RETICULATE EVOLUTION IN PHLOX AS SEEN THROUGH PROTEIN ELECTROPHORESIS

Seed protein profiles were used to document the genome constitutions of seven Phlox taxa believed to be stabilized derivatives of hybridization between P. pilosa and P. carolina, P. drummondii or P. amoena. These species have very distinctive seed protein profiles. Analyses of profiles from these and other phloxes not involved in the reticulum (P. maculata, P. glaberrima and their hybrid derivative P. maculata subsp. pyramidalis) demonstrated that profile components are inherited as co-dominant units, alteration of ploidal level does not alter the profile, profile components may undergo segregation and recombination, and profile components in hybrids do not re-combine in vivo or in vitro to produce new proteins. The putative derivatives of hybridization between P. pilosa and P. carolin, i.e., P. pilosa subsp. detonsa, P. pilosa subsp. sangamonensis, P. floridana and P. pulcherrima, display protein profiles which are nearly summations of the parental patterns as do the putative derivatives of P. pilosa and P. drummondii. i.e., P. villosissima and P. aspera. The putative derivative of P. pilosa and P. amoena, i.e., P. pilosa subsp. deamii, displays a completely additive profile. Genome constitutions judged from morphological, cytological, and chromatographical data are fully corroborated by the protein profiles.     

Dynamics of buried seed population and seedling cohorts of two dominant weeds in a hill agroecosystem of the humid subtropics of India

Dynamics of the buried seeds and plant population of two dominant weeds, viz.,Emilia sonchifolia (Linn.) DC. andRichardsonia pilosa HBK were studied in the crop fields of Meghalaya, north-east India during radish and maize cropping and intervening fallow periods. The total buried seed population ofR. pilosa was always larger than that ofE. sonchifolia, but the germinable fraction was invariably greater in the latter. A major portion (39–41%) of the viable (germinable+dormant) seed population in both weeds was confined to the surface soil layer (0–5 cm). The viable seed population ofE. sonchifolia peaked during April, while that ofR. pilosa showed two peaks (during August and December). The survival pattern and half-lives of seedling cohorts showed, some differences in the two weed species, but both being summer annuals, their populations behaved in a similar manner by showing higher seedling recruitment (K) and survivorship (p) rates in the summer crop (maize) than in the winter crop (radish). However, the density of plants that could attain adulthood was significantly higher inE. sonchifolia thanR. pilosa which might have resulted in greater seed input of the former to the soil leading to its greater abundance in the crop fields.     

Proteomics Analysis of Antitumor Activity of Agrimonia pilosa Ledeb. in Human Oral Squamous Cell Carcinoma Cells

Oral cancer is a malignant neoplasm of oral cavity. It accounts for approximately 5% of all malignant tumors. Approximately 97% of all oral cancers are squamous cell carcinomas, followed by adenocarcinomas, and rarely malignant melanomas. It occurs particularly in males (twice as common in males than in females) of middle age (above 40 years). Agrimonia pilosa Ledeb. has traditionally been known for its effective antitumor activity and is currently used in China for cancer therapy. A. pilosa Ledeb. has been traditionally used for the treatment of abdominal pain, sore throat, headache, blood discharge, parasitic infections, and eczema in Korea and other Asian countries. Most studies on A. pilosa Ledeb. are related to the leaves and a few investigated the roots of the plant. However, detailed mechanisms of antitumor activity of A. pilosa Ledeb. have not been fully elucidated. Furthermore, to date, there have been no reports on the antitumor effect of A. pilosa Ledeb. in oral squamous cells. In this study, we used proteomic technology to observe changes in proteins related to anticancer activity of A. pilosa Ledeb. and identified target proteins among altered proteins to reveal the underlying mechanism of action.     

Secondary intergradation and genome incompatibility in Phlox pilosa (polemoniaceae)

Phlox pilosa ssp.pilosa andP. pilosa ssp.fulgida form a zone of secondary intergradation extending from northern Illinois and Indiana along the prairie-forest border to southwestern Missouri. The structure of the zone was analyzed in Illinois and Indiana. Trichome morphology offered the only diagnostic characteristic. Character gradients in the zone are highly irregular and steeply sloping. Closely adjacent populations often have disparate phenotype distributions. Most populations are strongly skewed toward the parental ends of the morphological spectrum. The properties of the zone are attributed to genome incompatibility, which is seen in the crossability barriers between the two subspecies and between the subspecies and hybrids, a reduction in pollen fertility, anther size and fecundity in hybrids, and the formation of multiple nucleoli in the hybrids. Subspeciesfulgida appears to be the primary recipient of alien gene flow. However, the position of the zone ostensibly is stabilized by the presence of an ecological barrier.     

Notes on the ecology and distribution ofBidens pilosa L. in Cuba

The authors give the known distribution ofBidens pilosa L. in Cuba, discuss some features of its ecology, its diaspore production, and enumerate its parasites known in Cuba.     

Population size, pollination and reproductive success in two endangered Genista species

Habitat fragmentation often leads to small and isolated plant populations as well as decreased habitat quality. These processes can fundamentally disrupt the interactions between plants and pollinators and decrease reproductive success. This concerns especially self-incompatible, non-clonal species that depend on pollination for successful reproduction.In two rare and endangered heathland plant species, Genista anglica and G. pilosa, we examined pollination and reproduction in relation to population size. Eight populations of G. anglica and ten populations of G. pilosa were surveyed in the vicinity of Bremen, NW-Germany. We counted the visits of pollinators (honeybees, bumblebees, and other insects) and determined the reproductive output of the observed shoots.Contrary to our expectation to find increased pollinator visitation rates in larger populations of both Genista species, the number of flower-visiting insects was unrelated to the number of flowering shoots. Increasing shoot length had a positive and increasing temperature a negative impact on the number of visiting honeybees and bumblebees. Despite the general absence of population size effects on pollinator numbers, the number of fruits and seeds in G. anglica increased with increasing population size. Fruit and seed set in G. pilosa were negatively related to the number of ‘other insects’. Our field observations showed that larger populations of both Genista species flowered earlier than smaller populations and much earlier than reported in the literature. Flowering in large populations therefore tends to coincide less well with pollinator abundance, and this may cause a disruption of the temporal coincidence between flowering phenology and pollinator activity.     

A novel allele of the P-starvation tolerance gene OsPSTOL1 from African rice (Oryza glaberrima Steud) and its distribution in the genus Oryza

Key message

We have developed allele-specific markers for molecular breeding to transfer the PSTOL1 gene from Kasalath to African mega-varieties, including NERICAs, to improve their tolerance to P-deficient soil.

Abstract

The deficiency of phosphorus (P) in soil is a major problem in Sub-Saharan Africa due to general nutrient depletion and the presence of P-fixing soils. Developing rice cultivars with enhanced P efficiency would, therefore, represent a sustainable strategy to improve the livelihood of resource-poor farmers. Recently the Pup1 locus, a major QTL for tolerance to P deficiency in soil, was successfully narrowed-down to a major gene, the protein kinase OsPSTOL1 (P-starvation tolerance), which was found to be generally absent from modern irrigated rice varieties. Our target is to improve the tolerance of African mega-varieties to P deficiency through marker-assisted introgression of PSTOL1. As a first step, we have determined the Pup1 haplotype and surveyed the presence or absence of PSTOL1 and other genes of the Pup1 locus in African mega-varieties, NERICAs (New Rice for Africa) and their Oryza glaberrima parents. Here, we report the presence of a novel PSTOL1 allele in upland NERICAs that was inherited from the O. glaberrima parent CG14. This allele showed a 35 base-pair substitution when aligned to the Kasalath allele, but maintained a fully conserved kinase domain, and is present in most O. glaberrima accessions evaluated. In-silico and marker analysis indicated that many other genes of the Kasalath Pup1 locus were missing in the O. glaberrima genome, including the dirigent-like gene OsPupK20-2, which was shown to be downstream of PSTOL1. We have developed several allele-specific markers for the use for molecular breeding to transfer the PSTOL1 gene from Kasalath to African mega-varieties, including NERICAs.     

Karyotype,heterochromatin content and meiotic features ofPoecilocera picta (Orthoptera,Acrididae)

Cytological study of three distinctly separated populations ofPoecilocera picta revealed a chromosome number of 2N = 18 + XO/ XX. Except for the hemizygosity of a procentric heterochromatic block in the M₆ pair of the Bangalore population, the basic karyotype of the three populations is markedly similar. The autosomal karyotype formula is 2Lt + 4Mt + 1 Mst + 2S st and the telocentric X chromosome is the longest of the complement. All bivalents at pachytene carried procentric heterochromatic blocks. The M₄ is the nucleolus organiser with the NOR region situated interstitially but proximal to the centromere. About 11 μm (4%) of the total (290 μm) autosomal pachytene complement is heterochromatic; a major portion of it is contributed by the S₉ pair which is mostly heterochromatic. Chiasmata are localized proximally and distally and in the S₉ pair their formation is confined to the short procentric euchromatic segment of the long arm. Female meiosis did not reveal any chromomere pattern at pachytene and, unlike in the male, the sex bivalent in the female is indistinguishable from the autosomal bivalents. G- and C-banding patterns in males showed procentric bands in all the chromosomes. In addition there are eight telomeric and two interstitial bands which are C negative. The S₉ pair showed only two bands. The G-banding pattern of the sex chromosome in meiosis showed only a centric band while the heterochromatic body of the facultatively heterochromatic X remained G negative.     

Sesquiterpene and polyacetylene profile of the Bidens pilosa complex (Asteraceae: Heliantheae) from Southeast of Brazil

The pantropical weed Bidens pilosa (Asteraceae) is a species with several taxonomic problems. Recently it has been shown to be a complex of different species. To shed light on this problem in Brazil, dichloromethane extract of leaves of several populations corresponding to three proposed species for the complex in southeastern Brazil (B. pilosa, Bidens alba and Bidens subalternans) were analyzed by GC–MS. Twenty-four substances were detected, of which four resemble polyacetylenes, the others sesquiterpenes. Five sesquiterpenes tentatively identified as E-caryophyllene, α-humulene, germacrene-D, bicyclogermacrene and α-muurolene were found in all three Bidens species. The polyacetylene phenylhepta-1,3,5-triyne was identified only in B. alba. Multivariate analysis (cluster and principal component analyses) separated the three entities, suggesting that these compounds could represent a useful tool to distinguish species in the B. pilosa complex.     

Characterization of type II protein secretion (xcp) genes in the plant growth-stimulatingPseudomonas putida,strain WCS358

InPseudomonas aeruginosa, the products of thexcp genes are required for the secretion of exoproteins across the outer membrane. Despite structural conservation of the Xcp components, secretion of exoproteins via the Xcp pathway is generally not found in heterologous organisms. To study the specificity of this protein secretion pathway, thexcp genes of another fluorescent pseudomonad, the plant growth-promotingPseudomonas putida strain WCS358, were cloned and characterized. Nucleotide sequence analysis revealed the presence of at least five genes, i.e.,xcpP, Q, R, S, andT, with homology toxcp genes ofP. aeruginosa. Unlike the genetic organization inP. aeruginosa, where thexcp cluster consists of two divergently transcribed operons, thexcp genes inP. putida are all oriented in the same direction, and probably comprise a single operon. Upstream ofxcpP inP. putida, an additional open reading frame, with no homolog inP. aeruginosa, was identified, which possibly encodes a lipoprotein. Mutational inactivation ofxcp genes inP. putida did not affect secretion, indicating that no proteins are secreted via the Xcp system under the growth conditions tested, and that an alternative secretion system is operative. To obtain some insight into the secretory pathway involved, the amino acid sequence of the N-terminus of the major extracellular protein was determined. The protein could be identified as flagellin. Mutations in thexcpQ andR genes ofP. aeruginosa could not be complemented by introduction of the correspondingxcp genes ofP. putida. However, expression of a hybrid XcpR protein, composed of the N-terminal one-third ofP. aeruginosa XcpR and the C-terminal two-thirds ofP. putida XcpR, did restore protein secretion in aP. aeruginosa xcpR mutant.     

Taxonomic classification of asexual isolates ofPythium ultimum based on cultural characteristics and mitochondrial DNA restriction patterns

Taxonomic characteristics of 8 isolates ofPythium ultimum and 11 isolates ofPythium in the spherical hyphal swelling (HS) group of Van der Platts-Niterink were compared. Isolates in the two groups had identical temperature growth responses and morphological features of hyphal swellings. All isolates were pathogenic on sugar beet. Attempts to cross HS isolates among themselves and with opposite mating types ofP. heterothallicum andP. sylvaticum failed. HS isolates were not induced to form antheridia when paired with isolates ofP. ultimum using a polycarbonate membrane sandwich technique. In single culture, some HS isolates formed low numbers of spherical structures encompassed by swollen hyphal masses resembling antheridia. Comparisons of restriction banding patterns ofHindIII-digested mitochondrial DNA revealed that all but 1 isolate ofP. ultimum were identical; however, the variable isolate shared 80% of the bands in common with the others. Seven of 11 HS isolates had banding patterns identical to the predominantP. ultimum pattern and 1 isolate shared 96% comigrating bands. On the basis of the number of shared characteristics, these isolates appear to beP. ultimum which have lost the ability to reproduce sexually.     

An extensive analysis of the African rice genetic diversity through a global genotyping

Key message

We present here the first curated collection of wild and cultivated African rice species. For that, we designed specific SNPs and were able to structure these very low diverse species.

Abstract

Oryza glaberrima, the cultivated African rice, is endemic from Africa. This species and its direct ancestor, O. barthii, are valuable tool for improvement of Asian rice O. sativa in terms of abiotic and biotic stress resistance. However, only a few limited studies about the genetic diversity of these species were performed. In the present paper, and for the first time at such extend, we genotyped 279 O. glaberrima, selected both for their impact in current breeding and for their geographical distribution, and 101 O. barthii, chosen based on their geographic origin, using a set of 235 SNPs specifically designed for African rice diversity. Using those data, we were able to structure the individuals from our sample in three populations for O. barthii, related to geography, and two populations in O. glaberrima; these two last populations cannot be linked however to any currently phenotyped trait. Moreover, we were also able to identify misclassification in O. glaberrima as well as in O. barthii and identified new form of O. sativa from the set of African varieties.     

Alterations in amounts and covalent modifications of low-molecular-weight chromosomal proteins in Chinese hamster ovary cells during polyamine depletion

The effect of polyamine depletion on phosphorylation and ADP-ribosylation of low-M_r chromosomal proteins was studied in intact, mutant Chinese hamster ovary cells (CHO-P22) devoid of ornithine decarboxylase activity. When starved of polyamines for 6 days, severe polyamine deficiency develops and the cells gradually stop growing. The rate of DNA synthesis was retarded to 16% of the control value and to 29% in density-inhibited cells. The synthesis of high-mobility-group (HMG) proteins was decreased by 65% in polyamine-depleted cells and by 40% in density-inhibited cells. The synthesis of core histones was decreased by 40% both in polyamine-depleted and density-inhibited cells. In polyamine-depleted cells the molar ratio of the higher-M_r HMG proteins (HMG 1 + 2) to the lower-M_r HMG proteins (HMG 14 + P) was about one-half of that found in cells grown in the presence of putrescine or in density-inhibited cells. In contrast to HMG proteins, no major differences were found in the content of core histones in these cell populations. In the perchloric acid-soluble fraction of nuclear proteins, ³²P was incorporated mainly into histone H1, HMG P and a protein migrating more slowly than HMG 1 (protein P1). Specific changes in the ³²P-labeling and migration of a number of protein bands, including histone H1, was observed in polyamine-depleted cells as compared to cells grown in the presence of putrescine or to density-inhibited cells. ADP-ribosylation experiments using [³H]adenosine showed a different pattern of label distribution; the higher-M_r HMG proteins from polyamine-depleted cells contained about one-half the amount of label found in the proteins from control cells. The lower-M_r HMG proteins and histone H1 were the preferentially labeled proteins in polyamine-depleted cells. Labeling of core histones with [³²P]orthophosphate or [³H]adenosine did not differ markedly in the two cell populations. The results obtained using intact polyamine auxotrophic cells indicated that polyamine depletion is connected with more severe alterations in amounts and covalent modifications (phosphorylation and ADP-ribosylation) of HMG chromosomal proteins and histone H1 than core histones.     

Accessory nucleoli in microsporocytes of hybrid Phlox

Studies were conducted on the number of nucleoli present during diplotene and diakinesis in plants adjacent to, at the edge, and at the center of a zone of secondary intergradation between Phlox pilosa subsp. pilosa and P. pilosa subsp. fulgida. Assessory nucleoli were present in some cells of about 75% of the plants examined. Nucleolar numbers varied from 1 to 5. Where 2 or more nucleoli were present they were usually attached to different chromosomes. As the number of nucleoli increased, the volume of single nucleoli decreased so that volume of all nucleoli was roughly that of a normal single nucleolus. — Only 1% of the microsporocytes from populations adjacent to the zone had accessory nucleoli as compared to 4.6% of the cells in populations at the edge of the zone, and 7.7% in populations in the center. The correlation between population hybridity and incidence of accessory nucleoli is r = 0.80.     

The proteolytic systems and heterologous proteins degradation in the methylotrophic yeastPichia pastoris

ThePichia pastoris expression system has been successfully used for production of various recombinant heterogeneous proteins. The productivity ofP. pastoris can be improved substantially by bioreactor cultivations. However, heterologous proteins degradation increases as well in high-cell density culture. Proteolytic degradation is a serious problem since the yeast has been employed to express recombinant proteins. In this review, some of the recent developments, as well as strategies for reducing proteolytic degradation of the expressed recombinant protein at cultivation, cellular and protein levels on the cytosolic proteasome, vacuolar proteases, and proteases located within the secretory pathway inP. pastoris, are reviewed.     

An Arabidopsis VPS45p Homolog Implicated in Protein Transport to the Vacuole

The Sec1p family of proteins is required for vesicle-mediated protein trafficking between various organelles of the endomembrane system. This family includes Vps45p, which is required for transport to the vacuole in yeast (Saccharomyces cerevisiae). We have isolated a cDNA encoding a VPS45 homolog from Arabidopsis thaliana (AtVPS45). The cDNA is able to complement both the temperature-sensitive growth defect and the vacuolar-targeting defect of a yeast vps45 mutant, indicating that the two proteins are functionally related. AtVPS45p is a peripheral membrane protein that associates with microsomal membranes. Sucrose-density gradient fractionation demonstrated that AtVPS45p co-fractionates with AtELP, a potential vacuolar protein sorting receptor, implying that they may reside on the same membrane populations. These results indicate that AtVPS45p is likely to function in the transport of proteins to the vacuole in plants.     

Rescue of Defective Poliovirus RNA Replication by 3AB-Containing Precursor Polyproteins

This study demonstrates the in vitro complementation of an RNA replication-defective lesion in poliovirus RNA by providing a replicase/polymerase precursor polypeptide [P3(wt) {wild type}] in trans. The replication-defective mutation was a phenylalanine-to-histidine change (F69H) in the hydrophobic domain of the membrane-associated viral protein 3AB. RNAs encoding wild-type forms of protein 3AB or the P3 precursor polypeptide were cotranslated with full-length poliovirus RNAs containing the F69H mutation in a HeLa cell-free translation/replication assay in an attempt to trans complement the RNA replication defect exhibited by the 3AB(F69H) lesion. Unexpectedly, generation of 3AB(wt) in trans was not able to efficiently complement the defective replication complex; however, cotranslation of the large P3(wt) precursor protein allowed rescue of RNA replication. Furthermore, P3 proteins harboring mutations that resulted in either an inactive polymerase or an inactive proteinase domain displayed differential abilities to trans complement the RNA replication defect. Our results indicate that replication proteins like 3AB may need to be delivered to the poliovirus replication complex in the form of a larger 3AB-containing protein precursor prior to complex assembly rather than as the mature viral cleavage product.     

Two-dimensional protein electrophoretic analysis of postponed aging inDrosophila

Five populations ofDrosophila melanogaster that had been selected for postponed aging were compared with five control populations using two-dimensional protein gel electrophoresis. The goals of the study were to identify specific proteins associated with postponed aging and to survey the population genetics of the response to selection. A total of 321 proteins were resolvable per population; these proteins were scored according to their intensity. The resulting data were analyzed using resampling, combinatoric, and maximum parsimony methods. The analysis indicated that the populations with postponed aging were different from their controls with respect to specific proteins and with respect to the variation between populations. The populations selected for postponed aging were more heterogeneous between populations than were the control populations. Maximum parsimony trees separate the selected populations, as a group, from their controls, thereby exhibiting a homoplastic pattern.     

Inter- and intrapopulational variations in the peroxidase isozyme ofAsarum nipponicum F. Maekawa

Both the inter- and intrapopulational variations in five natural populations ofAsarum nipponicum F. Maekawa were demonstrated for peroxidase isozymes using gel electrofocusing. The patterns of isozymic variation suggested that a very short distance (100 m) effectively isolates populations. The reason why some bands were not detected in a particular population is discussed in terms of genetic drift.