were reminiscent of the conidial apparatus of other Aspergillus species and of related genera.

Construction and phenotypes of double sporulation deficient mutants in Streptomyces coelicolor A3(2).

In whiA, B, G and H mutants of Streptomyces coelicolor A3(2), aerial hyphae develop but sporulation speta are not formed. Septa are produced by whiI mutants but are spaced abnormally far apart. Mutants in each locus have a distinctive aerial mycelium morphology, except for whiA and B mutants which are closely similar. Seven strains were made with pairwise combinations of whiA and B mutations with whiG, H and I mutations and with each other. The genotypes of these strains were confirmed by suitable crosses and their aerial mycelium morphology examined. An indirect procedure was used to determine the aerial mycelium morphology of whiGH, GI and HI double mutants. The double mutants always closely resembled one of the single mutant parent strains in morphology and a consistent scheme of epistasis was obtained--whiG being epistatic to whiH, A, B and I; whiH to whiA, B and I; and whiA or B to whiI. These results point to the absence of any complex interactions between gene products, which might have been revealed by the occurrence of novel phenotypes in double mutants or by inconsistencies in the epistasis scheme.

Carotenoids of wild-type and mutant strains of tetTetranychus pacificus McGregor (Acari: Tetranychidae)

1. 1. The carotenoids of wild-type and of eight pigment mutants of the spider mite Tetranychus pacificus were investigated. 2. 2. Besides the plant pigments α-carotene, β-carotene, lutein, lutein-5,6-epoxide, violaxanthin and neoxanthin, wild-type T. pacificus was found to contain esterified 3-hydroxy-4-keto-β-carotene, 3-hydroxy-4,4't'-diketo-β-carotene and astaxanthin. 3. 3. In four mutants only unaltered plant pigments were found. Two other mutants showed an almost complete lack of astaxanthin. The two remaining mutants possessed the same hydroxy-keto-carotenoids as wild-type, but these were not or only partly esterified. 4. 4. From two-factor crosses between the different mutants, the double-mutant phenotypes were inferred. 5. 5. The possible nature of the pigment mutations is discussed with regard to the results of the pigment analyses and the genetic data.

Genetic Analysis of the First Steps of Sulphate Metabolism in Aspergillus nidulans

PHENOTYPIC analysis of interactions between different mutations in the same genome often helps elucidate the structure and function of the loci in question. If, in a series of mutations affecting a linear pathway, each non-allelic mutation has a characteristic phenotype, then the order of the steps they affect can be determined from the phenotypes of double mutants. A (“non-leaky”) mutation blocking one step in the pathway will be epistatic to all those blocking later steps. The minimum number of different double mutants sufficient to determine unequivocally the order of loci participation on a linear pathway equals the number of different loci (the total number of different double mutants equals n(n−l)/2 where n equals the number of different loci). This communication reports application of this method in Aspergillus nidulans to four loci where mutation can result in loss of ability to use sulphate but not sulphite as a sulphur source and indicates that these four loci are involved in the conversion of sulphate to a metabolite which interferes with utilization of exogenous choline O-sulphate.