Identification of a movement protein of the tenuivirus rice stripe virus

Rice stripe virus (RSV) is the type member of the genus Tenuivirus. RSV has four single-stranded RNAs and causes severe disease in rice fields in different parts of China. To date, no reports have described how RSV spreads within host plants or the viral and/or host factor(s) required for tenuivirus movement. We investigated functions of six RSV-encoded proteins using trans-complementation experiments and biolistic bombardment. We demonstrate that NSvc4, encoded by RSV RNA4, supports the intercellular trafficking of a movement-deficient Potato virus X in Nicotiana benthamiana leaves. We also determined that upon biolistic bombardment or agroinfiltration, NSvc4:enhanced green fluorescent protein (eGFP) fusion proteins localize predominantly near or within the walls of onion and tobacco epidermal cells. In addition, the NSvc4:eGFP fusion protein can move from initially bombarded cells to neighboring cells in Nicotiana benthamiana leaves. Immunocytochemistry using tissue sections from RSV-infected rice leaves and an RSV NSvc4-specific antibody showed that the NSvc4 protein accumulated in walls of RSV-infected leaf cells. Gel retardation assays revealed that the NSvc4 protein interacts with single-stranded RNA in vitro, a common feature of many reported plant viral movement proteins (MPs). RSV NSvc4 failed to interact with the RSV nucleocapsid protein using yeast two-hybrid assays. Taken together, our data indicate that RSV NSvc4 is likely an MP of the virus. This is the first report describing a tenuivirus MP.     

Characterisation of datura yellow vein virus, a newly described rhabdovirus from Australia

A previously undescribed sub-group 2 rhabdovirus was isolated in Queensland from Datura stramonium with symptoms of vein yellowing, leaf distortion and reduced leaf size. Particles accumulated in the perinuclear space of infected cells of D. stramonium and measured 77 × 166 nm in preparations from sap. The virus was named datura yellow vein virus (DYVV) and was graft-transmitted to several hosts in the Solanaceae including Lycopersicon esculentum, Nicotiana tabacum and Solanum melongena, but not to Capsicum annuum or Solanum tuberosum. DYVV was not transmitted by mechanical inoculation and no insect vector was found. Purified particles of DYVV contained four structural proteins with molecular weights of about 78, 47, 41 and 36 kd. The 78 kd protein bound the lectin concanavalin A, thus identifying it as the viral glycoprotein. DYVV was serologically distinct from 11 other rhabdoviruses belonging to both subgroups, including potato chlorotic stunt, potato yellow dwarf (2 isolates) and tomato vein yellowing viruses. The glycoprotein only of DYVV cross-reacted with a polyclonal antiserum to sonchus yellow net virus.     

Identification of vectors of rice yellow mottle virus in Tanzania

Rice yellow mottle virus (RYMV) is specific to Africa and has been reported in some countries in East Africa and almost all the countries in West Africa. At present, it is undoubtedly the most important disease of rice in Tanzania. It was first reported in the 1980's. It has spread fast and is now found in almost all the rice growing areas. In view of the increasing incidence and importance of RYMV on rice production in Tanzania, studies on the epidemiology of the disease were initiated in order to find ways of controlling the disease. Transmission studies were carried out on seventy‐seven species of beetles and grasshoppers collected from different rice growing locations to determine vector identity. Four vectors have been identified (three chrysomelids; Dactylispa sp., Chaetocnema sp. and Chaetocnema pulla) and one tetrigid grasshopper. The wide distribution of Chaetocnema spp. in the RYMV endemic areas suggests that the species are the most important vectors responsible for infections in these areas.     

Interaction between the movement protein of barley yellow dwarf virus and the cell nuclear envelope: role of a putative amphiphilic alpha-helix at the N-terminus of the movement protein

The open reading frame 4 (ORF 4) gene product of barley yellow dwarf virus (BYDV) may act as a movement protein (MP) by assisting the transport of viral genomic RNA across the nuclear envelope (NE) of host plant cells. To investigate interactions between BYDV MP and the NE, wild-type and mutant open reading frame (ORF 4)-green fluorescent protein (GFP) fusion cistrons were expressed in insect cells. A fusion protein expressed by the wild-type ORF 4-GFP cistron associated with the NE and caused protrusions from its surface. The fusion protein expressed by the mutant ORF 4-GFP cistron lacked a putative amphiphilic alpha-helix at its N-terminus and although associating with the NE, showed decreased levels of protrusions. A peptide homologue of this putative alpha-helix induced an increase of 7 degrees C in the phase transition temperature of dimyrystoyl phosphatidylserine (DMPS) membranes, accompanied by a decrease in membrane fluidity, but exhibited no significant interaction with either dimyristoyl phosphatidylcholine (DMPC) or dimyristoyl phosphatidylethanolamine (DMPE) membranes. These results strongly support the view that BYDV MP may interact with the NE to help transport viral genomic RNA into the nuclear compartment. This function of BYDV MP appears to involve protrusions on the surface of the NE and may require the presence of an N-terminal amphiphilic alpha-helix, which is speculated to destabilize membranes, thereby assisting the entry of BYDV-GAV into the nuclear compartment.     

Identification of a novel secretory leukocyte protease inhibitor-binding protein involved in membrane phospholipid movement

Previous studies have suggested that human salivary secretory leukocyte protease inhibitor (SLPI) inhibits HIV-1 by binding to a host cell surface protein of unknown identity. Using the yeast two-hybrid assay, we identified a gene sequence encoding a novel SLPI-binding protein (SLPI-BP). The 1.5-kb cDNA encodes a 318-amino acid protein with a predicted transmembrane segment near the C-terminus. Sequence analysis revealed that SLPI-BP is the human scramblase protein that is involved in the movement of membrane phospholipids. Co-expression of SLPI and SLPI-BP followed by an S-protein pulldown assay confirmed the specific interaction between these two proteins. Our data represent the first report for the identity of SLPI-BP.     

A structural comparison of concanavalin a and tomato bushy stunt virus protein

Summary Significant structural equivalence has been found among the polypeptide folds of the two tomato bushy stunt virus (TBSV) subunit domains and concanavalin A. This suggests gene duplication in the TBSV coat protein and leads to speculation on common functional properties of concanavalin A and viral coat proteins.Non-standard abbreviations TBSV tomato bushy stunt virus - SBMV southern bean mosaic virus     

Identification of the major protein components of rice egg cells

The female gamete, the egg cell, is a specially differentiated haploid cell that develops into an embryo following fertilization. In the present study, we analyzed egg cell lysates by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and subsequent mass spectrometry-based proteomics technology and identified the major proteins expressed in rice egg cells. The proteins identified included glyceraldehyde-3-phosphate dehydrogenase, ascorbate peroxidase and heat shock protein 90. The abundant existence of chaperons and antioxidant enzymes in plant egg cells indicates that the major protein components of plant egg cells are partly analogous to those of mammalian eggs and oocytes.     

Characterization of infectious hematopoietic necrosis virus mRNA species reveals a nonvirion rhabdovirus protein.

The genome RNA and six mRNA species of infectious hematopoietic necrosis virus were analyzed by denaturing gel electrophoresis. The following molecular weights were determined: genome RNA, 3.7 X 10(6); mRNA 1, 2.26 X 10(6); mRNA 2, 5.63 X 10(5); mRNA 3, 4.84 X 10(5); mRNA 4 (containing two different mRNA species), 3.00 X 10(5); and mRNA 5, 1.95 X 10(5). Densitometer analyses of gels were used to calculate the molar ratios of the intracellular mRNA species: mRNA 1, 0.02; mRNA 2, 0.49; mRNA 3, 1.0; mRNA 4, 2.52; and mRNA 5, 0.41. Hybrid selection studies determined the mRNA coding assignments as follows: mRNA 1 encodes the viral polymerase, L; mRNA 2 encodes the glycoprotein, G; mRNA 3 encodes the nucleocapsid protein, N; mRNA 4 is composed of two comigrating mRNA species which encode the matrix proteins, M1 and M2; and mRNA 5 encodes a previously unrecognized viral protein which is induced in infected cells but is not present in mature virions. This nonvirion protein has been designated the NV protein.     

Tomato bushy stunt virus genomic RNA accumulation is regulated by interdependent cis-acting elements within the movement protein open reading frames

This study on Tomato bushy stunt virus identified and defined three previously unknown regulatory sequences involved in RNA accumulation that are located within the 3'-proximal nested movement protein genes p22 and p19. The first is a 16-nucleotide (nt) element termed III-A that is positioned at the very 3' end of p22 and is essential for RNA accumulation. Approximately 300 nt upstream of III-A resides an approximately 80-nt inhibitory element (IE) that is obstructive to replication only in the absence of a third regulatory element of approximately 30 nt (SUR-III) that is positioned immediately upstream of III-A. Inspection of the nucleotide sequences predicted that III-A and SUR-III can form looped hairpins. A comparison of different tombusviruses showed, in each case, conservation for potential base pairing between the two predicted hairpin-loops. Insertion of a spacer adjacent to the predicted hairpins had no or a minimal effect on RNA accumulation, whereas an insertion in the putative III-A loop abolished genomic RNA multiplication. We conclude that the sequences composing the predicted III-A and SUR-III hairpin-loops are crucial for optimal RNA accumulation and that the inhibitory effect of IE surfaces when the alleged interaction between SUR-III and III-A is disturbed.     

Functional characterization of the stunt lemma palea 1 mutant allele in rice

The rice stunted lemma/palea 1 (slp1) mutant displays a dwarf, shortened panicle length, degenerated lemma and palea, and sterility. A previous study suggested that a missense mutation at the sixth amino acid of the OsSPL16 protein was likely to be responsible for the slp1 mutant phenotype. The current study shows that the overexpression of the wild-type OsSPL16 allele in slp1/slp1 and Slp1/slp1 mutants was unable to convert the slp1 mutant phenotype to normal. However, the introduction of the mutant OsSPL16 allele into a normal rice cultivar led to the slp1 mutant phenotype in transgenic plants. These results indicated that the missense mutation in OsSPL16 creates a neomorphic allele, which affects plant height and the development of the inflorescence and spikelet.     

Identification of rice genes induced in a rice blast-resistant mutant

To clarify mechanisms of rice blast resistance in rice plants we used suppression subtractive hybridization (SSH) to isolate genes induced upon rice blast inoculation in a rice blast-resistant mutant. A total of 26 rice cDNAs were isolated and found to have elevated expression upon rice blast infection in a rice blast-resistant derivative, SHM-11, of the rice cultivar, Sanghaehyanghyella. Sequencing of the cDNAs revealed that many of the proteins they encoded had been previously described as involved in plant responses against pathogen attack. Two interesting groups of the defense-related proteins consisted of three different PR5 homologues and four different protease inhibitors, all highly expressed in the rice blast mutant. Genes encoding proteins involved in signal transduction and regulation were also identified, including translation initiation factor eIF5A, C2 domain DNA binding protein, putative rice EDS and putative receptor like kinase. Most of the identified cDNAs were highly expressed 24 h after blast inoculation. Our results suggest that a pathway regulating defense gene expression may be altered in the mutant, resulting in early induction of the defense genes upon fungal infection.     

Thermal effects on movement patterns of yellow baboons

This paper examines the effect of thermal environment on movement patterns of free-ranging yellow baboons (Papio cynocephalus). For Amboseli baboons, one source of potential thermal stress is intense midday heat, and a plausible thermoregulatory response is for animals to simply move into the shade. I therefore examined the hypothesis that baboons would choose quadrats with higher shade availability (as measured by vegetation cover) in response to increasing midday heat loads (as measured by air temperature and solar radiation). Surprisingly, this was not the case—neither ambient air temperature, ambient solar radiation, nor quadrat plant species composition had a significant effect on shade availability of quadrat selected. Instead, thermal conditions affected a different aspect of baboon movements; namely, spatial displacement rates. At high air temperatures, baboons as a group traversed woodland habitats more slowly, and bare pans more quickly, than at lower air temperatures. I surmised that this relationship might reflect thermal effects on movement patterns at a smaller scale: if individuals exposed to high heat loads spent more time resting in shade under clumps of vegetation, they would thereby traverse densely-vegetated (hence shaded) quadrats more slowly. To address this question directly, I obtained focal sample data on activity and microhabitat budgets of individual baboons in relation to environmental temperature. The frequency of most combinations of activity state (e.g., grooming, social behavior) and microenvironment state (e.g., elevation, proximity to vegetation) did not vary monotonically with air temperature. However, baboons in shaded locations (but not those in unshaded locations) spent more time resting and less time moving at high air temperatures than low. In other words, baboon activity budgets depended on both microclimate and microhabitat—animals reduced their activity, particularly movement, when they encountered shade under hot conditions. This pattern of microhabitat choice in turn led to temperature-dependent changes in travel rate at the habitat level. These observational studies of movement patterns suggest that Amboseli baboons employ opportunistic thermoregulation—they do not seek out densely-shaded habitats or individual patches of shade at high air temperatures. Instead, they respond to environmental heat loads by resting, and thereby slowing down, when they happen to encounter plant shade. Aspects of baboon ecology that favor such an opportunistic mode of thermoregulation include large body size and non-thermal constraints on movement patterns.     

Reversible inhibition of spreading of in vitro infection and imbalance of viral protein accumulation at low pH in viral hemorrhagic septicemia rhabdovirus, a salmonid rhabdovirus

The inhibition of viral hemorrhagic septicemia rhabdovirus (VHSV) in vitro infection by pHs of <7 (low pH) has been previously reported. Nevertheless, the details of the mechanism underlying this effect remain obscure. We present evidence showing that low-pH inhibition occurs during a viral postadsorption step. Thus, while VHSV bound, replicated within single cells, and presented its G protein on the membranes of infected cells at both low and physiological pHs, both cell-to-cell spreading of infection (as estimated by the appearance of foci of infected cells) and fusion (as estimated by a syncytium assay) were inhibited by this low pH. The decreased VHSV titers and the inhibition of both cell-to-cell spreading of infection and fusion could be reversed by adjusting the pH to 7.5 at any time during infection. This effect should be taken into account to avoid false negatives in the diagnosis of VHSV by cell culture. On the other hand, the cell-to-cell spreading of infection at pH 7.5 could be stopped at any time by reducing the pH to 6.5. Since at low pH there were changes in the protein G conformation and smaller and imbalanced amounts of N with respect to M1, M2, and G viral proteins, alterations of the assembly and/or budding of VHSV are most probably involved in the absence of newly released infective virions.     

三化螟幼虫的田间分布型

通过采用聚集度指标的方法,对三化螟幼虫在稻田的空间格局进行了系统的研究。测定了幼虫个体群大小、分析聚集原因,为田间抽样、《查定》、及防治提供了依据。