Mutant

About: Mutant is a research topic. Over the lifetime, 74520 publications have been published within this topic receiving 3477079 citations.

Alterations in Growth, Photosynthesis, and Respiration in a Starchless Mutant of Arabidopsis thaliana (L.) Deficient in Chloroplast Phosphoglucomutase Activity

Abstract: A mutant of Arabidopsis thaliana (L.) Heynh. which lacks leaf starch was isolated by screening for plants which did not stain with iodine. The starchless phenotype, confirmed by quantitative enzymic analysis, is caused by a single recessive nuclear mutation which results in a deficiency of the chloroplast isozyme of phosphoglucomutase. When grown in a 12-h photoperiod, leaves of the wild-type accumulated substantial amounts of starch but lower levels of soluble sugars. Under these conditions, the mutant accumulated relatively high levels of soluble sugars. Rates of growth and net photosynthesis of the mutant and wild-type were indistinguishable when the plants were grown in constant illumination. However, in a short photoperiod, the growth of the mutant was severely impaired, the rate of photosynthesis was depressed relative to the wild-type, and the rate of dark respiration, which was high following the onset of darkness, exhibited an uncharacteristic decay throughout the dark period. The altered control of respiration by the mutant, which may be related to the relatively high levels of soluble carbohydrate that accumulate in the leaf and stem tissue, is believed to be partially responsible for the low growth rate of the mutant in short days. The depressed photosynthetic capacity of the mutant may also reflect a metabolic adaptation to the accumulation of high levels of soluble carbohydrate which mimics the effects of alterations in source/sink ratio. The activities of sucrose phosphate synthase and acid invertase are significantly higher in the mutant than in the wild-type whereas ADP-glucose pyrophosphorylase activity is lower. This suggests that the activities of these enzymes may be modulated in response to metabolite concentrations or flux through the pathways.

hCTR1: A human gene for copper uptake identified by complementation in yeast

Abstract: The molecular mechanisms responsible for the cellular uptake of copper in mammalian cells are unknown. We describe isolation of a human gene involved in this process by complementation of the yeast high-affinity copper uptake mutant, ctr1. Besides complementing ctr1 growth defect on nonfermentable media, the human gene also rescues iron transport and SOD1 defects in ctr1 yeast. Overexpression of the gene in yeast leads to vulnerability to the toxicity of copper overload. In addition, its expression in ctr1 yeast significantly increases the level of cellular copper, as demonstrated by atomic absorption. We propose this gene as a candidate for high-affinity copper uptake in humans and by analogy have named it hCTR1. The hCTR1 and yeast CTR1 predicted transmembrane proteins are 29% identical, but the human protein is substantially smaller in both the extracellular metal-binding and intracellular domains. An additional human gene similar to hCTR1, here named hCTR2, was identified in a database search. Both hCTR1 and hCTR2 are expressed in all human tissues examined, and both genes are located in 9q31/32. These studies, together with the previously recognized functional and sequence similarity between the Menkes/Wilson copper export proteins and CCC2 in yeast, demonstrate that similar copper homeostatic mechanisms are used in these evolutionarily divergent organisms.

Enhanced Rice Blast Resistance by CRISPR/Cas9-Targeted Mutagenesis of the ERF Transcription Factor Gene OsERF922

Abstract: Rice blast is one of the most destructive diseases affecting rice worldwide. The adoption of host resistance has proven to be the most economical and effective approach to control rice blast. In recent years, sequence-specific nucleases (SSNs) have been demonstrated to be powerful tools for the improvement of crops via gene-specific genome editing, and CRISPR/Cas9 is thought to be the most effective SSN. Here, we report the improvement of rice blast resistance by engineering a CRISPR/Cas9 SSN (C-ERF922) targeting the OsERF922 gene in rice. Twenty-one C-ERF922-induced mutant plants (42.0%) were identified from 50 T0 transgenic plants. Sanger sequencing revealed that these plants harbored various insertion or deletion (InDel) mutations at the target site. We showed that all of the C-ERF922-induced allele mutations were transmitted to subsequent generations. Mutant plants harboring the desired gene modification but not containing the transferred DNA were obtained by segregation in the T1 and T2 generations. Six T2 homozygous mutant lines were further examined for a blast resistance phenotype and agronomic traits, such as plant height, flag leaf length and width, number of productive panicles, panicle length, number of grains per panicle, seed setting percentage and thousand seed weight. The results revealed that the number of blast lesions formed following pathogen infection was significantly decreased in all 6 mutant lines compared with wild-type plants at both the seedling and tillering stages. Furthermore, there were no significant differences between any of the 6 T2 mutant lines and the wild-type plants with regard to the agronomic traits tested. We also simultaneously targeted multiple sites within OsERF922 by using Cas9/Multi-target-sgRNAs (C-ERF922S1S2 and C-ERF922S1S2S3) to obtain plants harboring mutations at two or three sites. Our results indicate that gene modification via CRISPR/Cas9 is a useful approach for enhancing blast resistance in rice.

The Arabidopsis GA1 locus encodes the cyclase ent-kaurene synthetase A of gibberellin biosynthesis.

Abstract: The first committed step in the gibberellin (GA) biosynthetic pathway is the conversion of geranylgeranyl pyrophosphate (GGPP) through copalyl pyrophosphate (CPP) to ent-kaurene catalyzed by ent-kaurene synthetases A and B. The ga1 mutants of Arabidopsis are gibberellin-responsive male-sterile dwarfs. Biochemical studies indicate that biosynthesis of GAs in the ga1 mutants is blocked prior to the synthesis of ent-kaurene. The GA1 locus was cloned previously using the technique of genomic subtraction. Here, we report the isolation of a nearly full-length GA1 cDNA clone from wild-type Arabidopsis. This cDNA clone encodes an active protein and is able to complement the dwarf phenotype in ga1-3 mutants by Agrobacterium-mediated transformation. In Escherichia coli cells that express both the Arabidopsis GA1 gene and the Erwinia uredovora gene encoding GGPP synthase, CPP was accumulated. This result indicates that the GA1 gene encodes the enzyme ent-kaurene synthetase A, which catalyzes the conversion of GGPP to CPP. Subcellular localization of the GA1 protein was studied using 35S-labeled GA1 protein and isolated pea chloroplasts. The results showed that the GA1 protein is imported into and processed in pea chloroplasts in vitro.

Targeted disruption of the housekeeping gene encoding glucose 6-phosphate dehydrogenase (G6PD): G6PD is dispensable for pentose synthesis but essential for defense against oxidative stress

Abstract: Glucose 6-phosphate dehydrogenase (G6PD) is a housekeeping enzyme encoded in mammals by an X-linked gene. It has important functions in intermediary metabolism because it catalyzes the first step in the pentose phosphate pathway and provides reductive potential in the form of NADPH. In human populations, many mutant G6PD alleles (some present at polymorphic frequencies) cause a partial loss of G6PD activity and a variety of hemolytic anemias, which vary from mild to severe. All these mutants have some residual enzyme activity, and no large deletions in the G6PD gene have ever been found. To test which, if any, function of G6PD is essential, we have disrupted the G6PD gene in male mouse embryonic stem cells by targeted homologous recombination. We have isolated numerous clones, shown to be recombinant by Southern blot analysis, in which G6PD activity is undetectable. We have extensively characterized individual clones and found that they are extremely sensitive to H2O2 and to the sulfydryl group oxidizing agent, diamide. Their markedly impaired cloning efficiency is restored by reducing the oxygen tension. We conclude that G6PD activity is dispensable for pentose synthesis, but is essential to protect cells against even mild oxidative stress.