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SERK and APOSTART. Candidate genes for apomixis in Poa pratensis.

Seed production generally requires the mating of opposite sex gametes. Apomixis, an asexual mode of reproduction, avoids both meiotic reduction and egg fertilization. The essential feature of apomixis is that an embryo is formed autonomously by parthenogenesis from an unreduced egg of an embryo sac generated through apomeiosis. If apomixis were well understood and harnessed, it could be exploited to indefinitely propagate superior hybrids or specific genotypes bearing complex gene sets. A more profound knowledge of the mechanisms that regulate reproductive events would contribute fundamentally to understanding the genetic control of the apomictic pathway. In Poa pratensis, we isolated and characterized two genes, PpSERK (SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE) and APOSTART. These full-length genes were recovered by rapid amplification of cDNA ends and their temporal and spatial expression patterns were assessed by reverse transcription-polymerase chain reaction and in situ hybridization, respectively. The expression of PpSERK and APOSTART differed in apomictic and sexual genotypes. Their putative role in cell-signaling transduction cascades and trafficking events required during sporogenesis, gametogenesis, and embryogenesis in plants is reported and discussed. We propose that, in nucellar cells of apomictic genotypes, PpSERK is the switch that channels embryo sac development and that it may also redirect signaling gene products to compartments other than their typical ones. The involvement of APOSTART in meiosis and programmed cell death is also discussed.

Amino Acid Sequence↗

The Inheritance of apomixis in Poa pratensis confirms a five locus model with differences in gene expressivity and penetrance.

The genetic control of apomixis was studied in numerous segregating progenies originated from intercrossing and selfing of obligate sexual and facultative apomictic parents in Poa pratensis by means of the flow cytometric seed screen. The data support a novel model with five major genes required to control asexual seed formation: the Apospory initiator (Ait) gene, the Apospory preventer (Apv) gene, a Megaspore development (Mdv) gene, the Parthenogenesis initiator (Pit) gene, and the Parthenogenesis preventer (Ppv) gene. Differences in expressivity and interactions of these genes are responsible for the wide variation of the mode of reproduction. Apospory and parthenogenesis as well as the initiator and preventer genes of these components segregate independently. The genotypes with the highest expressivity of apospory and parthenogenesis were assigned as Ait-/apvapv/Pit-/ppvppv, those with intermediate expressivity as Ait-/Apv-/Pit-/Ppv-, and those with low expressivity as aitait/apvapv/pitpit/ppvppv. Among the self progenies of obligate sexual individuals, plants with a low capacity for apospory and/or parthenogenesis occurred, indicating that the sexual parents were heterozygous for the preventer genes and homozygous for the recessive initiator alleles (aitait/Apv-/pitpit/Ppv-). The dominant allele Ait exhibits incomplete penetrance. The degree of expressivity of apospory and parthenogenesis was constant among several harvest years of F1 plants.

Gene Expression Regulation, Plant↗

Microbial succession of nitrate-reducing bacteria in the rhizosphere of Poa alpina across a glacier foreland in the Central Alps.

Changes in community structure and activity of the dissimilatory nitrate-reducing community were investigated across a glacier foreland in the Central Alps to gain insight into the successional pattern of this functional group and the driving environmental factors. Bulk soil and rhizosphere soil of Poa alpina was sampled in five replicates in August during the flowering stage and in September after the first snowfalls along a gradient from 25 to 129 years after deglaciation and at a reference site outside the glacier foreland (>2000 years deglaciated). In a laboratory-based assay, nitrate reductase activity was determined colorimetrically after 24 h of anaerobic incubation. In selected rhizosphere soil samples, the community structure of nitrate-reducing microorganisms was analysed by restriction fragment length polymorphism (RFLP) analysis using degenerate primers for the narG gene encoding the active site of the membrane-bound nitrate reductase. Clone libraries of the early (25 years) and late (129 years) succession were constructed and representative clones sequenced. The activity of the nitrate-reducing community increased significantly with age mainly due to higher carbon and nitrate availability in the late succession. The community structure, however, only showed a small shift over the 100 years of soil formation with pH explaining a major part (19%) of the observed variance. Clone library analysis of the early and late succession pointed to a trend of declining diversity with progressing age. Presumably, the pressure of competition on the nitrate reducers was relatively low in the early successional stage due to minor densities of microorganisms compared with the late stage; hence, a higher diversity could persist in this sparse environment. These results suggest that the nitrate reductase activity is regulated by environmental factors other than those shaping the genetic structure of the nitrate-reducing community.

Bacteria↗

[Variation in seed yield components among varieties of Poa pratensis].

Seed yield and its characteristics of 28 varieties of Poa pratensis in field plots at Lanzhou, China were evaluated in 2001. The results showed that the seed yields among the varieties were significantly different (P < 0.01) by LSD test. The highest yield was 643.4-650.2 kg.hm-2 obtained from varieties of Balin and Monoply, the lowest was 90.1 kg.hm-2 obtained from Elysee, and the other varieties were in the middle groups with the yields varied from 114.7 kg.hm-2 to 435.1 kg.hm-2. Correlation analysis between seed yield and yielding characteristics indicated that the main factors affecting seed yield were the number of fertile tillers per plant (r = 0.785), the height of fertile tiller (r = 0.712), the length of inflorescence (r = 0.743), and the number of spikelets/fertile tiller (r = 0.786). Cluster analysis for seed production capacity based on seed yield and above-mentioned 4 main characteristics showed that the 28 varieties could be divided into 6 grades.

Cluster Analysis↗

[Identification of the apomixis in Poa pratensis L. using SDS-electrophoresis of endosperm reserve proteins].

To determine the characteristics of variability of seed reproduction in the Kentucky bluegrass (Poa pratensis), individual seed variability with respect to the composition of endospermal reserve proteins was studied. Comparative analysis of caryopses obtained by self-fertilization and free fertilization of plants I1 (no. 2-4) and I2 (no. 2-4-7) of the wild-type specimen Murmanskii-95 was performed using SDS-PAGE. Using a cytoembryological express method, we demonstrated that facultative stimulation-autonomous apomeiotic apomixis, along with the formation of meiotic megasporocytes, is characteristic of the Kentucky bluegrass. This method made it possible to determine the consequences of meiotic processes in the maternal plant and to reveal the hybrid nature of seed endosperm.

Crosses, Genetic↗

Effects of traffic pollution on the genetic structure of Poa annua L. populations.

The genetic composition of Poa annua L. populations with a series of traffic pollution was studied by starch electrophoresis. Five enzyme systems were stained. The results showed that: (1) Traffic pollution can dramatically change genotypic frequencies at some loci of P. annua populations. Significant deviations from Hardy-Weinberg equilibrium were observed on loci Fe-1 and Me due to the excess of heterozygotes in some populations. (2) The effective number of alleles per locus and the observed and expected heterozygosity were higher in the pollution series than in the clear control site (Botanic Park population), but the increase was not related with the pollution extent. (3) Most genetic variation was found within populations, and only 6.21% was among populations of the polluted series. Slightly higher differentiation (FST = 7.98%) was observed when the control population was included. (4) The calculated gene flow (Nm) is 2.8841 per generation. The mean of genetic identity is 0.9864 and the genetic distance average to 0.0138.

Air Pollutants↗

No difference in activity of Sod-1 genotypes of Poa annua to short-term treatment of ambient gaseous organic pollution.

Our previous studies indicated that genotypes at locus Sod-1 of Poa annua changed clinally along a gradient of gaseous organic pollution. In the present study, we aimed to know whether there were differential responses of activities of different superoxide dismutase (SOD) genotypes to short-term treatment of ambient gaseous organic pollution. Significant bias from Hardy-Weinberg equibrium was observed on locus Sod-1, and no genotype Sod-1-BB was found. Significantly increased activities were observed in most treatments for genotype Sod-1-AA and in one treatment for genotype Sod-1-AB. However, no significant difference in SOD activities was found between the two genotypes. It was interpreted that fitness difference between the two genotypes was small and treatment duration was too short or extent of ambient organic pollution was too low to lead to differential responses. Other environmental factor effects on activities of superoxide dismutases can also explain the results.

Air Pollutants↗

Development of a field bioassay for the side-effects of herbicides on vascular plants using Brassica napus and Poa annua.

A field bioassay method has been developed for assessing side-effects of herbicides on Brassica napus and Poa annua. The test method aims at effects outside the target area, up to 20 m from the treated parcel. Experiments were conducted in the field and in a glasshouse with the herbicides atrazin, bentazone, diquat, glyphosate, and r(4-chloro-2-methylphenoxyacetic acid) MCPA. An iterative procedure was used with the aim of developing a sound test procedure. The resulting method appeared to be very sensitive. Pesticide drift was measured using water-sensitive paper. Effects were sometimes found below the detection limit of this method (at 8 m from the treated parcel with moderate wind speeds of 3-5 m/s). The sensitivity of the test and the test conditions are discussed and an optimized method is presented.

Air Movements↗

Comparisons of the genomic cis-elements and coding regions in RNA beta components of the hordeiviruses barley stripe mosaic virus, lychnis ringspot virus, and poa semilatent virus.

Nucleotide sequences of the genomic RNA beta components of hordeiviruses poa semilatent virus (PSLV) and lychnis ringspot virus (LRSV) were determined. PSLV and LRSV closely resemble barley stripe mosaic virus (BSMV), type hordeivirus, in the gene arrangement of their RNAs beta, comprising 5'-proximal beta a (coat protein) gene and downstream triple gene block (TGB) coding for the beta b, beta c, and beta d putative transport proteins. The beta a, beta b, beta c, and beta d proteins of the three hordeiviruses showed significant sequence similarity, with the respective proteins of PSLV and BSMV being closer to each other than to their counterparts of LSRV. Comparisons of the TGB-encoded proteins of hordeiviruses, potexviruses, carlaviruses, and furoviruses indicate that the first and second TGB genes belong to the monophyletic groups, whereas the third gene may have multiple ancestry. LRSV, PSLV, and BSMV showed remarkable variation in the 3'-untranslated regions of their genomic RNAs. Among the three hordeiviruses, LRSV has the shortest 3'-noncoding region that lacks tentative pseudoknot-forming elements conserved upstream of the 3'-tRNA-like structure in the BSMV and PSLV genomes. On the other hand, LRSV RNA beta, like that of BSMV, contained the internal poly(A) sequence that is absent from PSLV RNA.

Adaptation, Physiological↗

Tremorgenic mycotoxins produced by strains of Penicillium spp. isolated from toxic Poa huecu parodi.

Seventeen strains of Penicillium spp. have been isolated from Poa huecu Parodi from the Zapala zone, exhibiting toxicity to sheet. The following strains have been identified: P. crustosum, cyclopium, notatum, palitans, puberulum, verrucosum, viridicatum and Penicillium spp. The toxigenic capacity of the strains was studied after growing them under suitable conditions. Toxins produced were analysed by thin layer chromatography (TLC). Penitrem A (PA) and Penitrem B (PB) neurotoxins were identified and quantitated in twelve strains; verruculogen (VERR) and fumitremorgen B (FTB) being present in one of them. The effect of these mycotoxins was studied in mice. Neurological symptoms characteristic of the intoxication by tremorgenic toxins and similar to those observed in sheep suffering from 'huecu's disease' were observed. The possible role of these toxins as causative agents of 'huecu's disease' is discussed.

Animals↗

Linkage mapping in apomictic and sexual Kentucky bluegrass ( Poa pratensis L.) genotypes using a two way pseudo-testcross strategy based on AFLP and SAMPL markers.

The high versatility of the mode of reproduction and the retention of a pollen recognition system are the factors responsible for the extreme complexity of the genome in Poa pratensis L. Two genetic maps, one of an apomictic and one of a sexual genotype, were constructed using a two-way pseudo-testcross strategy and multiplex PCR-based molecular markers (AFLP and SAMPL). Due to the high ploidy level and the uncertainty of chromosome pairing-behavior at meiosis, only parent-specific single-dose markers (SDMs) that segregated 1:1 in an F(1) mapping population (161 out of 299 SAMPLs, and 70 out of 275 AFLPs) were used for linkage analysis. A total of 41 paternal (33 SAMPLs and 8 AFLPs) and 47 maternal (33 SAMPLs and 14 AFLPs) SDMs, tested to be linked in coupling phase, were mapped to 7+7 linkage groups covering 367 and 338.4 cM, respectively. The comparison between the two marker systems revealed that SAMPL markers were statistically more efficient than AFLP ones in detecting parent-specific SDMs (75% vs 32.4%). There were no significant differences in the percentages of distorted marker alleles detected by the two marker systems (27.8% of SAMPLs vs 21.3% of AFLPs). The pairwise comparison of co-segregational groups for linkage detection between marker loci suggested that at least some of the P. pratensis chromosomes pair preferentially at meiosis-I.

Journal Article↗

Complete genome sequence of the first alphahypovirus from the phytopathogenic fungus Curvularia americana on Poa pratensis in China.

In the present study, a novel positive-sense single-stranded RNA virus was identified from Curvularia americana strain 32&#x2009;-&#x2009;2, a phytopathogenic fungus infecting Poa pratensis in China, which was designated as "Curvularia americana hypovirus 1" (CaHV1). The complete genome of CaHV1 consists of 14,386 nucleotides (nt) with a single large open reading frame (ORF) (nt position 593 to 14035). The ORF encodes a putative polyprotein (4,480 amino acids) containing two potential conserved domains, which includes a Domain of Unknown Function 3525 (DUF3525) and Helicase. BLASTp analysis showed that amino acid sequence of the polyprotein of CaHV1 shared the highest amino acid sequence identity with that of Bipolaris oryzae hypovirus 1 (63.52%). Phylogenetic analysis based on amino acid sequence of the polyprotein indicated that CaHV1 clustered together with members of the genus Alphahypovirus within the family Hypoviridae. To the best of our knowledge, this is the first report of an alphahypovirus infecting C. americana.

Genome, Viral↗

Genome sequences of poa semilatent and lychnis ringspot hordeiviruses.

The nucleotide sequences of the genomic RNAs of two hordeiviruses, poa semilatent virus (PSLV) and lychnis ringspot virus (LRSV), were determined. The genome organization of both viruses is similar to that of the type hordeivirus, barley stripe mosaic virus (BSMV). Comparisons of the amino acid sequences of the hordeivirus replicase components revealed a high level of sequence similarity to those of soil-borne wheat mosaic virus (SBWMV), peanut clump virus (PCV), and Indian peanut clump virus. The cysteine-rich gamma b proteins of hordeiviruses are similar to those of SBWMV, PCV, and tobraviruses. Analysis of cis-acting elements in the genomes of the three hordeiviruses revealed conservation of putative core elements of subgenomic RNA promoters. Complex patterns of conservation were also found in the 5'-untranslated regions of hordeivirus genomic RNAs alpha, beta, and gamma.

Amino Acid Sequence↗

The lethal principle of Poa huecu (coiron blanco): a plant indigenous to Argentina.

The lethality of Poa huecu, a plant toxic to cattle and sheep, was followed by injection of chromatographic fractions in mice. The lethal aqueous extract was administered i.p. to Rockland mice of either sex and produced motor incoordination, transient ataxia, rough hair coat, tremors and muscle contractions and, occasionally, blindness. Doses greater than 1.5 g/kg mouse were always lethal. Fractionation of this lethal extract included dialysis, column chromatography on Sephadex G-25 and fractional precipitation with ethanol. Precipitates obtained with 70% and 85% ethanol were further purified on a DEAE-cellulose column. Eight fractions were obtained, each was injected into mice. Only fractions 3-6 were toxic. Fraction 3 produced slight hepatosis and hyperemia in the liver and gliosis in the brain. None of the other tissues exhibited histological lesions. Fractions 4 and 5 caused death of all animals within 30 min to 4 hr after injection. Polyacrylamide gel electrophoresis and acid hydrolysis showed that fractions 4 and 5 contained a glycoprotein of nearly the same mol. wt (67,000-94,000). Microscopic pathology in the mice treated with the lethal glycoprotein of fraction 4 included hyperemia in the kidneys, megakaryocytes in the spleen, slight hepatosis and focal coagulative necrosis with nuclear pyknosis and karyonexis in the liver, gliosis, intracellular brain edema with axon degeneration and swollen astrocytes in the brain. These brain injuries may relate to the motor incoordination of cattle that causes a delayed righting reflex. The major monosaccharides of the lethal glycoprotein are glucose and mannose, while rhamnose, arabinose, xylose and galactose are present in low percentages. Proline and the acidic amino acids (glutamic and aspartic acids) are the most abundant in the peptidic residue.

Amino Acids↗

Mapping of antibody binding epitopes of a recombinant Poa p IX allergen.

Antibody binding epitopes of a recombinant Poa p IX allergen were delineated using recombinant DNA and solid-phase peptide synthesis procedures. The full-length cDNA clone KBG60 and its four overlapping recombinant fragments, KBG60.1, KBG60.2, KBG8.3 and KBG10 which spanned the entire molecule were synthesized in E. coli with aid of the plasmid expression vector, pWR590.1. The antigenic and allergenic sites of these recombinant proteins were analyzed by ELISA using human IgE and murine IgG antibodies. It was thus demonstrated that although the epitopes were found on all the fragments tested, the majority of these were located on a C-terminal fragment, rKBG8.3. Furthermore, synthetic peptides were also employed to identify the epitopes of rKBG60 protein. The use of antisera raised against native KBG pollen extract and the recombinant KBG8.3 protein to scan a total of 56 overlapping deca-penta peptides, covering the entire rKBG60 protein, revealed that 10 positive peptides involved in the antibody-binding site(s). Taken together, the results of these studies indicate that rKBG60 protein possesses at least 10 antibody binding epitopes.

Allergens↗

Effects of carbon dioxide and nitrogen fertilization on phenolic content in Poa annua L.

Different but partially overlapping hypotheses have been developed to predict the allocation of phenolics in elevated atmospheric CO(2). The carbon-nutrient balance hypothesis predicts increased allocation to phenolics due to reduced relative availability of nitrogen. The growth-differentiation balance hypothesis states that allocation will depend on source and sink strength, while the protein competition model predicts that allocation will remain unchanged. We grew Poa annua at two CO(2) concentrations in soils of three different nutrient levels. Although plant-tissue nitrogen levels were reduced in high CO(2) and photosynthetic rate increased, phenolic concentration and biomass allocation remained unchanged. We discuss these data in the context of the three models' predictions of phenolic allocation in conditions of elevated CO(2).

Journal Article↗

The alternative oxidase in roots of poa annua after transfer from high-light to low-light conditions.

The activity of the alternative pathway can be affected by a number of factors, including the amount and reduction state of the alternative oxidase protein, and the reduction state of the ubiquinone pool. To investigate the importance of these factors in vivo, we manipulated the rate of root respiration by transferring the annual grass Poa annua L. from high-light to low-light conditions, and at the same time from long-day to short-day conditions for four days. As a result of the low-light treatment, the total respiration rate of the roots decreased by 45%, in vitro cytochrome c oxidase capacity decreased by 49%, sugar concentration decreased by 90% and the ubiquinone concentration increased by 31%, relative to control values. The absolute rate of oxygen uptake via the alternative pathway, as determined using the 18O-isotope fractionation technique, did not change. Conversely, the cytochrome pathway activity decreased during the low-light treatment; its activity increased upon addition of exogenous sugars to the roots. Interestingly, no change was observed in the concentration of the alternative oxidase protein or in the reduction state of the protein. Also, there was no change in the reduction state of the ubiquinone pool. In conclusion, the concentration and activity of the alternative oxidase were not changed, even under severe light deprivation.

Carbohydrate Metabolism↗

The influence of secondary senescence processes within the culm of a pseudoviviparous grass (Poa alpina var. vivipara L.) on the supply of water to propagules.

An anatomical investigation of the culm of pseudoviviparous alpine meadow grass (Poa alpina var. vivipara L.) revealed that transpiration flow, as delimited by Lucifer Yellow tracer dye, was maintained despite advanced senescence (as evidenced by loss of chlorophyll and chloroplasts), with leafy spikelets driving transpiration flow. Transpiration flow was not hindered by cavitation or tylosis in older culms, the low frequencies of these senescence processes being bypassed via nodal plexi. Despite this, water content of plantlets declined over time and water stress became apparent, suggesting that water supply via the determinate culm was not sufficient for the increasing transpirational demand of indeterminate plantlets. The implications of declining water content on the biomechanical properties of the culm, and concomitant limitations on the pseudoviviparous reproductive strategy, are discussed. Nomenclature of grass follows Hubbard.

Fluorescent Dyes↗