Genetic control of malate dehydrogenase isozymes in maize

At least six nuclear loci are responsible for the genetic control of malate dehydrogenase (L-malate: NAD oxidoreductase; EC 1.1.1.37; MDH) in coleoptiles of maize. Three independently segregating loci (Mdh1, Mdh2, Mdh3) govern the production of MDH isozymes resistant to inactivation by ascorbic acid and found largely or solely in the mitochondria. A rare recessive allele found at a fourth nuclear locus (mmm) causes increased electrophoretic mobility of the MDH isozymes governed by the Mdh1, Mdh2 and Mdh3 loci.—Two loci (Mdh4, Mdh5) govern MDH isozymes that are selectively inactivated by homogenization in an ascorbic acid solution and that appear to be nonmitochondrial (soluble). Mdh4 and Mdh5 segregate independently of each other and independently of Mdh1, Mdh2 and Mdh3. However, there is close linkage between the migration modifier and Mdh4.——Multiple alleles have been found for all of the Mdh loci except the migration modifier, and electrophoretically "null" or near "null" alleles (as expressed in standardized sections of maize coleoptile) have been found for all loci except Mdh4. Duplicate inheritance commonly occurs for Mdh1 and Mdh2 and also for Mdh4 and Mdh5.——Inter- and intragenic heterodimers are formed between sub-units specified by the three loci governing the mitochondrial MDH isozymes. The same is true of the alleles and nonalleles at the two loci governing the soluble variants. No such heterodimers are formed by interactions between mitochondrial and soluble MDH isozymes.     

Linkage relationships of 19 enzyme Loci in maize

Linkage relationships of 19 enzyme loci have been examined. The chromosomal locations of eight of these loci are formally reported for the first time in this paper. These localizations should assist in the construction of additional useful chromosome marker stocks, especially since several of these enzyme loci lie in regions that were previously poorly mapped. Six loci are on the long arm of chromosome 1. The arrangement is (centromere)—Mdh4-mmm-Pgm1-Adh1-Phi-Gdh1, with about 46% recombination between Mdh4 and Gdh1.—Linkage studies with a2 and pr have resulted in the localization of four enzyme genes to chromosome 5 with arrangement Pgm2-Mdh5-Got3-a2-(centromere)-pr-Got2. Pgm2 lies approximately 35 map units distal to a2 in a previously unmapped region of the short arm of 5, beyond ameiotic.—Approximately 23% recombination was observed between Mdh4 and Pgm1 on chromosome 1, while 17% recombination occurred between Mdh5 and Pgm2 on chromosome 5. Similarly, linkages between Idh1 and Mdh1, about 22 map units apart on chromosome 8, and between Mdh2 and Idh2, less than 5 map units apart on chromosome 6, were observed. Thus, segments of chromosomes 1 and 5 and segments of 6 and 8 may represent duplications on nonhomologous chromosomes.     

Chromosomal Location of Two Mitochondrial Malate Dehydrogenase Structural Genes in ZEA MAYS Using Trisomics and B-A Translocations

Maize mitochondrial malate dehydrogenase is coded by four genetic loci, Mdh1, Mdh2, Mdh3 and Mdh4. Two of the four loci have been located on the long arm of chromosome 6, using trisomic analysis and B-A translocations.     

Malate Dehydrogenase, Alcohol Dehydrogenase, and 6-Phosphogluconate Dehydrogenase Isozymes of Zea mays L. × Tripsacum dactyloides L. Hybrids and Parents

Electrophoretic patterns of malate dehydrogenase (Mdh), alcohol dehydrogenase (Adh), and 6-phosphogluconate dehydrogenase (Pgd) of Zea mays L. × Tripsacum dactyloides L. hybrids and their parents were compared. The components of enzymes specific to T. dactyloides may be used as markers to identify the following T. dactyloides chromosomes in the hybrids: Tr 16 (Mdh 2 and Pdg 1), Tr 7, and/or Tr 13 (Adh 2). The isozymes of Mdh 2 are supposed as a possible biochemical marker to evaluate the introgression of genes, determining an apomictic mode of reproduction from T. dactyloides (localized on Tripsacum 16 chromosome) into Z. mays. The isozymes may be used as markers for the identification of maize chromosomes 1 and 6 in the hybrids as well. Chromosome count taken on the examined hybrids showed the addition of 9 to 13 chromosomes of T. dactyloides to maize chromosome complement.     

Gene-Centromere Mapping in Rainbow Trout: High Interference over Long Map Distances

Ten enzyme loci were mapped in relation to their centromeres in gynogenetic diploid rainbow trout. Gene-centromere map distances, calculated under the assumption of complete interference, range from 1.1 cM for Ldh4 to 50 cM for Sod1. The Idh2 and Est1 loci are linked on the same chromosome arm.—The observation of close to 100% heterozygous gynogenetic diploids for the Sod1 and Mdh3,4 loci suggests that near-complete interference occurs on the chromosome arms carrying these loci. The high interference observed in this study and in several other species of fish may be related to the small size of fish chromosome arms.—Comparisons of map locations for the Ldh3 and Ldh4 and the Mdh3 and Mdh4 loci, which were duplicated by a tetraploid event in the evolution of salmonid fish, reveal that they are located at similar distances from their centromeres. Comparative mapping of loci duplicated longer ago shows more variation in map location.—The high proportion of heterozygotes for some loci after gynogenesis involving second polar body retention demonstrates that this is not a practical method for producing homozygous inbred lines in rainbow trout; treatments suppressing the first cell division are more promising for this purpose.     

Association of Isozymes with a Reciprocal Translocation in Cultivated Rye (SECALE CEREALE L.)

In rye (Secale cereale L. cv. "Ailés") the progeny of a cross between a structural heterozygote for a reciprocal translocation (involving the 1R chromosome) and a homozygote for the standard chromosome arrangement were analyzed for the electrophoretic patterns of eight different leaf isozymes and also for their meiotic configuration at metaphase I.——The Got-3 and Mdh-2b loci are linked to each other and also to the reciprocal translocation. The Mdh-2b locus is located in the interstitial segment of the 3Rq chromosome arm, with an estimated distance of 8 cM to the breakpoint. Therefore, the reciprocal translocation involves the 1R and 3R chromosomes.——Also, the Mdh-1 and 6-Pgd-2 loci are linked (16 ± 3 cM) and have been located on the 2Rq arm. Finally, the Per-3 and Per-4 loci are located on the 2Rp chromosome arm at an estimated distance of 26 ± 4 cM.     

Subcellular localization of isozymes of NAD-dependent malate dehydrogenase in sugar beet <Emphasis Type="Italic">Beta vulgaris</Emphasis> L.

Subcellular localization of isozymes of NAD-dependent malate dehydrogenase (MDH) in sugar beet was studied. Isozymes ss and ll controlled by loci Mdh2 and Mdh3, respectively, were shown to locate in mitochondria, whereas isozyme pp controlled by locus Mdh1, in microbodies. All examined samples lack hybrid MDH isozymes, which could testify to the interaction between products of nonallelic Mdh genes. This can be explained by the localization of nonallelic isozymes in various compartments of the cell and organelles.     

Genetic Control and Linkage Relationships among Aminopeptidases in Maize

Maize aminopeptidase is coded by four genes. Amp1 and Amp2 have been localized to chromosome 1. A three-point cross shows the gene order to be Amp2—15%—Amp1—33%—Adh2 (alcohol dehydrogenase). Amp3 and Amp4 assort independently of each other and of chromosome 1 aminopeptidases. Another linkage relationship among the maize genes Amy2 (amylase), Cat1 (catalase), and Amp3 exists, but the chromosome location has yet to be established unequivocally.     

Isozyme Analysis in a Genetic Collection of Amaranths (<Emphasis Type="Italic">Amaranthus</Emphasis> L.)

In various populations of the cultivated and weedy amaranth species, the electrophoretic patterns of alcohol dehydrogenase (ADH), glutamate dehydrogenase (GDH), malate dehydrogenase (MDH), isocitrate dehydrogenase (IDH) and malic enzyme (Me) were studied. In total, 52 populations and two varieties (Cherginskii and Valentina) have been examined. Allozyme variation of this material was low. Irrespective of species affiliation, 26 populations and two varieties were monomorphic for five enzymes; a slight polymorphism of three, two, and one enzymes was revealed in three, nine, and fourteen populations, respectively. A single amaranth locus, Adh, with two alleles, Adh F and Adh S, controls amaranth ADH. Two alleles, common Gdh S and rare Gdh F, control GDH; no heterozygotes at this locus were found. The MDH pattern has two, the fast- and slow-migrating, zones of activity (I and II, respectively). Under the given electrophoresis conditions, the fast zone is diffuse, whereas slow zone is controlled by two nonallelic genes, monomorphic Mdh 1 and polymorphic Mdh 2 that includes three alleles: Mdh 2-F, Mdh 2-N, and Mdh 2-S. Low polymorphism of IDH and Me was also found, though their genetic control remains unknown.     

Malate Dehydrogenase Isozymes (MDH; EC 1.1.1.37) in Long-Term Callus Culture of Cereus peruvianus (Cactaceae) Exposed to Sugar and Temperature Stress

Malate dehydrogenase (MDH; EC 1.1.1.37) isozymes in long-term callus tissue culture of Cereus peruvianus were studied in starch gel electrophoresis to investigate the control of differential Mdh gene expression under sugar and temperature stress. While two cytosol MDH isozymes showed an unchanged phenotype when the callus tissues were transferred to medium maintained at 22 or 37°C and containing different concentrations of sucrose, glucose, and fructose, the different combinations of five mitochondrial MDH (mtMDH) and two microbody MDH (mbMDH) showed different MDH isozyme patterns in the callus populations. Differential expression of mtMDH isozymes seems to be modulated at the posttranslational level in callus tissues exposed to different concentrations and types of sugar and to high-temperature and low-temperature stress. An inductor effect on the expression of mbMDH isozymes was observed under stress conditions and in long-term callus tissue, and they may also present different responses.     

New isozyme systems for maize (Zea mays L.): Aconitate hydratase,adenylate kinase,NADH dehydrogenase,and shikimate dehydrogenase

Electrophoretic variation and inheritance of four novel enzyme systems were studied in maize (Zea mays L.). A minimum of 10 genetic loci collectively encodes isozymes of aconitate hydratase (ACO; EC 4.2.1.3.), adenylate kinase (ADK; EC 2.7.4.3), NADH dehydrogenase (DIA; EC 1.6.99.—), and shikimate dehydrogenase (SAD; EC 1.1.1.25). At least four loci are responsible for the genetic control of ACO. Genetic data for two of the encoding loci,Aco1 andAco4, demonstrated that at least two maize ACOs are active as monomers. Analysis of organellar preparations suggests that ACO1 and ACO4 are localized in the cytosolic and mitochondrial subcellular fractions, respectively. Maize ADK is encoded by a single nuclear locus,Adk1, governing monomeric enzymes that are located in the chloroplasts. Two cytosolic and two mitochondrial forms of DIA were electrophoretically resolved. Segregation analyses demonstrated that the two cytosolic isozymes are controlled by separate loci,Dia1 andDia2, coding for products that are functional as monomers (DIA1) and dimers (DIA2). The major isozyme of SAD is apparently cytosolic, although an additional faintly staining plastid form may be present. Alleles atSad1 are each associated with two bands that cosegregate in controlled crosses. Linkage analyses and crosses with B-A translocation stocks were effective in determining the map locations of six loci, including the previously described but unmapped locusAcp4. Several of these loci were localized to sparsely mapped regions of the genome.Dia2 andAcp4 were placed on the distal portion of the long arm of chromosome 1, 12.6 map units apart.Dia1 was localized to chromosome 2, 22.2 centimorgans (cM) fromB1. Aco1 was mapped to chromosome 4, 6.2 cM fromsu1. Adk1 was placed on the poorly marked short arm of chromosome 6, 8.1 map units fromrgd1. Less than 1% recombination was observed betweenGlu1 (on chromosome 10) andSad1. In contrast to many other maize isozyme systems, there was little evidence of gene duplication or of parallel linkage relationships for these allozyme loci. This work was supported by grants from Pioneer Hi-Bred International, Inc., of Johnston, Iowa, the National Institute of Health (Research Grant GM11546), and the United States Department of Agriculture (Competitive Research Grant 83-CRCR-1-1273). This is Paper No. 11372 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh.     

Chromosome polymorphism of teosinte

Florida teosinte from Honduras and Guanajuato teosinte have most of their knobs internally located and a common inversion in the short arm of chromosome 8. Teosintes from northern Guatemala, Huixta and Monajil, have all their knobs terminally located and do not have any inversions. Therefore, Florida teosinte from Honduras appears to be phylogenetically closer to Guanajuato teosinte than it is to Guatemalan teosintes.——The presence of B chromosomes and an In 8 in Guanajuato teosinte and Florida teosinte from Honduras may constitute evidence of introgression between maize and teosinte.     

Temperature-Sensitive Lethal Mutations on Yeast Chromosome I Appear to Define Only a Small Number of Genes

A method was developed for isolating large numbers of mutations on chromosome I of the yeast Saccharomyces cerevisiae. A strain monosomic for chromosome I (i.e., haploid for chromosome I and diploid for all other chromosomes) was mutagenized with either ethyl methanesulfonate or N-methyl-N'-nitro-N -nitrosoguanidine and screened for temperature-sensitive (Ts^- ) mutants capable of growth on rich, glucose-containing medium at 25° but not at 37°. Recessive mutations induced on chromosome I are expressed, whereas those on the diploid chromosomes are usually not expressed because of the presence of wild-type alleles on the homologous chromosomes. Dominant ts mutations on all chromosomes should also be expressed, but these appeared rarely. — Of the 41 ts mutations analyzed, 32 mapped on chromosome I. These 32 mutations fell into only three complementation groups, which proved to be the previously described genes CDC15, CDC24 and PYK1 (or CDC19). We recovered 16 or 17 independent mutations in CDC15, 12 independent mutations in CDC24 and three independent mutations in PYK1. A fourth gene on chromosome I, MAK16, is known to be capable of giving rise to a ts-lethal allele, but we recovered no mutations in this gene. The remaining nine mutations isolated using the monosomic strain appeared not to map on chromosome I and were apparently expressed in the original mutants because they had become homozygous or hemizygous by mitotic recombination or chromosome loss. — The available information about the size of chromosome I suggests that it should contain approximately 60–100 genes. However, our isolation in the monosomic strain of multiple, independent alleles of just three genes suggests that only a small proportion of the genes on chromosome I is easily mutable to give a Ts^--lethal phenotype. — During these studies, we located CDC24 on chromosome I and determined that it is centromere distal to PYK1 on the left arm of the chromosome.     

Two Chromosomal Genes Required for Killing Expression in Killer Strains of SACCHAROMYCES CEREVISIAE

The killer character of yeast is determined by a 1.4 x 10⁶ molecular weight double-stranded RNA plasmid and at least 12 chromosomal genes. Wild-type strains of yeast that carry this plasmid (killers) secrete a toxin which is lethal only to strains not carrying this plasmid (sensitives). ——— We have isolated 28 independent recessive chromosomal mutants of a killer strain that have lost the ability to secrete an active toxin but remain resistant to the effects of the toxin and continue to carry the complete cytoplasmic killer genome. These mutants define two complementation groups, kex1 and kex2. Kex1 is located on chromosome VII between ade5 and lys5. Kex2 is located on chromosome XIV, but it does not show meiotic linkage to any gene previously located on this chromosome. ——— When the killer plasmid of kex1 or kex2 strains is eliminated by curing with heat or cycloheximide, the strains become sensitive to killing. The mutant phenotype reappears among the meiotic segregants in a cross with a normal killer. Thus, the kex phenotype does not require an alteration of the killer plasmid. ——— Kex1 and kex2 strains each contain near-normal levels of the 1.4 x 10⁶ molecular weight double-stranded RNA, whose presence is correlated with the presence of the killer genome.     

Purification and biochemical properties of genetically defined malate dehydrogenase in maize

Malate dehydrogenase of maize exists in multiple molecular forms (isozymes). In strain W64A, two soluble forms (s-MDH), five mitochondrial forms (m-MDH), and two glyoxysomal forms (g-MDH) were found in etiolated seedlings. The s-MDHs and m-MDHs were prepared in highly purified form. Using these purified isozymes, experiments with reducing agents (100 mm mercaptoethanol), low pH (2.0), and high salt cocn (7.5 m guanidine-HCl), along with genetic data, have eliminated the possibility of conformational alterations as an explanation for MDH multiplicity in maize; the MDH isozymes are genetically determined. Biochemical properties for each of the seven MDH isozymes were examined. Molecular weight, pI, pH optimum, thermolability, and K_m for oxaloacetate, malate, NAD, and NADH at different pH values were determined for each isozyme. Different kinetics of substrate inhibition (oxaloacetate) and coenzyme inhibition (NAD) were observed for the different isozymes. Effects of NAD analogs, chelating agents, reducing agents, metal ions, and TCA cycle acids on the enzymatic activity of these isozymes were tested. Based on the physical and kinetic properties observed, the maize malate dehydrogenase isozymes can be classified into four groups: s-MDH¹; s-MDH²; the two most anodal m-MDHs; and the three most cathodal m-MDHs. Since strain W64A is highly inbred, our data along with our previous and simultaneous genetic analysis suggest that multiple genes are involved in the expression of maize malate dehydrogenase isozymes.     

Developmental genetics of mutants that specify knotted leaves in maize

Of seven dominant knotted-leaf mutants tested, six mapped at or near Kn1 on the long arm of chromosome 1, and one was not linked to Kn1. Comparisons of phenotypes among these mutants allowed us to focus on a systematic abnormality: the parenchyma cells associated with lateral veins do not fully differentiate into bundle sheath, mesophyll or upper sclerenchyma. The more dramatic expression of Kn1 mutants—knots, ligule alterations and twisting—is sporadic and dependent on the time when the mutant acts in leaf primordium development. Using lw to mark leaf sectors that lose Kn1 following X-irradiation, we show that the knotted-leaf phenotype encoded by chromosome 1L is autonomous. Analysis of sectors lacking a particular Kn1 gene ( Kn1-N2) suggests that Kn1 itself, rather than a linked modifier gene, is autonomous in the leaf primordium. Aneuploid studies using various translocations involving 1L and marked by Adh1 allozymes are compared. The Kn1 mutant appears to encode a "new" function or a considerable overproduction of an extant product in the leaf. Kn1/- 1L hypoploids either express knotted poorly or not at all; transvection is ruled out, but the cause for this modification of Kn1 expression is not yet known.—Our working hypothesis is that Kn1 mutants permit the expression of a product that is usually not produced in leaf primordial cells. We suggest that this product interferes with the early cell-type commitments of cells near lateral veins. Thus, relatively uncommitted cells are present in more mature blades, where they may divide unexpectedly into knots or may induce bits of ligule.     

The cytogenetic localization of the alcohol dehydrogenase-1 locus in maize

The alcohol dehydrogenase-1 (Adh) locus in maize has been positioned relative to thirteen reciprocal translocations that have breakpoints in the long arm of chromosome 1(1L). The methods of Gopinath and Burnham (1956) to produce interstitial segmental trisomy with overlapping translocations and of Rakha and Robertson (1970) to produce compound B-A translocations were coupled with the co-dominant nature of the ADH isozymes to allow the cytological placement. The results of several crosses are consistent with Adh being in the region of 0.80-0.90 of 1L.--The duplication that results from the overlap of translocations 1-3(5267) and 1-3(5242) and that includes Adh was studied with respect to meiotic segregation and pollen transmission. When heterozygous with normal chromosomes, a low level of recombination within the duplicated regions is detectable and the duplication and normals are recovered with equal frequencies through the female. In the pollen, the hyperploid grains cannot compete equally with the euploids in achieving fertilization.--The use of co-dominant heteromultimeric isozymes as genetic markers for the development of a series of interstitial segmental trisomics in maize is discussed.     

A study of enzyme activities in a dosage series of the long arm of chromosome one in maize

The enzyme activity levels of alcohol, malate, isocitrate, glucose-6-phosphate and 6-phosphogluconate dehydrogenases were determined in mature maize scutella in a series of one to four doses of the long arm of chromosome 1, produced by the B-A translocation 1La. Although the Adh structural locus was varied, ADH levels did not exhibit a gene-dosage effect. The levels of G6PDH, 6PGDH and IDH were negatively correlated with the dosage of 1L. MDH was unresponsive. The esterase-8 enzyme, whose structural locus was demonstrated to be elsewhere in the genome, was also negatively correlated with 1L dosage. The portion of the B chromosome involved in the translocation was shown to have no effect on the enzyme levels. Measurements of cell size and hydrolysable DNA per mg dry weight revealed no change in the number of cells through the one, two and three dose series. The topic of enzyme alterations in aneuploids is reviewed.     

A Chromosomal Translocation Causing Overproduction of Iso-2-Cytochrome c in Yeast

The CYC7–1 mutation in the yeast Saccharomyces cerevisiae causes the production of approximately 30 times the normal amount of iso-2-cytochrome c. Genetic analysis established that the CYC7–1 mutation is a reciprocal translocation involving the left arm of chromosome V and the right arm of chromosome XVI. The chromosome V arm was broken adjacent to the gene CYC7, which determines the primary structure of iso-2-cytochrome c, and this fragment containing the CYC7 gene was joined to the segment of chromosome XVI. It appears as though the elevation of iso-2-cytochrome c is caused by an abnormal controlling region adjacent to the structural region of the CYC7 gene.