On Antimutagenesis Mechanism in Plants

The effect of nitrosylmethylurea (NMU) on the mitotic index and the frequency of cells with aberrations, as well as the effects of pre- and posttreatment with antioxidant ambiol on the NMU effects were studied on seedlings of common winter wheat Triticum aestivum, cultivar Moskovskaya 39. Both pre- and posttreatment with ambiol resulted in antimutagenic effect but after posttreatment, the effect was lower. Irrespective of type of seedling treatment with ambiol and the time of their fixation (45, 48, and 51 h), when mitotic index is plotted versus frequency of cells with aberrations, all experimental points fall on the same regression line with coefficient of correlation of −0.82 (P < 0.001). This implies that the same mechanism underlies antimutagenic effect irrespective of when the antimutagen was applied, before or after the knockout mutagen dose. This also suggests that the antimutagenic effect is independent of the degree of the mutagen-induced damage, because by the time of posttreatment, the volume of genome damage is already determined and the antimutagen fails to change it. Finally, this suggests that irrespective of time of antimutagen treatment, the mutation frequency is reduced by the mechanism of stimulated repopulation.__________Translated from Genetika, Vol. 41, No. 5, 2005, pp. 676–679.Original Russian Text Copyright © 2005 by Serebryanyi, Zoz, Morozova.     

Numerous transposed sequences of mitochondrial cytochrome oxidase I-II in aphids of the genus Sitobion (Hemiptera: Aphididae)

Polymerase chain reaction (PCR) products corresponding to 803 bp of the cytochrome oxidase subunits I and II region of mitochondrial DNA (mtDNA COI-II) were deduced to consist of multiple haplotypes in three Sitobion species. We investigated the molecular basis of these observations. PCR products were cloned, and six clones from one individual per species were sequenced. In each individual, one sequence was found commonly, but also two or three divergent sequences were seen. The divergent sequences were shown to be nonmitochondrial by sequencing from purified mtDNA and Southern blotting experiments. All seven nonmitochondrial clones sequenced to completion were unique. Nonmitochondrial sequences have a high proportion of unique sites, and very few characters are shared between nonmitochondrial clones to the exclusion of mtDNA. From these data, we infer that fragments of mtDNA have been transposed separately (probably into aphid chromosomes), at a frequency only known to be equalled in humans. The transposition phenomenon appears to occur infrequently or not at all in closely related genera and other aphids investigated. Patterns of nucleotide substitution in mtDNA inferred over a parsimony tree are very different from those in transposed sequences. Compared with mtDNA, nonmitochondrial sequences have less codon position bias, more even exchanges between A, G, C and T, and a higher proportion of nonsynonymous replacements. Although these data are consistent with the transposed sequences being under less constraint than mtDNA, changes in the nonmitochondrial sequences are not random: there remains significant position bias, and probable excesses of synonymous replacements and of conservative inferred amino acid replacements. We conclude that a proportion of the inferred change in the nonmitochondrial sequences occurred before transposition. We believe that Sitobion aphids (and other species exhibiting mtDNA transposition) may be important for studying the molecular evolution of mtDNA and pseudogenes. However, our data highlight the need to establish the true evolutionary relationships between sequences in comparative investigations.      

Initial verification of the resistance management strategy for Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae) in Australia

Shortly after the initial detection of western flower thrips (WFT), Frankiniella occidentalis (Pergande), in Australia during 1993 a resistance management strategy based on the alternation of chemical groups was implemented. This study aimed to verify this strategy by field testing α-cypermethrin against WFT with and without chemical alternation. Up to 114 times α-cypermethrin resistance (at LC50) was detected and resistance increased with and without chemical alternation; however, chemical alternation did significantly reduce numbers of thrips compared with a nonalternation strategy. Resistance has the potential to undermine the sustainable use of those chemicals to which there is no current detectable resistance. Consequently, chemicals with a high frequency and level of resistance against WFT need to be identified through monitoring and quickly eliminated from WFT chemical control recommendations.     

Three-iron-sulfur centers of pea mitochondria

EPR signals of three distinct types of three-iron-sulfur center were observed in pea mitochondria: the signal of Center S-3 (low-field peak at g = 2.016), the signal of Center ISP-1 (low-field peak at g = 2.024) and the signal of the axial Center ISP-2 with two maxima, at g = 2.027 and 2.016. Succinate increases the signal amplitude of Center ISP-1 and diminishes that of Center ISP-2; malate has an opposite effect. Membrane damage enhances the effect of malate and decreases that of succinate.     

Alveolar epithelial cells undergo epithelial-to-mesenchymal transition in response to endoplasmic reticulum stress

Expression of mutant surfactant protein C (SFTPC) results in endoplasmic reticulum (ER) stress in type II alveolar epithelial cells (AECs). AECs have been implicated as a source of lung fibroblasts via epithelial-to-mesenchymal transition (EMT); therefore, we investigated whether ER stress contributes to EMT as a possible mechanism for fibrotic remodeling. ER stress was induced by tunicamyin administration or stable expression of mutant (L188Q) SFTPC in type II AEC lines. Both tunicamycin treatment and mutant SFTPC expression induced ER stress and the unfolded protein response. With tunicamycin or mutant SFTPC expression, phase contrast imaging revealed a change to a fibroblast-like appearance. During ER stress, expression of epithelial markers E-cadherin and Zonula occludens-1 decreased while expression of mesenchymal markers S100A4 and α-smooth muscle actin increased. Following induction of ER stress, we found activation of a number of pathways, including MAPK, Smad, β-catenin, and Src kinase. Using specific inhibitors, the combination of a Smad2/3 inhibitor (SB431542) and a Src kinase inhibitor (PP2) blocked EMT with maintenance of epithelial appearance and epithelial marker expression. Similar results were noted with siRNA targeting Smad2 and Src kinase. Together, these studies reveal that induction of ER stress leads to EMT in lung epithelial cells, suggesting possible cross-talk between Smad and Src kinase pathways. Dissecting pathways involved in ER stress-induced EMT may lead to new treatment strategies to limit fibrosis.     

A simple method for studying whole sections of rice grain

We developed a new and simple method to collect sections of a whole brown rice kernel for investigation of histological properties. A single kernel of rice was dehydrated through a graded ethanol series, transferred to xylene, and embedded in paraffin. During sectioning of the blocks using a rotary microtome, we used a special adhesive tape to collect and place the sections on slides so they remained flat. The use of the adhesive tape technique combined with autofluorescence characteristics allowed us to visualize cell walls throughout an entire longitudinal or transverse section of a whole rice kernel. We obtained scanning electron microscopy images of the sections to determine section quality.     

On antimutagenesis mechanism in plants

The effect of nitrosylmethylurea (NMU) on the mitotic index and the frequency of cells with aberrations, as well as the effects of pre- and posttreatment with antioxidant ambiol on the NMU effects were studied on seedlings of common winter wheat Triticum aestivum, cultivar Moskovskaya 39. Both pre- and posttreatment with ambiol resulted in antimutagenic effect but after posttreatment, the effect was lower. Irrespective of type of seedling treatment with ambiol and the time of their fixation (45, 48, and 51 h), when mitotic index is plotted versus frequency of cells with aberrations, all experimental points fall on the same regression line with coefficient of correlation of -0.82 (P < 0.001). This implies that the same mechanism underlies antimutagenic effect irrespective of when the antimutagen was applied, before or after the knockout mutagen dose. This also suggests that the antimutagenic effect is independent of the degree of the mutagen-induced damage, because by the time of posttreatment, the volume of genome damage is already determined and the antimutagen fails to change it. Finally, this suggests that irrespective of time of antimutagen treatment, the mutation frequency is reduced by the mechanism of stimulated repopulation.