Overexpression of <Emphasis Type="Italic">PSP1</Emphasis> enhances growth of transgenic Arabidopsis plants under ambient air conditions

Cellulose biosynthesis is mediated by cellulose synthases (CesAs), which constitute into rosette-like cellulose synthase complexe (CSC) on the plasma membrane. Two types of CSCs in Arabidopsis are believed to be involved in cellulose synthesis in the primary cell wall and secondary cell walls, respectively. In this work, we found that the two type CSCs participated cellulose biosynthesis in differentiating xylem cells undergoing secondary cell wall thickening in Populus. During the cell wall thickening process, expression of one type CSC genes increased while expression of the other type CSC genes decreased. Suppression of different type CSC genes both affected the wall-thickening and disrupted the multilaminar structure of the secondary cell walls. When CesA7A was suppressed, crystalline cellulose content was reduced, which, however, showed an increase when CesA3D was suppressed. The CesA suppression also affected cellulose digestibility of the wood cell walls. The results suggest that two type CSCs are involved in coordinating the cellulose biosynthesis in formation of the multilaminar structure in Populus wood secondary cell walls.     

Regulation of secondary xylem formation in young hybrid poplars by modifying the expression levels of the <Emphasis Type="Italic">PtaGLIMa</Emphasis> gene

Due to the importance of wood in many industrial applications, a tremendous amount of research has focused on the regulation of secondary xylem formation and wood properties. In this study, we performed functional analysis of PtaGLIM1a, a LIM gene that is predominantly expressed in the differentiation of secondary xylem of the hybrid poplar (Populus tremula × P. alba). With no growth retardation, transgenic poplar plants with increased and reduced expression levels of PtaGlim1a exhibited enhanced and diminished secondary growth, respectively, accompanied by a corresponding change in their lignin abundance. This study demonstrates that the wood-associated PtaGlim1a acts as a positive regulator of secondary xylem formation in poplar trees and could potentially be utilized in modifying the synthesis of plant secondary wall lignin.     

Analysis of <Emphasis Type="Italic">Streptomyces coelicolor</Emphasis> M145 genes <Emphasis Type="Italic">SCO4164</Emphasis> and <Emphasis Type="Italic">SCO5854</Emphasis> encoding putative rhodaneses

Streptomyces coelicolor genome carries two apparently paralogous genes, SCO4164 and SCO5854, that encode putative thiosulfate sulfurtransferases (rhodaneses). These genes (and their presumed translation products) are highly conserved and widely distributed across actinobacterial genomes. The SCO4164 knockout strain was unable to grow on minimal media with either sulfate or sulfite as the sole sulfur source. The SCO5854 mutant had no growth defects in the presence of various sulfur sources; however, it produced significantly less amounts of actinorhodin. Furthermore, we discuss possible links between basic interconversions of inorganic sulfur species and secondary metabolism in S. coelicolor.     

Localization of gene expression,tissue specificity of <Emphasis Type="Italic">Populus</Emphasis> xylosyltransferase genes by isolation and functional characterization of their promoters

Efficient bioconversion of cellulose into glucose using lignocellulose as the feedstock requires improved cell wall traits. Xylan is part of the matrix covering cellulose and linkages of xylan and lignin inhibit cellulases access. Xylose, an uncommon sugar in the yeast fermentation process, represents 20% of the lignocellulose mass fraction, thus xylan related genes are important targets for biotechnology. Initially, the isolation of candidate genes and their functional characterization is a prerequisite. A common strategy is to perform exhaustive promoter characterizations for candidate genes. Here, we report on the characterization of two xylosyltransferases-related gene promoters that were isolated upstream of the respective candidate genes in poplar, a promising feedstock for bioenergy that is characterized by its short-rotation. The species is known to have undergone recent whole genome duplication, thus finding duplicated gene sequences for xylosyltransferases based on phylogeny reconstruction but with distinct expression patterns is expected. To investigate these two genes closely related to the single, well-characterised Arabidopsis thaliana (At) irx10 gene, we constructed two binary vectors, each studying the full-length promoter of the respective gene. For the poplar (Ptr) gene, most likely the functionally closest to the At irx10 gene, localization of gene expression was studied using the green fluorescence protein (GFP). Poplar’s glucuronoxylan glucuronosyltransferase protein, PtrGUT2B, expression was localized in the inflorescence, and specifically in the xylem and along the fibres, suggesting that PtrGUT2B gene expression is associated with fibre production similar to the At irx10 gene. Conversely, the second, duplicated, poplar gene termed PtrGUT2A showed expression in the cambium/phloem, vascular bundles, in the primary xylem and the central cylinder that actively undergo cell divisions and differentiation as evidenced by driving ß-glucuronidase (GUS) expression in these specific tissues. There was no staining detected in the mature secondary xylem or interfascicular fibres. Our study demonstrated that the PtrGUT2B is the true ortholog of irx10 and the two isolated promoters can be used to target tissue specific expression in plant biotechnology assays.     

Defence reactions in the apoplastic proteome of oilseed rape (<Emphasis Type="Italic">Brassica napus</Emphasis> var. <Emphasis Type="Italic">napus</Emphasis>) attenuate <Emphasis Type="Italic">Verticillium longisporum</Emphasis>growth but not disease symptoms

Background

Verticillium longisporum is one of the most important pathogens of Brassicaceae that remains strictly in the xylem during most stages of its development. It has been suggested that disease symptoms are associated with clogging of xylem vessels. The aim of our study was to investigate extracellular defence reactions induced by V. longisporum in the xylem sap and leaf apoplast of Brassica napus var. napus in relation to the development of disease symptoms, photosynthesis and nutrient status.

Results

V. longisporum (strain VL43) did not overcome the hypocotyl barrier until 3 weeks after infection although the plants showed massive stunting of the stem and mild leaf chlorosis. During this initial infection phase photosynthetic carbon assimilation, transpiration rate and nutrient elements in leaves were not affected in VL43-infected compared to non-infected plants. Proteome analysis of the leaf apoplast revealed 170 spots after 2-D-protein separation, of which 12 were significantly enhanced in response to VL43-infection. LS-MS/MS analysis and data base searches revealed matches of VL43-responsive proteins to an endochitinase, a peroxidase, a PR-4 protein and a β-1,3-glucanase. In xylem sap three up-regulated proteins were found of which two were identified as PR-4 and β-1,3-glucanase. Xylem sap of infected plants inhibited the growth of V. longisporum.

Conclusion

V. longisporum infection did not result in drought stress or nutrient limitations. Stunting and mild chlorosis were, therefore, not consequences of insufficient water and nutrient supply due to VL43-caused xylem obstruction. A distinct array of extracellular PR-proteins was activated that might have limited Verticillium spreading above the hypocotyl. In silico analysis suggested that ethylene was involved in up-regulating VL43-responsive proteins.

     

Identification of a novel <Emphasis Type="Italic">SPLIT</Emphasis>-<Emphasis Type="Italic">HULL</Emphasis> (<Emphasis Type="Italic">SPH</Emphasis>) gene associated with hull splitting in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Key message

The split-hull phenotype caused by reduced lemma width and low lignin content is under control of SPH encoding a type-2 13-lipoxygenase and contributes to high dehulling efficiency.

Abstract

Rice hulls consist of two bract-like structures, the lemma and palea. The hull is an important organ that helps to protect seeds from environmental stress, determines seed shape, and ensures grain filling. Achieving optimal hull size and morphology is beneficial for seed development. We characterized the split-hull (sph) mutant in rice, which exhibits hull splitting in the interlocking part between lemma and palea and/or the folded part of the lemma during the grain filling stage. Morphological and chemical analysis revealed that reduction in the width of the lemma and lignin content of the hull in the sph mutant might be the cause of hull splitting. Genetic analysis indicated that the mutant phenotype was controlled by a single recessive gene, sph (Os04g0447100), which encodes a type-2 13-lipoxygenase. SPH knockout and knockdown transgenic plants displayed the same split-hull phenotype as in the mutant. The sph mutant showed significantly higher linoleic and linolenic acid (substrates of lipoxygenase) contents in spikelets compared to the wild type. It is probably due to the genetic defect of SPH and subsequent decrease in lipoxygenase activity. In dehulling experiment, the sph mutant showed high dehulling efficiency even by a weak tearing force in a dehulling machine. Collectively, the results provide a basis for understanding of the functional role of lipoxygenase in structure and maintenance of hulls, and would facilitate breeding of easy-dehulling rice.

     

Deletion of <Emphasis Type="Italic">ldh</Emphasis>A and <Emphasis Type="Italic">ald</Emphasis>H genes in <Emphasis Type="Italic">Klebsiella</Emphasis><Emphasis Type="Italic">pneumoniae</Emphasis> to enhance 1,3-propanediol production

Objectives

To improve 1,3-propanediol (1,3-PD) production and reduce byproduct concentration during the fermentation of Klebsiella pneumonia.

Results

Klebsiella. pneumonia 2-1ΔldhA, K. pneumonia 2-1ΔaldH and K. pneumonia 2-1ΔldhAΔaldH mutant strains were obtained through deletion of the ldhA gene encoding lactate dehydrogenase required for lactate synthesis and the aldH gene encoding acetaldehyde dehydrogenase involved in the synthesis of ethanol. After fed-batch fermentation, the production of 1,3-PD from glycerol was enhanced and the concentrations of byproducts were reduced compared with the original strain K. pneumonia 2-1. The maximum yields of 1,3-PD were 85.7, 82.5 and 87.5 g/l in the respective mutant strains.

Conclusion

Deletion of either aldH or ldhA promoted 1,3-PD production in K. pneumonia.

     

Role of Arabidopsis <Emphasis Type="Italic">ABF1</Emphasis>/<Emphasis Type="Italic">3</Emphasis>/<Emphasis Type="Italic">4</Emphasis> during <Emphasis Type="Italic">det1</Emphasis> germination in salt and osmotic stress conditions

Key message

Arabidopsis det1 mutants exhibit salt and osmotic stress resistant germination. This phenotype requires HY5, ABF1, ABF3, and ABF4.

Abstract

While DE-ETIOLATED 1 (DET1) is well known as a negative regulator of light development, here we describe how det1 mutants also exhibit altered responses to salt and osmotic stress, specifically salt and mannitol resistant germination. LONG HYPOCOTYL 5 (HY5) positively regulates both light and abscisic acid (ABA) signalling. We found that hy5 suppressed the det1 salt and mannitol resistant germination phenotype, thus, det1 stress resistant germination requires HY5. We then queried publically available microarray datasets to identify genes downstream of HY5 that were differentially expressed in det1 mutants. Our analysis revealed that ABA regulated genes, including ABA RESPONSIVE ELEMENT BINDING FACTOR 3 (ABF3), are downregulated in det1 seedlings. We found that ABF3 is induced by salt in wildtype seeds, while homologues ABF4 and ABF1 are repressed, and all three genes are underexpressed in det1 seeds. We then investigated the role of ABF3, ABF4, and ABF1 in det1 phenotypes. Double mutant analysis showed that abf3, abf4, and abf1 all suppress the det1 salt/osmotic stress resistant germination phenotype. In addition, abf1 suppressed det1 rapid water loss and open stomata phenotypes. Thus interactions between ABF genes contribute to det1 salt/osmotic stress response phenotypes.

     

<Emphasis Type="Italic">Quinua</Emphasis> and Relatives (<Emphasis Type="Italic">Chenopodium</Emphasis> sect.<Emphasis Type="Italic">Chenopodium</Emphasis> subsect.<Emphasis Type="Italic">Celluloid</Emphasis>)

Traditionally viewed as an Andean grain crop,Chenopodium quinoa Willd. includes domesticated populations that are not Andean, and Andean populations that are not domesticated. Comparative analysis of leaf morphology and allozyme frequencies have demonstrated that Andean populations, both domesticated(quinua) and free-living(ajara), represent an exceptionally homogeneous unit that is well differentiated from allied domesticates of coastal Chile(quingua) and freeliving populations of the Argentine lowlands(C. hircinum). This pattern of relationships indicates that Andean populations represent a monophyletic crop/weed system that has possibly developed through cyclic differentiation (natural vs. human selection) and introgressive hybridization. Relative levels of variation suggest that this complex originated in the southern Andes, possibly from wild types allied withC. hircinum, with subsequent dispersal north to Colombia and south to the Chilean coast. Coastal populations were apparently isolated from post-dispersal differentiation and homogenization that occurred in the Andes. Other data point toward a center of origin in the northern Andes with secondary centers of genetic diversity subsequently developing in the southern Andes and the plains of Argentina. Comparative linkage of South American taxa, all tetraploid, with North American tetraploids of the subsection will eventually clarify this problem. While the possibility of a direct phyletic connection betweenC. quinoa and the Mexican domesticate(C. berlandieri subsp. nuttalliae,) cannot be excluded, available evidence indicates that the latter represents an autonomous lineage that is associated with the basal tetraploid, C. b. subsp.berlandieri, through var.sinuatum, whereas South American taxa show possible affinities to either var. zschackei or var.berlandieri. An extinct domesticate of eastern North America,C. b. subsp.jonesianum, represents either another instance of independent domestication, possibly from subsp. b. var.zschackei, or a northeastern outlier of subsp.nuttalliae.     

<Emphasis Type="Italic">R</Emphasis>-acetoin accumulation and dissimilation in <Emphasis Type="Italic">Klebsiella pneumoniae</Emphasis>

Klebsiella pneumoniae is a 2,3-butanediol producer, and R-acetoin is an intermediate of 2,3-butanediol production. R-acetoin accumulation and dissimilation in K. pneumoniae was studied here. A budC mutant, which has lost 2,3-butanediol dehydrogenase activity, accumulated high levels of R-acetoin in culture broth. However, after glucose was exhausted, the accumulated R-acetoin could be reused by the cells as a carbon source. Acetoin dehydrogenase enzyme system, encoded by acoABCD, was responsible for R-acetoin dissimilation. acoABCD mutants lost the ability to grow on acetoin as the sole carbon source, and the acetoin accumulated could not be dissimilated. However, in the presence of another carbon source, the acetoin accumulated in broth of acoABCD mutants was converted to 2,3-butanediol. Parameters of R-acetoin production by budC mutants were optimized in batch culture. Aerobic culture and mildly acidic conditions (pH 6–6.5) favored R-acetoin accumulation. At the optimized conditions, in fed-batch fermentation, 62.3 g/L R-acetoin was produced by budC and acoABCD double mutant in 57 h culture, with an optical purity of 98.0 %, and a substrate conversion ratio of 28.7 %.     

The <Emphasis Type="Italic">angora</Emphasis> gene weakens the effect of interaction of the mutant genes <Emphasis Type="Italic">wellhaarig</Emphasis> and <Emphasis Type="Italic">waved alopecia</Emphasis> in mice

The interaction of the mutant genes wellhaarig (we) and waved alopecia (wal) in mice was earlier demonstrated in our laboratory. The we gene significantly accelerates the appearance of alopecia in double we/wewal/wal homozygotes as compared to that in single +/+wal/wal homozygotes. It has been found in this work that the mutant gene angora-Y (Fgf5 ^go-Y ) weakens the effect of interaction of the we and wal genes. The first signs of alopecia appear in mice of the we/wewal/wal genotype at the age of 14 days, in triple Fgf5 ^go-Y /Fgf5 ^go-Y we/wewal/wal homozygotes alopecia is observed seven days later, i. e., in 21-day-old animals. The progression of alopecia in triple homozygotes is expressed to a lesser degree than in double +/+we/wewal/wal homozygotes. A single dose of the Fgf5 ^go-Y gene also decreases the effect of interaction of the we and wal genes, but less than a double dose of this gene. The first signs of alopecia in mice of the +/Fgf5 ^go-Y we/wewal/wal genotype appear only three days later than in double +/+we/wewal/wal homozygotes. The data obtained demonstrate that the Fgf5 ^go-Y gene is a powerful modifier of mutant genes determining the process of alopecia.     

The <Emphasis Type="Italic">Adh</Emphasis> locus and adaptation of <Emphasis Type="Italic">cn</Emphasis> and <Emphasis Type="Italic">vg</Emphasis> mutants in experimental populations of <Emphasis Type="Italic">Drosophila melanogaster MEIG</Emphasis>

A complex study on the adaptation of cn and vn mutants and the allozymes of alcoholdehydrogenase (ADH) was carried out in initially pure lines, and their panmixia populations during exchange of the mutant genotype with that of wild-type flies (C-S) and D) through saturating crossings. The relative adaptation of the genotypes was estimated by their effect on reproductive efficiency in the experimentally obtained population. Fecundity, lifespan, and the resistance of the studied genotypes to hyperthermia were investigated individually. It was shown that the high level of adaptation of the cn mutants and the low level of adaptation of the vg mutants was correlated with the presence of different ADH allozymes. In the studied population, the F-allozyme of ADH accompanied the vg mutation, while the S-allozyme of the enzyme was detected in cn mutants. Saturating crossings of C-S(Adh ^S)×vg(Adh ^F) and D(Adh ^F) × cn(Adh ^S), along with the parallel determination of the allele composition of the Adh locus, demonstrated that the complete substitution of the F-allozyme of ADH in the vg mutants by the S-allozyme in D flies, as well as the substitution of the S-allozyme of ADH in the cn mutants by the F-allozyme in D flies was realized only after the 15th–20th backcrosses. These results favor the coadaptation of cn and vg marker genes with alleles of the Adh locus and indicate the important role of the latter in the adaptation of genotypes. In the studied population, selection acted primarily against the vg mutants, which were inferior to the cn mutants, and heterozygote genotypes in indices of the main adaptation components.     

Characterization and fine mapping of <Emphasis Type="Italic">osh15</Emphasis>(<Emphasis Type="Italic">t</Emphasis>), a novel dwarf mutant gene in rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.)

Plant height is one of the most important agronomic traits of plant architecture, and also affects grain yield in rice. In this study, we obtained a novel dwarf rice mutant of japonica variety Shennong9816, designated Shennong9816d. Compared with wild-type, the Shennong9816d plant height was significantly reduced, and the tiller number significantly increased. Additionally, the mutant yield component, and the number of large and small vascular bundles were significantly decreased compared with wild-type. Genetic analysis indicated that the Shennong9816d dwarf phenotype was controlled by a recessive nuclear gene, while the plant was shown to be sensitive to gibberellic acid. Using a large F₂ population derived from a cross between Shennong9816d and the indica rice variety Habataki, the osh15(t) gene was fine mapped between RM20891 and RM20898, within a physical distance of 73.78 kb. Sequencing analysis showed that Shennong9816d carries a 1 bp mutation and a 30 bp insertion in the OSH15 region. These results suggest that osh15(t) is a novel allelic mutant originally derived from japonica variety Shennong9816, which may be useful for introducing the semi-dwarf phenotype to improve plant architecture in rice breeding practice.