Viruses face a double defense by plant small RNAs

Plants fight viral infections with enzymes that digest viral RNA, but viruses retaliate with proteins that suppress these enzymes. To boost their antiviral response plants deploy enzymes with redundant functions.

Small RNA viruses of insects : expression in plants and RNA silencing

Interest in insect small RNA viruses (SRVs) has grown slowly but steadily. A number of new viruses have been analyzed at the sequence level, adding to our knowledge of their diversity at the level of both individual virus species and families. In particular, a number of possible new virus families have emerged. This research has largely been driven by interest in their potential for pest control, as well as in their importance as the causal agents of disease in beneficial arthropods. At the same time, research into known viruses has made valuable contributions to our understanding of an emerging new field of central importance to molecular biology-the existence of RNA-based gene silencing, developmental control, and adaptive immune systems in eukaryotes. Subject to RNA-based adaptive immune responses in their hosts, viruses have evolved a variety of genes encoding proteins capable of suppressing the immune response. Such genes were first identified in plant viruses, but the first examples known from animal viruses were identified in insect RNA viruses. This chapter will address the diversity of insect SRVs, and attempts to harness their simplicity in the engineering of transgenic plants expressing viruses for resistance to insect pests. We also describe RNA interference and antiviral pathways identified in plants and animals, how they have led viruses to evolve genes capable of suppressing such adaptive immunity, and the problems presented by these pathways for the strategy of expressing viruses in transgenic plants. Approaches for countering these problems are also discussed. © 2006 Elsevier Inc. All rights reserved.

Production of transgenic rice with rice ragged stunt virus synthetic resistance genes

Rice ragged stunt virus (RRSV) is an important pathogen of rice affecting its cultivation in South and South East Asia. An approach based on pathogen derived resistance (PDR) was used to produce RRSV resistant rice cultivars. Sequences from the coding region of RRSV genome segments 7 and 10 (non-structural genes), and 5, 8 and 9 (structural genes) were placed in sense or antisense orientation behind the plant expression promoters CaMV35S, RolC, Ubil, Actl and RBTV. Rice cultivars Taipei 309 and Chinsurah Boro II were transformed by biolistic and/or Agrobacterium-mediated delivery of one or more of these PDR gene constructs. A large number of transgenic lines were produced from calli derived from mature or immature embryos, co-bombarded with the marker gene hph encoding hygromycin resistance and RRSV PDR genes or co-cultivated with strains having the binary vector containing these two genes. Both Mendelian and non-Mendelian segregations were observed in transgenic progeny, especially with transgenic lines produced by biolistics. Preliminary tests conducted in China on selected transgenic lines indicate that plants with RRSV segment 5 antisense PDR gene confer RRSV resistance.

A 22-nt artificial microRNA mediates widespread RNA silencing in Arabidopsis

It is known that 22-nucleotide (nt) microRNAs (miRNAs) derived from asymmetric duplexes trigger phased small-interfering RNA (phasiRNA) production from complementary targets. Here we investigate the efficacy of 22-nt artificial miRNA (amiRNA)-mediated RNA silencing relative to conventional hairpin RNA (hpRNA) and 21-nt amiRNA-mediated RNA silencing. CHALCONE SYNTHASE (CHS) was selected as a target in Arabidopsis thaliana due to the obvious and non-lethal loss of anthocyanin accumulation upon widespread RNA silencing. Over-expression of CHS in the pap1-D background facilitated visual detection of both local and systemic RNA silencing. RNA silencing was initiated in leaf tissues from hpRNA and amiRNA plant expression vectors under the control of an Arabidopsis RuBisCo small subunit 1A promoter (SSU). In this system, hpRNA expression triggered CHS silencing in most leaf tissues but not in roots or seed coats. Similarly, 21-nt amiRNA expression from symmetric miRNA/miRNA* duplexes triggered CHS silencing in all leaf tissues but not in roots or seed coats. However, 22-nt amiRNA expression from an asymmetric duplex triggered CHS silencing in all tissues, including roots and seed coats, in the majority of plant lines. This widespread CHS silencing required RNA-DEPENDENT RNA POLYMERASE6-mediated accumulation of phasiRNAs from the endogenous CHS transcript. These results demonstrate the efficacy of asymmetric 22-nt amiRNA-directed RNA silencing and associated phasiRNA production and activity, in mediating widespread RNA silencing of an endogenous target gene. Asymmetric 22-nt amiRNA-directed RNA silencing requires little modification of existing amiRNA technology and is expected to be effective in suppressing other genes and/or members of gene families.

Complete genomic sequence of a Rubus yellow net virus isolate and detection of genome-wide pararetrovirus-derived small RNAs

Rubus yellow net virus (RYNV) was cloned and sequenced from a red raspberry (Rubus idaeus L.) plant exhibiting symptoms of mosaic and mottling in the leaves. Its genomic sequence indicates that it is a distinct member of the genus Badnavirus, with 7932. bp and seven ORFs, the first three corresponding in size and location to the ORFs found in the type member Commelina yellow mottle virus. Bioinformatic analysis of the genomic sequence detected several features including nucleic acid binding motifs, multiple zinc finger-like sequences and domains associated with cellular signaling. Subsequent sequencing of the small RNAs (sRNAs) from RYNV-infected R. idaeus leaf tissue was used to determine any RYNV sequences targeted by RNA silencing and identified abundant virus-derived small RNAs (vsRNAs). The majority of the vsRNAs were 22-nt in length. We observed a highly uneven genome-wide distribution of vsRNAs with strong clustering to small defined regions distributed over both strands of the RYNV genome. Together, our data show that sequences of the aphid-transmitted pararetrovirus RYNV are targeted in red raspberry by the interfering RNA pathway, a predominant antiviral defense mechanism in plants. © 2013.

Small RNA sequencing of potato leafroll virus-infected plants reveals an additional subgenomic RNA encoding a sequence-specific RNA-binding protein

Potato leafroll virus (PLRV) is a positive-strand RNA virus that generates subgenomic RNAs (sgRNA) for expression of 3' proximal genes. Small RNA (sRNA) sequencing and mapping of the PLRV-derived sRNAs revealed coverage of the entire viral genome with the exception of four distinctive gaps. Remarkably, these gaps mapped to areas of PLRV genome with extensive secondary structures, such as the internal ribosome entry site and 5' transcriptional start site of sgRNA1 and sgRNA2. The last gap mapped to ~500. nt from the 3' terminus of PLRV genome and suggested the possible presence of an additional sgRNA for PLRV. Quantitative real-time PCR and northern blot analysis confirmed the expression of sgRNA3 and subsequent analyses placed its 5' transcriptional start site at position 5347 of PLRV genome. A regulatory role is proposed for the PLRV sgRNA3 as it encodes for an RNA-binding protein with specificity to the 5' of PLRV genomic RNA. © 2013.

The roles of plant dsRNA-binding proteins in RNAi-like pathways

Dicers are associated with double-stranded RNA-binding proteins (dsRBPs) in animals. In the plant, Arabidopsis, there are four dicer-like (DCL) proteins and five potential dsRBPs. These DCLs act redundantly and hierarchically. However, we show there is little or no redundancy or hierarchy amongst the DRBs in their DCL interactions. DCL1 operates exclusively with DRB1 to produce micro (mi)RNAs, DCL4 operates exclusively with DRB4 to produce trans-acting (ta) siRNAs and 21nt siRNAs from viral RNA. DCL2 and DCL3 produce viral siRNAs without requiring assistance from any dsRBP. DRB2, DRB3 and DRB5 appear unnecessary for mi-, tasi-, viral si-, or heterochromatinising siRNA production but act redundantly in a developmental pathway. © 2008 Federation of European Biochemical Societies.

Posttranscriptional gene silencing is not compromised in the Arabidopsis CARPEL FACTORY (DICER-LIKE1) mutant, a homolog of dicer-1 from Drosophila

Posttranscriptional silencing (PTGS) in plants, nematodes, Drosophila, and perhaps all eukaryotes operates by sequence-specific degradation or translational inhibition of the target mRNA. These processes are mediated by duplexed RNA. In Drosophila and nematodes, double-stranded (ds)RNA or self-complementary RNA is processed into fragments of approximately 21 nt by Dicer-1 [1, 2]. These small interfering RNAs (siRNAs) serve as guides to target degradation of homologous single-stranded (ss)RNA [1, 3]. In some cases, the approximately 21 nt guide fragments derived from endogenous, imperfectly self-complementary RNAs cause translational inhibition of their target mRNAs, with which they have substantial, but not perfect sequence complementarity [4-6]. These small temporal RNAs (stRNAs) belong to a class of noncoding microRNAs (miRNAs), 20-24 nt in length, that are found in flies, plants, nematodes, and mammals [4, 6-12]. In nematodes, the Dicer-1 enzyme catalyzes the production of both siRNA and stRNA [2, 13-15]. Mutation of the Arabidopsis Dicer-1 homolog, CARPEL FACTORY (CAF), blocks miRNA production [1, 4, 16-18]. Here, we report that the same caf mutant does not block either PTGS or siRNA production induced by self-complementary hairpin RNA. This suggests either that this mutation only impairs miRNA formation or, more interestingly, that plants have two distinct dicer-like enzymes, one for miRNA and another for siRNAi production.

Co-creating knowledge online : approaches for community artists

Forming peer alliances to share and build knowledge is an important aspect of community arts practice, and these co-creation processes are increasingly being mediated by the internet. This paper offers guidance for practitioners who are interested in better utilising the internet to connect, share, and make new knowledge. It argues that new approaches are required to foster the organising activities that underpin online co-creation, building from the premise that people have become increasingly networked as individuals rather than in groups (Rainie and Wellman 2012: 6), and that these new ways of connecting enable new modes of peer-to-peer production and exchange. This position advocates that practitioners move beyond situating the internet as a platform for dissemination and a tool for co-creating media, to embrace its knowledge collaboration potential. Drawing on a design experiment I developed to promote online knowledge co-creation, this paper suggests three development phases – developing connections, developing ideas, and developing agility – to ground six methods. They are: switching and routing, engaging in small trades of ideas with networked individuals; organising, co-ordinating networked individuals and their data; beta-release, offering ‘beta’ artifacts as knowledge trades; beta-testing, trialing and modifying other peoples ‘beta’ ideas; adapting, responding to technological disruption; and, reconfiguring, embracing opportunities offered by technological disruption. These approaches position knowledge co-creation as another capability of the community artist, along with co-creating art and media.

Diabetes, hyperinsulinemia, and hyperlipidemia in small aboriginal community in northern Australia

A small rural Aboriginal community in northern Australia was surveyed for diabetes, impaired glucose tolerance (IGT), hyperinsulinemia, and lipid levels. Of the 122 adults >17 yr of age who participated (95% response rate), 11.5% had diabetes, 7.4% had IGT, and the remaining 81.1% had normal glucose tolerance. Both diabetes and IGT were strongly age related. This high frequency of diabetes occurred, despite the population being relatively lean. Although the body mass index (BMI) increased with age in both men and women, only 25% of the population overall had BMI >25 kg/m2. There were wide ranges of insulin responses to glucose, with the upper fertile of 2-h insulin levels being more than seven times higher than the lower fertile (144 ± 13 vs. 19 ± 1 mLI/L). Hyperinsulinemia was associated with IGT, elevated triglycerides, and lower high-density lipoprotein cholesterol levels. Lipid abnormalities were much more frequent among men than women. Cholesterol levels were an average of 0.55 mM higher and triglycerides an average of 1.05 mM higher in men than in women, and both increased with age. In conclusion, this small isolated Aboriginal population from northern Australia had an unexpectedly high frequency of diabetes (in view of their relative leanness) in association with a high frequency of metabolic abnormalities indicative of insulin resistance (hyperinsulinemia, IGT, hypertriglyceridemia).

Application of lean production to reducing operational waste in a tile manufacturing process

Roofing tile manufacturing is a mass production process with high operational and inventory wastes and costs. Due to huge operational costs, excessive inventory and wastes, and quality problems, roofing tile manufacturers are trying to implement lean manufacturing practice in their operations in order to remain competitive in an ncreasingly competitive global market. The aim of this research is to evaluate the possibility of reducing the operational and inventory costs of the tile manufacturing process through waste minimization. This paper analyses the current waste situation in a tile manufacturing process and develops current and future value stream mapping for such a process with a view to implementing lean principles in manufacturing. The focus of the approach is on cost reduction by eliminating non-value-added activities.

Pornography as creative industry

Much academic writing on pornography focuses on the genre’s exceptionalism (the ways in which it is different from other forms of culture). This paper rather focuses on its typicality as a Creative Industry. As with other Creative Industries, in the production and distribution of pornography it is producers who make most money. While a small number of on-camera performers become wealthy and powerful the majority lack creative control and are poorly paid. The careers of performers are typically short examples of “nomadic labour”. Many creative workers are involved in the production of pornography apart those in front of the camera, and the skills of these behind-the-camera creatives can be transferable across sectors. Like other Creative Industries, the pornography business is facing challenges from increased digitalization and globalization; and like other Creative Industries the solutions to these challenges lie in branding, niche marketing, and exploiting new technological possibilities.

Small signal stability analysis of power systems with high penetration of wind power

The integration of large amount of wind power into a power system imposes a new challenge for the secure and economic operation of the system. It is necessary to investigate the impacts of wind power generation on the dynamic behavior of the power system concerned. This paper investigates the impacts of large amount of wind power on small signal stability and the corresponding control strategies to mitigate the negative effects. The concepts of different types of wind turbine generators (WTGs) and the principles of the grid-connected structures of wind power generation systems are first briefly introduced. Then, the state-of-the-art of the studies on the impacts of WTGs on small signal stability as well as potential problems to be studied are clarified. Finally, the control strategies on WTGs to enhance power system damping characteristics are presented.

Queensland small towns Documentary project

This photographic documentary project explores diverse stories of Queensland mining towns. Images compare the interplay between communities, commerce and natural resources. The project is a collaboration between three Brisbane universities (Queensland University of Technology, Griffith University and University of Queensland) and is supported by the Brisbane Powerhouse and the State Library of Queensland. Funded by a $50,000 Arts Queensland grant.

Intranasal immunization with C. muridarum major outer membrane protein (MOMP) and cholera toxin elicits local production of neutralising IgA in the prostate

Successful control of sexually transmitted diseases (STDs) through vaccination will require the development of vaccine strategies that target protective immunity to both the female and male reproductive tracts (MRT). In the male, the immune privileged nature of the male reproductive tract provides a barrier to entry of serum immunoglobulins into the male reproductive ducts, thereby preventing the induction of protective immunity using conventional injectable vaccination techniques. In this study we investigated the potential of intranasal (IN) immunization to elicit anti-chlamydial immunity in BALB/c male mice. Intranasal immunization with Chlamydia muridarum major outer membrane protein (MOMP) admixed with cholera toxin (CT) resulted in high levels of MOMP-specific IgA in prostatic fluids (PF) and MOMP-specific IgA-secreting cells in the prostate. Prostatic fluid IgA inhibited in vitro infection of McCoy cells with C. muridarum. Using RT-PCR we also show that mRNA for the polymeric immunoglobulin receptor (PIgR), which transports IgA across mucosal epithelia, is expressed only in the prostate but not in other regions of the male reproductive ducts upstream of the prostate. These data suggest that using intranasal immunization to target IgA to the prostate may protect males against STDs while at the same time maintaining the state of immune privilege within the MRT.