Induction of the AOX1D Isoform of Alternative Oxidase in A. thaliana T-DNA Insertion Lines Lacking Isoform AOX1A Is Insufficient to Optimize Photosynthesis when Treated with Antimycin A

Plant respiration is characterized by two pathways for electron transfer to O2, namely the cytochrome pathway （CP） that is linked to ATP production, and the alternative pathway （AP）, where electrons from ubiquinol are directly transferred to O2 via an alternative oxidase （AOX） without concomitant ATP production. This latter pathway is well suited to dispose of excess electrons in the light, leading to optimized photosynthetic performance. We have characterized T- DNA-insertion mutant lines of Arabidopsis thaliana that do not express the major isoform, AOXIA. In standard growth conditions, these plants did not show any phenotype, but restriction of electron flow through CP by antimycin A, which induces AOXIA expression in the wild-type, led to an increased expression of AOXID in leaves of the aoxla-knockout mutant. Despite the increased presence of the AOX1D isoform in the mutant, antimycin A caused inhibition of photosyn- thesis, increased ROS, and ultimately resulted in amplified membrane leakage and necrosis when compared to the wild- type, which was only marginally affected by the inhibitor. It thus appears that AOX1 D was unable to fully compensate for the loss of AOXIA when electron flow via the CP is restricted. A combination of inhibition studies, coupled to metabolite profiling and targeted expression analysis of the P-protein of glycine decarboxylase complex （GDC）, suggests that the aoxla mutants attempt to increase their capacity for photorespiration. However, given their deficiency, it is intriguing that increase in expression neither of AOX1D nor of GDC could fully compensate for the lack of AOXIA to optimize pho- tosynthesis when treated with antimycin A. We suggest that the aoxla mutants can further be used to substantiate the current models concerning the influence of mitochondrial redox on photosynthetic performance and gene expression.     

Oxygation Enhances Growth,Gas Exchange and Salt Tolerance of Vegetable Soybean and Cotton in a Saline Vertisol

Impacts of salinity become severe when the soil is deficient in oxygen. OxygaUon （using aerated water for subsurface drip irrigation of crop） could minimize the impact of salinity on plants under oxygen-limiting soil environments. Pot experiments were conducted to evaluate the effects of oxygation （12% air volume/volume of water） on vegetable soybean （moderately salt tolerant） and cotton （salt tolerant） in a salinized vertisol at 2, 8, 14, 20 dS/m ECe. In vegetable soybean, oxygation increased above ground biomass yield and water use efficiency （WUE） by 13% and 22%, respectively, compared with the control. Higher yield with oxygation was accompanied by greater plant height and stem diameter and reduced specific leaf area and leaf Na^＋ and CI^- concentrations. In cotton, oxygation increased lint yield and WUE by 18% and 16%, respectively, compared with the control, and was accompanied by greater canopy light interception, plant height and stem diameter. Oxygation also led to a greater rate of photosynthesis, higher relative water content in the leaf, reduced crop water stress index and lower leaf water potential. It did not, however, affect leaf Na^＋ or CI^- concentration. Oxygation invariably increased, whereas salinity reduced the K^＋： Na^＋ ratio in the leaves of both species. Oxygation improved yield and WUE performance of salt tolerant and moderately tolerant crops under saline soil environments, and this may have a significant impact for irrigated agriculture where saline soils pose constraints to crop production.     

Interaction between a Dark Septate Endophytic Isolate from Dendrobium sp. and Roots of D. nobile Seedlings

Interactions between an isolate of dark septate endophytes （DSE） and roots of Dendrobium nobile Lindl. seedlings are reported in this paper. The isolate was obtained from orchid mycorrhizas on Dendrobium sp. in subtropical forest. The fungus formed typical orchid mycorrhiza in aseptic co-culture with D. nobile seedlings on modified Murashige-Skoog （MMS） medium. Anatomic observations of the infected roots showed that the DSE hyphae invaded the velamen layer, passed through passage cells in exodermis, entered the cortex cells, and then formed fungal pelotons of orchid mycorrhiza. D. nobile seedlings＇ plant height, stem diameter, new roots number and biomass were greatly enhanced by inoculating the fungus to seedlings. The fungus was identified as Leptodontidium by sequencing the polymerase chain reaction-amplified rDNA ITS1-5,8S-ITS2 （internal transcribed spacer （ITS）） regions and comparison with similar taxa.     

Molecular Characterization of the Calvin Cycle Enzyme Phosphoribulokinase in the Stramenopile Alga Vaucheria litorea and the Plastid Hosting Mollusc Elysia chlorotica

Phosphoribulokinase （PRK）, a nuclear-encoded plastid-localized enzyme unique to the photosynthetic carbon reduction （Calvin） cycle, was cloned and characterized from the stramenopile alga Vaucheria litorea. This alga is the source of plastids for the mollusc （sea slug） Elysia chlorotica which enable the animal to survive for months solely by photoautotrophic CO2 fixation. The 1633-bp V. litorea prk gene was cloned and the coding region, found to be interrupted by four introns, encodes a 405-amino acid protein. This protein contains the typical bipartite target sequence expected of nuclearencoded proteins that are directed to complex （i.e. four membrane-bound） algal plastids. De novo synthesis of PRK and enzyme activity were detected in E. chlorotica in spite of having been starved of V. litorea for several months. Unlike the algal enzyme, PRK in the sea slug did not exhibit redox regulation. Two copies of partial PRK-encoding genes were isolated from both sea slug and aposymbiotic sea slug egg DNA using PCR. Each copy contains the nucleotide region spanning exon 1 and part of exon 2 of V litorea prk, including the bipartite targeting peptide. However, the larger prk fragment also includes intron 1. The exon and intron sequences of prk in E. chlorotica and V/itorea are nearly identical. These data suggest that PRK is differentially regulated in V. litorea and E. chlorotica and at least a portion of the V. litorea nuclear PRK gene is present in sea slugs that have been starved for several months.     

Crossability Barriers in the Interspecific Hybridization between Oryza sativa and O. meyeriana

Oryza meyeriana Baill （GG genome） is a precious germplasm in the tertiary gene pool of cultivated rice （AA genome）, and possesses important traits such as resistance and tolerance to biotic and abiotic stress. However, interspeciflc crossability barrier, a critical bottleneck restricting genes transfer from O. meyeriana to cultivars has led to no hybrids through conventional reproduction. Therefore, the reasons underlying incrossability were investigated in the present report. The results showed that： （i） at 3-7 d after pollination （DAP）, many hybrid embryos degenerated at the earlier globular-shaped stage, and could not develop into the later pear-shaped stage. Meanwhile, free endosperm nuclei started to degenerate at 1 DAP, and cellular endosperm could not form at 3 DAP, leading to nutrition starvation for young embryo development; （ii） at 11-13 DAP, almost all hybrid ovaries aborted. Even though 72.22% of hybrid young embryos were produced in the interspecific hybridization between O. sativa and O. meyeriana, young embryos were not able to further develop into hybrid plantlets via culturing in vitro. The main reason for the incrossability was hybrid embryo inviability, presenting as embryo development stagnation and degeneration since 3 DAP. Some possible approaches to overcome the crossability barriers in the interspecific hybridization between O. sativa and O. meyeriana are discussed.     

Vegetation Mapping of the Mond Protected Area of Bushehr Province （South-west Iran）

Arid regions of the world occupy up to 35% of the earth＇s surface, the basis of various definitions of climatic conditions, vegetation types or potential for food production. Due to their high ecological value, monitoring of arid regions is necessary and modern vegetation studies can help in the conservation and management of these areas. The use of remote sensing for mapping of desert vegetation is difficult due to mixing of the spectral reflectance of bright desert soils with the weak spectral response of sparse vegetation. We studied the vegetation types in the semiarid to arid region of Mond Protected Area, south-west Iran, based on unsupervised classification of the Spot XS bands and then produced updated maps. Sixteen map units covering 12 vegetation types were recognized in the area based on both field works and satellite mapping. Halocnemum strobilaceum and Suaeda fruticosa vegetation types were the dominant types and Ephedra foliata, Salicornia europaea-Suaeda heterophylla vegetation types were the smallest. Vegetation coverage decreased sharply with the increase in salinity towards the coastal areas of the Persian Gulf. The highest vegetation coverage belonged to the riparian vegetation along the Mond River, which represents the northern boundary of the protected area. The location of vegetation types was studied on the separate soil and habitat diversity maps of the study area, which helped in final refinements of the vegetation map produced.     

Identification,expression, and characterization of the highly conserved D-xylose isomerase in animals

D-xylose is a necessary sugar for animals. The xylanase from a mollusk, Ampullaria crossean, was previously reported by our laboratory. This xylanase can degrade the xylan into D-xylose. But there is still a gap in our knowledge on its metabolic pathway. The question is how does the xylose enter the pentose pathway？ With the help of genomic databases and bioinformatic tools, we found that some animals, such as bacteria, have a highly conserved D-xylose isomerase （EC 5.3.1.5）. The xyiose isomerase from a sea squirt, Ciona intestinali, was heterogeneously expressed in Escherichia coli and purified to confirm its function. The recombinant enzyme had good thermal stability in the presence of Mg^2＋. At the optimum temperature and optimum pH environment, its specific activity on D-xylose was 0.331 μmol/mg/min. This enzyme exists broadly in many animals, but it disappeared in the genome of Amphibia-like Xenopus laevis. Its sequence was highly conserved. The xylose isomerases from animals are very interesting proteins for the study of evolution.     

Pitt,a Novel Tetratricopeptide Repeat Protein Involved in Light-Dependent Chlorophyll Biosynthesis and Thylakoid Membrane Biogenesis in Synechocystis sp. PCC 6803

Biogenesis of photosynthetic pigment/protein complexes is a highly regulated process that requires various assisting factors. Here, we report on the molecular analysis of the Pitt gene （sir1644） from the cyanobacterium Synechocystis sp. PCC 6803 （Synechocystis 6803） that encodes a membrane-bound tetratricopeptide repeat （TPR） protein of formerly unknown function. Targeted inactivation of Pitt affected photosynthetic performance and light-dependent chlorophyll synthesis. Yeast two-hybrid analyses and native PAGE strongly suggest a complex formation between Pitt and the light-dependent protochlorophyllide oxidoreductase （POR）. Consistently, POR levels are approximately threefold reduced in the pitt insertion mutant. The membrane sublocalization of Pitt was found to be dependent on the presence of the periplasmic photosystem Ⅱ （PSⅡ） biogenesis factor PratA, supporting the idea that Pitt is involved in the early steps of photosynthetic pigment/protein complex formation.     

The Translational Apparatus of Plastids and Its Role in Plant Development

Chloroplasts （plastids） possess a genome and their own machinery to express it. Translation in plastids occurs on bacterial-type 70S ribosomes utilizing a set of tRNAs that is entirely encoded in the plastid genome. In recent years, the components of the chloroplast translational apparatus have been intensely studied by proteomic approaches and by reverse genetics in the model systems tobacco （plastid-encoded components） and Arabidopsis （nucleus-encoded components）. This work has provided important new insights into the structure, function, and biogenesis of chloroplast ribosomes, and also has shed fresh light on the molecular mechanisms of the translation process in plastids. In addition, mutants affected in plastid translation have yielded strong genetic evidence for chloroplast genes and gene products influencing plant develop- ment at various levels, presumably via retrograde signaling pathway（s）. In this review, we describe recent progress with the functional analysis of components of the chloroplast translational machinery and discuss the currently available evidence that supports a significant impact of plastid translational activity on plant anatomy and morphology.     

The Response Difference of Mitochondria in Recalcitrant Antiaris toxicaria Axes and Orthodox Zea mays Embryos to Dehydration Injury

Long-term preservation of recalcitrant seeds is very difficult because the physiological basis on their desiccation sensitivity is poorly understood. Survival of Antiaris toxicaria axes rapidly decreased and that of immature maize embryos very slowly decreased with dehydration. To understand their different responses to dehydration, we examined the changes in mitochondria activity during dehydration. Although activities of cytochrome （Cyt） c oxidase and malate dehydrogenase of the A. toxicaria axis and maize embryo mitochondria decreased with dehydration, the parameters of maize embryo mitochondria were much higher than those of A. toxicaria, showing that the damage was more severe for the A. toxicaria axis mitochondria than for those of maize embryo. The state I and III respiration of the A. toxicaria axis mitochondria were higher than those of maize embryo, the former rapidly decreased, and the latter slowly decreased with dehydration. The proportion of Cyt c pathway to state III respiration for the A. toxicaria axis mitochondria was low and rapidly decreased with dehydration, and the proportion of alternative oxidase pathway was high and slightly increased with dehydration. In contrast, the proportion of Cyt c pathway for maize embryo mitochondria was high, and that of alternative oxidase pathway was low. Both pathways decreased slowly with dehydration.     

Molecular Characterization of Two Silenced y-type Genes for Glu-B1 in Triticum aestivum ssp. yunnanese and ssp. tibetanum

The high molecular weight glutenin subunits （HMW-GSs） are a major class of common wheat storage proteins. The breadmaking quality of common wheat flour is influenced by the composition of HMW-GSs. In the present study, two unexpressed 1 By genes from Triticum aesitvum L.ssp.yunnanese AS332 and T. aesitvum ssp.tibetanurn AS908 were respectively cloned and characterized. The results indicated that both of the silenced 1By genes in AS332 and AS908 were 1Byg. In contrast to previously reported mechanisms for silenced genes lAx and lay, which was due to the insertion of transposon elements or the presence of premature stop codon via base substitution of C→T transition in trinucleotides CAA or CAG, the silence of 1By9 genes was caused by premature stop codons via the deletion of base A in trinucleotide CA.A, which lead to frameshift mutation and indirectly produced several premature stop codons （TAG） downstream of the coding sequence.     

An Improved Strategy for Efficient Expression and Purification of Soluble HIV-1 Tat Protein in E.coli

Although the endogenous function of Tat has been elucidated in the past twenty years, the study of its exogenous activity has been hampered due to the difficulty of large scale preparation of the active Tat protein. To express the full-length Tat protein in E.coli, the tat gene was cloned from an HIV infected patient by overlapping PCR. Rare codon usage analysis showed that rare E.coli codons, especially consecutive rare codons for Arg, account for 14% （14 of 101） rare E.coli codons in the tat gene. The expression of the HIV-1 tat gene was verified to be very poor in strain BL21 （DE3） due to the abundance of rare codons; however, tat gene expression was found to be very efficient in the host strain of Rosetta-gami B （DE3）, which was supplemented with six rare tRNAs for Arg, Leu, Ile and Pro. Subsequent purification revealed that the proteins are soluble and unusually, the tagged Tat can form dimers independent of cystine disulfide bonds. The purity, integrity and molecular weight of the Tat protein were demonstrated by MALDI-TOF mass spectrometry. Reporter gene activating assay was further confirmed by investigating the transactivation activity of the recombinant Tat protein. Our improved strategy for efficient high level expression and purification of soluble Tat protein has paved the way to fully investigate its exogenous function.     

Paramutation： A Heritable Change in Gene Expression by Allelic Interactions In Trans

Epigenetic gene regulation involves the stable propagation of gene activity states through mitotic, and sometimes even meiotic, cell divisions without changes in DNA sequence. Paramutation is an epigenetic phenomenon involving changes in gene expression that are stably transmitted through mitosis as well as meiosis. These heritable changes are mediated by in trans interactions between homologous DNA sequences on different chromosomes. During these in trans interactions, epigenetic information is transferred from one allele of a gene to another allele of the same gene, resulting in a change in gene expression. Although paramutation was initially discovered in plants, it has recently been observed in mammals as well, suggesting that the mechanisms underlying paramutation might be evolutionarily conserved. Recent findings point to a crucial role for small RNAs in the paramutation process. In mice, small RNAs appear sufficient to induce paramutation, whereas in maize, it seems not to be the only player in the process. In this review, potential mechanisms are discussed in relation to the various paramutation phenomena.     

Defining the Functional Network of Epigenetic Regulators in Arabidopsis thaliana

Development of ChiP-chip and ChlP-seq technologies has allowed genome-wide high-resolution profiling of chromatin-associated marks and binding sites for epigenetic regulators. However, signals for directing epigenetic modifiers to their target sites are not understood. In this paper, we tested the hypothesis that genome location can affect the involvement of epigenetic regulators using Chromatin Charting （CC） Lines, which have an identical transgene construct inserted at different locations in the Arabidopsis genome. Four CC lines that showed evidence for epigenetic silencing of the luciferase reporter gene were transformed with RNAi vectors individually targeting epigenetic regulators LHP1, MOM1, CMT3, DRD1, DRM2, SUVH2, CLF, and HD1. Involvement of a particular epigenetic regulator in silencing the transgene locus in a CC line was determined by significant alterations in luciferase expression after suppression of the regulator＇s expression. Our results suggest that the targeting of epigenetic regulators can be influenced by genome location as well as sequence context. In addition, the relative importance of an epigenetic regulator can be influenced by tissue identity. We also report a novel approach to predict interactions between epigenetic regulators through clustering analysis of the regulators using alterations in gene expression of putative downstream targets, including endogenous loci and transgenes, in epigenetic mutants or RNAi lines. Our data support the existence of a complex and dynamic network of epigenetic regulators that serves to coordinate and control global gene expression in higher plants.     

Feruloylation in Grasses： Current and Future Perspectives

In the cell walls of forage grasses, ferulic acid is esterified to arabinoxylans and participates with lignin monomers in oxidative coupling pathways to generate ferulate-polysaccharide-lignin complexes that cross-link the cell wall. The accumulation of ferulates and the cross-linking of arabinoxylans via diferulate esters are hypothesized to function in various processes in plants. The specific roles of arabinoxylan feruloylation as well as the nature, cellular localization, and substrate for arabinoxylans feruloylation of cell walls are reviewed. The various approaches that have been used for assessing the specific roles of feruloylation are described and assessed. I argue that, until recently, the specific role of feruloylation in these various processes has been established largely by indirect experiments and, although these studies reached similar conclusions about the potential importance of wall feruloylation, they suffer from a common problem： namely they depend on correlations between two processes and do not stem from a detailed understanding of the mechanisms of feruloylation. I also argue that the nature of arabinoxylan feruloylation remains uncertain.     

XPA A23G polymorphism is associated with the elevated response to platinum-based chemotherapy in advanced non-small cell lung cancer

DNA repair capacity （DRC） is correlated with sensitivity of cancer cells toward platinum-based chemotherapy. We hypothesize that genetic polymorphisms in DNA repair gene XPA （xeroderma pigmentosum group A） and XPG （xeroderma pigmentosum group G） （ERCC5, excision repair cross-complementation group 5）, which result in inter-individual differences in DNA repair efficiency, may predict clinical response to platinum agents in advanced non-small cell lung cancer （NSCLC） patients. In this study, we find that the A → G change of XPA A23G polymorphism significantly increased response to platinum-based chemotherapy. Polymorphism in XPG His46His was associated with a decreased treatment response, but was not statistically significant.