Mutant Plastids Research Articles

Sorting of mitochondrial and plastid heteroplasmy in Arabidopsis is extremely rapid and depends on MSH1 activity

The fate of new mitochondrial and plastid mutations depends on their ability to persist and spread among the numerous organellar genome copies within a cell (heteroplasmy). The extent to which heteroplasmies are transmitted across generations or eliminated through genetic bottlenecks is not well understood in plants, in part because their low mutation rates make these variants so infrequent. Disruption of MutS Homolog 1 (MSH1), a gene involved in plant organellar DNA repair, results in numerous de novo point mutations, which we used to quantitatively track the inheritance of single nucleotide variants in mitochondrial and plastid genomes in Arabidopsis. We found that heteroplasmic sorting (the fixation or loss of a variant) was rapid for both organelles, greatly exceeding rates observed in animals. In msh1 mutants, plastid variants sorted faster than those in mitochondria and were typically fixed or lost within a single generation. Effective transmission bottleneck sizes (N) for plastids and mitochondria were N ∼ 1 and 4, respectively. Restoring MSH1 function further increased the rate of heteroplasmic sorting in mitochondria (N ∼ 1.3), potentially because of its hypothesized role in promoting gene conversion as a mechanism of DNA repair, which is expected to homogenize genome copies within a cell. Heteroplasmic sorting also favored GC base pairs. Therefore, recombinational repair and gene conversion in plant organellar genomes can potentially accelerate the elimination of heteroplasmies and bias the outcome of this sorting process.

Roles for the chloroplast-localized pentatricopeptide repeat protein 30 and the 'mitochondrial' transcription termination factor 9 in chloroplast quality control.

Chloroplasts constantly experience photo-oxidative stress while performing photosynthesis. This is particularly true under abiotic stresses that lead to the accumulation of reactive oxygen species (ROS) which oxidize DNA, proteins and lipids. Reactive oxygen species can also act as signals to induce acclimation through chloroplast degradation, cell death and nuclear gene expression. To better understand the mechanisms behind ROS signaling from chloroplasts, we have used the Arabidopsis thaliana mutant plastid ferrochelatase two (fc2) that conditionally accumulates the ROS singlet oxygen (1 O2 ) leading to chloroplast degradation and eventually cell death. Here we have mapped mutations that suppress chloroplast degradation in the fc2 mutant and demonstrate that they affect two independent loci (PPR30 and mTERF9) encoding chloroplast proteins predicted to be involved in post-transcriptional gene expression. These mutants exhibited broadly reduced chloroplast gene expression, impaired chloroplast development and reduced chloroplast stress signaling. Levels of 1 O2 , however, could be uncoupled from chloroplast degradation, suggesting that PPR30 and mTERF9 are involved in ROS signaling pathways. In the wild-type background, ppr30 and mTERF9 mutants were also observed to be less susceptible to cell death induced by excess light stress. While broad inhibition of plastid transcription with rifampicin was also able to suppress cell death in fc2 mutants, specific reductions in plastid gene expression using other mutations was not always sufficient. Together these results suggest that plastid gene expression, or the expression of specific plastid genes by PPR30 and mTERF0, is a necessary prerequisite for chloroplasts to activate the 1 O2 signaling pathways to induce chloroplast quality control pathways and/or cell death.

MSH1 is required for maintenance of the low mutation rates in plant mitochondrial and plastid genomes

Mitochondrial and plastid genomes in land plants exhibit some of the slowest rates of sequence evolution observed in any eukaryotic genome, suggesting an exceptional ability to prevent or correct mutations. However, the mechanisms responsible for this extreme fidelity remain unclear. We tested seven candidate genes involved in cytoplasmic DNA replication, recombination, and repair (POLIA, POLIB, MSH1, RECA3, UNG, FPG, and OGG1) for effects on mutation rates in the model angiosperm Arabidopsis thaliana by applying a highly accurate DNA sequencing technique (duplex sequencing) that can detect newly arisen mitochondrial and plastid mutations even at low heteroplasmic frequencies. We find that disrupting MSH1 (but not the other candidate genes) leads to massive increases in the frequency of point mutations and small indels and changes to the mutation spectrum in mitochondrial and plastid DNA. We also used droplet digital PCR to show transmission of de novo heteroplasmies across generations in msh1 mutants, confirming a contribution to heritable mutation rates. This dual-targeted gene is part of an enigmatic lineage within the mutS mismatch repair family that we find is also present outside of green plants in multiple eukaryotic groups (stramenopiles, alveolates, haptophytes, and cryptomonads), as well as certain bacteria and viruses. MSH1 has previously been shown to limit ectopic recombination in plant cytoplasmic genomes. Our results point to a broader role in recognition and correction of errors in plant mitochondrial and plastid DNA sequence, leading to greatly suppressed mutation rates perhaps via initiation of double-stranded breaks and repair pathways based on faithful homologous recombination.

Exploring the Limits and Causes of Plastid Genome Expansion in Volvocine Green Algae.

Plastid genomes are not normally celebrated for being large. But researchers are steadily uncovering algal lineages with big and, in rare cases, enormous plastid DNAs (ptDNAs), such as volvocine green algae. Plastome sequencing of five different volvocine species has revealed some of the largest, most repeat-dense plastomes on record, including that of Volvox carteri (∼525 kb). Volvocine algae have also been used as models for testing leading hypotheses on organelle genome evolution (e.g., the mutational hazard hypothesis), and it has been suggested that ptDNA inflation within this group might be a consequence of low mutation rates and/or the transition from a unicellular to multicellular existence. Here, we further our understanding of plastome size variation in the volvocine line by examining the ptDNA sequences of the colonial species Yamagishiella unicocca and Eudorina sp. NIES-3984 and the multicellular Volvox africanus, which are phylogenetically situated between species with known ptDNA sizes. Although V. africanus is closely related and similar in multicellular organization to V. carteri, its ptDNA was much less inflated than that of V. carteri. Synonymous- and noncoding-site nucleotide substitution rate analyses of these two Volvox ptDNAs suggest that there are drastically different plastid mutation rates operating in the coding versus intergenic regions, supporting the idea that error-prone DNA repair in repeat-rich intergenic spacers is contributing to genome expansion. Our results reinforce the idea that the volvocine line harbors extremes in plastome size but ultimately shed doubt on some of the previously proposed hypotheses for ptDNA inflation within the lineage.

Effects of mutant cytoplasmic factors on the expression of agronomic traits in cowpea, <i>Vigna unguiculata</i> (L.) Walp.

Nuclear genes have to a large extent been exploited in genetic cowpea improvement, but little is known about the effects of cytoplasmic mutations on agronomic traits. This study investigated the effect of cytoplasmic factors agronomic traits in crosses between two cytoplasmic mutants and some nuclear cowpea lines. The experiment was conducted at Department of Crop Protection Environmental Biology, University of Ibadan. A yellow variant derived from the sorting-out of mutant plastids of mixed cells lineage (IB-Y-Cyt.) was crossed to four normal green lines and a variegated mutant in all possible combinations. F 1 and reciprocal-F 1 progenies from the crosses and six generations comprising parents, F 1 , F 2 and their reciprocals from four selected crosses were evaluated in pots using Randomized Complete Block Design with four replicates. Data were collected on the following quantitative traits; days to flowering and ripe pod, pod length and seed traits. Data were analyzed using t -test and correlation coefficient analysis. Crosses involving the cytoplasmic mutants as female parents had negative effects on most of the traits studied. The mutant cytoplasms had negative effects on traits affecting the productive capacity of the plants as well as related traits in the F 1 generation, these effects in turn transcend to most of the quantitative traits leading to poor seed formation especially in the yellow cytoplasmic mutant. Keywords: Cytoplasmic mutation, Agronomic traits, Cowpea, Uniparental, Reciprocal differences

Rescue of Deletion Mutants to Isolate Plastid Transformants in Higher Plants.

Plastid transformation is an attractive alternative to nuclear transformation enabling manipulation of native plastid genes and the insertion of foreign genes into plastids for applications in agriculture and industrial biotechnology. Transformation is achieved using dominant positive selection markers that confer resistance to antibiotics. The very high copy number of plastid DNA means that a prolonged selection step is required to obtain a uniform population of transgenic plastid genomes. Repair of mutant plastid genes with the corresponding functional allele allows selection based on restoration of the wild type phenotype. The use of deletion rather than point mutants avoids spontaneous reversion back to wild type. Combining antibiotic resistance markers with native plastid genes speeds up the attainment of homoplasmy and allows early transfer of transplastomic lines to soil where antibiotic selection is replaced by selection for photoautotrophic growth. Here we describe our method using the wild type rbcL gene as a plastid transformation marker to restore pigmentation and photosynthesis to a pale green heterotrophic rbcLmutant.

Significance of accumulation of the alarmone (p)ppGpp in chloroplasts for controlling photosynthesis and metabolite balance during nitrogen starvation in Arabidopsis.

The regulatory nucleotides, guanosine 5'-triphosphate 3'-diphosphate (pppGpp) and guanosine 5'-diphosphate 3'-diphosphate (ppGpp), were originally identified in Escherichia coli, and control a large set of gene expression and enzyme activities. The (p)ppGpp-dependent control of cell activities is referred to as the stringent response. A growing number of (p)ppGpp synthase/hydrolase homologs have been identified in plants, which are localized in plastids in Arabidopsis thaliana. We recently reported that the Arabidopsis mutant overproducing ppGpp in plastids showed dwarf chloroplasts, and transcript levels in the mutant plastids were significantly suppressed. Furthermore, the mutant showed more robust growth than the wild type (WT), especially under nutrient-deficient conditions, although the mechanisms are unclear. To better understand the impact of the ppGpp accumulation on plant responses to nutrient deficiency, photosynthetic activities and metabolic changes in the ppGpp-overproducing mutant were characterized here. Upon transition to the nitrogen-deficient conditions, the mutant showed reduction of ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) contents, and effective and maximum quantum yield of photosystem II compared with WT. The mutant also showed more obvious changes in key metabolite levels including some amino acid contents than WT; similar metabolic change is known to be critical for plants to maintain carbon-nitrogen balance in their cells. These results suggest that artificially overproducing ppGpp modulates the organelle functions that play an important role in controlling photosynthetic performance and metabolite balance during nitrogen starvation.

Multiple fates of non‐mature lumenal proteins in thylakoids

Most proteins found in the thylakoid lumen are synthesized in the cytosol with an N-terminal extension consisting of transient signals for chloroplast import and thylakoid transfer in tandem. The thylakoid-transfer signal is required for protein sorting from the stroma to thylakoids, mainly via the cpSEC or cpTAT pathway, and is removed by the thylakoidal processing peptidase in the lumen. An Arabidopsis mutant lacking one of the thylakoidal processing peptidase homologs, Plsp1, contains plastids with anomalous thylakoids and is seedling-lethal. Furthermore, the mutant plastids accumulate two cpSEC substrates (PsbO and PetE) and one cpTAT substrate (PsbP) as intermediate forms. These properties of plsp1-null plastids suggest that complete maturation of lumenal proteins is a critical step for proper thylakoid assembly. Here we tested the effects of inhibition of thylakoid-transfer signal removal on protein targeting and accumulation by examining the localization of non-mature lumenal proteins in the Arabidopsis plsp1-null mutant and performing a protein import assay using pea chloroplasts. In plsp1-null plastids, the two cpSEC substrates were shown to be tightly associated with the membrane, while non-mature PsbP was found in the stroma. The import assay revealed that inhibition of thylakoid-transfer signal removal did not disrupt cpSEC- and cpTAT-dependent translocation, but prevented release of proteins from the membrane. Interestingly, non-mature PetE2 was quickly degraded under light, and unprocessed PsbO1 and PsbP1 were found in a 440-kDa complex and as a monomer, respectively. These results indicate that the cpTAT pathway may be disrupted in the plsp1-null mutant, and that there are multiple mechanisms to control unprocessed lumenal proteins in thylakoids.

The 2013 Plant and Cell Physiology Database Issue

We are pleased to announce the publication of this third Database Issue of Plant and Cell Physiology (PCP). It contains four new databases and seven updated databases (Tables 1, 2). Our aim with this issue is to provide a forum for discussion of bioinformatics research, in particular the development and maintenance of the infrastructure of web databases for plant science (Matsuoka and Yano 2010). The databases described in this issue cover a broad range of omics topics. The genome and transcriptome databases permit management of the flood of data from recent high-throughput sequencers, and have been rapidly extended to apply to non-model plants. On the other hand, metabolome and phenome data still require databases and web tools to store, annotate and compare the data. In the following paragraphs, we briefly introduce the 11 databases in this issue and broadly describe their functions. Databases of physiological data are particularly interesting from a PCP standpoint. The Chloroplast Function Database (Myouga et al. 2013) offers phenotype data obtained from 2,495 Arabidopsis mutants. The data include visually identifiable mutant phenotypes and plastid ultrastructures. UniVIO (Sakakibara et al. 2013) combines hormone concentration (hormonome) and transcriptome data for 14 organ parts of rice plants at the reproductive stage, and of seedling shoots of three gibberellin signaling mutants. RAP-DB has played a fundamental role in rice research (Sakai et al. 2013). Gene structures and annotations have been updated in it ever since 2005. The database now incorporates literature-based annotations, and RNA-Seq data and single nucleotide polymorphism (SNPs) can be queried using a customized browser. Similar genomeand expressed sequence tag (EST)-based databases have been developed for orchid (Su et al. 2013, Tsai et al. 2013) and radish (Shen et al. 2013). Although the amount of information available is rather limited in these non-model plants, these databases generate a new impetus for elucidating the biological mechanisms in these species and their relatives. In addition to databases based purely on experimental data, this issue also includes databases populated by computational predictions. To unravel the stress regulome, STIFDB2 (Naika et al. 2013) combines stress-responsive transcription factors with binding sites that have been predicted by Hidden Markov models for Arabidopsis and rice. ERISdb (Szcześniak et al. 2013) predicts cis-regulatory motifs for splicing events in eight plant species. KNApSAcK-3D (Nakamura et al. 2013) and PRIMe (Sakurai et al. 2013) both provide new tools for metabolome research. KNApSAcK-3D predicts the three-dimensional structure of all the metabolic compounds in the KNApSAcK database. PRIMe provides a series of tools

The Significance of Protein Maturation by Plastidic Type I Signal Peptidase 1 for Thylakoid Development in Arabidopsis Chloroplasts

Thylakoids are the chloroplast internal membrane systems that house light-harvesting and electron transport reactions. Despite the important functions and well-studied constituents of thylakoids, the molecular mechanism of their development remains largely elusive. A recent genetic study has demonstrated that plastidic type I signal peptidase 1 (Plsp1) is vital for proper thylakoid development in Arabidopsis (Arabidopsis thaliana) chloroplasts. Plsp1 was also shown to be necessary for processing of an envelope protein, Toc75, and a thylakoid lumenal protein, OE33; however, the relevance of the protein maturation in both of the two distinct subcompartments for proper chloroplast development remained unknown. Here, we conducted an extensive analysis of the plsp1-null mutant to address the significance of lumenal protein maturation in thylakoid development. Plastids that lack Plsp1 were found to accumulate vesicles of variable sizes in the stroma. Analyses of the mutant plastids revealed that the lack of Plsp1 causes a reduction in accumulation of thylakoid proteins and that Plsp1 is involved in maturation of two additional lumenal proteins, OE23 and plastocyanin. Further immunoblotting and electron microscopy immunolocalization studies showed that OE33 associates with the stromal vesicles of the mutant plastids. Finally, we used a genetic complementation system to demonstrate that accumulation of improperly processed forms of Toc75 in the plastid envelope does not disrupt normal plant development. These results suggest that proper maturation of lumenal proteins may be a key process for correct assembly of thylakoids.

Cytoplasmic male sterility in barley II. Physiology and anther cytology of msml

The cytoplasmic male sterile, msml (male sterile maternal 1), of barley was studied along with the restored form and with the nearly isogenic normal barley. Under light microscopy and TEM, the male sterile was found to have an abnormally productive and uncontrolled secretion of sporopollenin in its anthers. The energy of the anther is probably wasted in the biosynthesis of sporopollenin precursors. Consequently, several abnormalities are discernable in the cytological appearance of the anther at different developmental stages. The sporopollenin of the male sterile or partially restored male sterile displays a Fast blue B stainability different from that of the normal type. As a result of several observations, the msml stock would appear to be a plastid mutant. In the leaf of the unrestored male sterile plant the ratio of xanthophylls to β-carotene is about one tenth higher than that of the fertile plant. The restored msml plants were found to have a somewhat higher proportion of chlorophyll b in their chlorophylls than the unrestored material. The male sterile was found to be affected by the Helminthosporium maydis race T toxin in a way similar to that of the normal barley. This male sterility is expected to be beneficial to hybrid breeding of barley.

Plastid tRNA Genes trnC‐GCA and trnN‐GUU are essential for plant cell development

Higher plant chloroplast genomes code for a conserved set of 30 tRNAs. This set is believed to be sufficient to support translation, although import of cytosolic tRNA has been proposed to provide additional tRNA species to the chloroplast. Previous knock-outs of tRNA genes, or the pronounced reduction of the level of selected tRNAs, has not led to severe phenotypes. We deleted the two tRNA genes trnN-GUU and trnC-GCA independently from the plastid chromosome of tobacco. No homoplastomic tissue of either DeltatrnN or DeltatrnC plants could be isolated. Both mutants exhibit occasional loss of leaf sectors, and mutant plastid chromosomes are rapidly lost upon relief of selective pressure. This suggests that the knock-out of both trn genes is lethal, and that both tRNA species are required for cell survival. Surprisingly, the impact on chloroplast and cell development differs pronouncedly between the two mutants. Heteroplastomic DeltatrnC and DeltatrnN tissue exhibit different aberrations of the internal membrane systems and, more importantly, heteroplastomic DeltatrnN plants are variegated. Accumulation of tRNA-N and plastid-encoded proteins is reduced in white sectors of DeltatrnN plants, and differentiation of palisade cells is abolished. Our data demonstrate that plastid tRNAs are essential, i.e. not complemented by cytosolic tRNA, and have a differential impact on chloroplast and plant cell development.

Spectral features and structure of chloroplasts under an early block of chlorophyll synthesis

A xantha mutant of cotton (Gossypium hirsutum L.) with blocked synthesis of 5-aminolevulinic acid in light accumulates 30 times less chlorophyll than the parental strain. Formation of the chloroplast membrane system in the mutant stops at very early stages, mostly vesicles and single short thylakoids. The mutant plastid membranes contain only light-harvesting chlorophyll-a/b-protein complexes I and II with fluorescence maxima at 728 and 681 nm, respectively. Thus, an early block of chlorophyll synthesis impairs the formation and function of photosystem reaction centers and retards the development of the chloroplast membrane system at the stage of proplastids.

A Second Thylakoid Membrane-localized Alb3/OxaI/YidC Homologue Is Involved in Proper Chloroplast Biogenesis in Arabidopsis thaliana

The integral membrane proteins Alb3, OxaI, and YidC belong to an evolutionary conserved protein family mediating protein insertion into the thylakoid membrane of chloroplasts, the inner membrane of mitochondria, and bacteria, respectively. Whereas OxaI and YidC are involved in the insertion of a wide range of membrane proteins, the function of Alb3 seems to be limited to the insertion of a subset of the light-harvesting chlorophyll-binding proteins. In this study, we identified a second chloroplast homologue of the Alb3/OxaI/YidC family, named Alb4. Alb4 is almost identical to the Alb3/OxaI/YidC domain of the previously described 110-kDa inner envelope protein Artemis. We show that Alb4 is expressed as a separate 55-kDa protein and that Artemis was identified mistakenly. Alb4 is located in the thylakoid membrane of Arabidopsis thaliana chloroplasts. Analysis of an Arabidopsis mutant (Salk_136199) and RNA interference lines with a reduced level of Alb4 revealed chloroplasts with an altered ultrastructure. Mutant plastids are larger and more spherical in appearance, and the grana stacks within the mutant lines are less appressed than in the wild-type chloroplasts. These data indicate that Alb4 is required for proper chloroplast biogenesis.

Delayed embryo development in the ARABIDOPSIS TREHALOSE‐6‐PHOSPHATE SYNTHASE 1 mutant is associated with altered cell wall structure, decreased cell division and starch accumulation

The tps1 mutant, which is disrupted in the TREHALOSE-6-PHOSPHATE SYNTHASE 1 gene, has been previously characterized as a recessive embryo lethal. tps1 embryos do not develop past late torpedo or early cotyledon stage. We report here that at the ultrastructural, biochemical, and transcriptional levels tps1 exhibits many features typically associated with the maturation phase. The appearance of storage reserve transcripts and organelles follows the same temporal pattern in tps1 and wild-type (WT) embryos in the same silique as does accumulation of storage lipid and protein. The mutant plastids accumulate large starch granules that persist until the end of seed development, in contrast with WT plastids where starch accumulation is transient. The transcriptome of tps1 embryos shows a coordinate downregulation of genes involved in starch and sucrose degradation. Interestingly, genes involved in lipid mobilization and gluconeogenesis are induced in tps1 embryos. The cell walls of tps1 embryos show a remarkable degree of thickening at the ultrastructural level and immunodetection of cell wall components shows that altered deposition of pectins accounts for this altered morphology. Consistent with this at the transcriptome level, genes involved in sugar nucleotide and pectin metabolism are altered in the mutant. The frequency of cell division in tps1 embryos is half that of the wild type at the heart and torpedo stages. These results suggest that TPS1 may play a major role in coordinating cell wall biosynthesis and cell division with cellular metabolism during embryo development.

Observation of High-frequency Occurrence of Chimeral Adventitious Shoots in Tissue Culture from the Chimeral Tissues of Pelargonium zonale

In Pelargonium, the plastid mutation in three independent cell layers L1, L2, and L3, can produce plastid chimeras with visible shoot colour difference such as GWG (green-white-green) and GGW (green-green-white). Chimera can be used to trace the relationship between the cell layers of different genotypes during shoot development and the effect of the mutated genes on shoot development. In this study, we have obtained different adventitious shoots with GGG, GWG, GGW, and WWW combinations of cell layers through tissue culture of petioles and internodes from GGW and GWG chimeras of Pelargonium zonale `Mrs Pollock'. Much higher percentage (14.9%) of chimeral adventitious shoots was obtained from GGW tissues than from GWG tissues (4.2%). Of the 10.8% chimeral adventitious shoots regenerated in this experiment, 8.6% are different from the original type of explants. This result indicated that cells at least in both L2 and L3 of the explants were involved in the regeneration of the adventitious shoots. The number of shoot types regenerated is likely dependent on the number and the type of cells that were in direct contact with the culture medium. It is suggested that the mixed cells can be used to produce the chimera by tissue culture. Three possible ways to form the chimeras in vitro culture were discussed. Chemical names used: TDZ =1-phenyl-3-(1,2,3-thiadiazol-5-yl)urea (Thidiazuron); IAA = Indole-3-acetic acid; PVP = polyvinylpyrrolidone.

The effect of oxidative stress on nitrosomethylurea-induced mutagenesis in sunflower Helianthus annuus L.

The effect of hyberbaric oxygenation on mutagenicity of nitrosomethylurea (NMU) was examined. It was shown that in the regimes studied, hyperbaric oxygenation enhances the NMU mutagenic effect both on the nuclear and on the plastid genetic material of sunflower, but do not increase intensity of free-radical reactions and do not induce plastid mutations and chromosome aberrations per se. At high concentrations inducing chromosome aberrations (0.03%), NMU was shown to enhance the level of free-radical processes. Possible mechanisms of the increase of NMU-induced mutagenesis by hyperbaric oxygenation are discussed.

Induced of plastid mutations in soybean plant ( Glycine max L. Merrill) with gamma radiation and determination with RAPD

The aim of our study was to induce with radiation of atrazine resistant and tolerated mutants in Coles, Amsoy-71 and 1937 soybean varieties. Atrazine that is photosynthetic inhibitor is the most important herbicide of S-triazin group, and shows toxic effect on soybean plant. For the improvement of the atrazine resistant plants with mutation breeding, the seeds belonging to the three varieties were irradiated with 200 Gy of gamma radiation dose. The irradiated seeds were sown in the field and at the end of harvesting season, every pod at node situated on the main stem was picked up separately and M 2 generations were obtained. At the plants, which were obtained from M 2 generation, chlorophyll mutants were determined and atrazine selection was made. The percentage of chlorophyll mutants for Amsoy-71, Coles and 1937 soybean varieties were found as 1.07, 1.48 and 1.32, respectively. At the end of atrazine selection, the percentages of atrazine resistant plants for Amsoy-71, Coles and 1937 soybean varieties were 0.80, 0.60 and 0.53, respectively. The percentages of atrazine tolerated plants were 1.07, 1.18 and 1.05, respectively as well. In our research; the differences among the mutants replying to atrazine in various concentrations were examined by using RAPD procedure as the molecular marker techniques in comparison with polymorphism. In the study done by using 14 primers; according to the amplification results, the differences between atrazine resistant plants were shown.

Structural and Functional Characteristics of the Mutant Plastids from Extranuclear Variegated Forms of Sunflower

The electron-microscopic analysis of extranuclear variegated forms of sunflower demonstrated that plastids from white leaf areas are practically devoid of inner membrane structures. The mutants and the initial (wild-type) green line were compared by the contents of chlorophyll (Chl), carotenoids, and 70S ribosomes and by the activity of Rubisco. A mutant Var10 line was used to demonstrate that the primary characteristic manifestations of Chl deficiency include the synchronous retardation of the synthesis of a specific Chl precursor, 5-aminolevulinate, a decrease in the Chl a/b ratio to the level below that in the wild type, and the reduction of photosystem (PS) I Chl luminance. The progress of photodestructive processes in the mutant aggravated the listed disturbances; as a result, PSII complexes and light-harvesting complexes gradually degraded. The manifestation of the trait of Chl-deficiency notably varied depending on plant growth conditions (primarily, temperature and illumination regimes). The dependence of this manifestation on light and temperature was observed only at the early stage of development of pigment-containing tissues. When plants terminated this stage under low irradiation, they did not become Chl-deficient, and when their leaves were subsequently transferred to the conditions promoting the maximum pigment anomaly, the destructive processes did not advance in the mutant tissue. The authors assume that the plastom mutations under study impair the control over the expression of the structural plastogenes that determine the synthesis of pigment and protein components of the photosynthetic apparatus, rather than directly damage to the primary structures of plastogenes.

Spontaneous and Nitrosomethylurea-Induced Reversions in Plastome Chlorophyll Mutants of Sunflower Helianthus annuus L.

Spontaneous and induced revertants obtained from plastome chlorophyll mutants of sunflower were subjected to genetic analysis. The mechanism of reversion (nuclear suppression, plastid suppression, true reversion, or mitochondrial suppression) of the plastid mutation was shown to affect the degree of restoration of morphological and physiological traits of sunflower plants.