Transcriptional profiling of pea ABR17 mediated changes in gene expression in Arabidopsis thaliana

<p>Abstract</p> <p>Background</p> <p>Pathogenesis-related proteins belonging to group 10 (PR10) are elevated in response to biotic and abiotic stresses in plants. Previously, we have shown a drastic salinity-induced increase in the levels of ABR17, a member of the PR10 family, in pea. Furthermore, we have also demonstrated that the constitutive expression of pea <it>ABR17 </it>cDNA in <it>Arabidopsis thaliana </it>and <it>Brassica napus </it>enhances their germination and early seedling growth under stress. Although it has been reported that several members of the PR10 family including ABR17 possess RNase activity, the exact mechanism by which the aforementioned characteristics are conferred by ABR17 is unknown at this time. We hypothesized that a study of differences in transcriptome between wild type (WT) and <it>ABR17 </it>transgenic <it>A. thaliana </it>may shed light on this process.</p> <p>Results</p> <p>The molecular changes brought about by the expression of pea <it>ABR17 </it>cDNA in <it>A. thaliana </it>in the presence or absence of salt stress were investigated using microarrays consisting of 70-mer oligonucleotide probes representing 23,686 <it>Arabidopsis </it>genes. Statistical analysis identified number of genes which were over represented among up- or down-regulated transcripts in the transgenic line. Our results highlight the important roles of many abscisic acid (ABA) and cytokinin (CK) responsive genes in <it>ABR17 </it>transgenic lines. Although the transcriptional changes followed a general salt response theme in both WT and transgenic seedlings under salt stress, many genes exhibited differential expression patterns when the transgenic and WT lines were compared. These genes include plant defensins, heat shock proteins, other defense related genes, and several transcriptional factors. Our microarray results for selected genes were validated using quantitative real-time PCR.</p> <p>Conclusion</p> <p>Transcriptional analysis in <it>ABR17 </it>transgenic <it>Arabidopsis </it>plants, both under normal and saline conditions, revealed significant changes in abundance of transcripts for many stress responsive genes, as well as those related to plant growth and development. Our results also suggest that <it>ABR17 </it>may mediate stress tolerance through the modulation of many ABA- and CK-responsive genes and may further our understanding of the role of ABR17 in mediating plant stress responses.</p

The POM Monoclonals: A Comprehensive Set of Antibodies to Non-Overlapping Prion Protein Epitopes

PrPSc, a misfolded and aggregated form of the cellular prion protein PrPC, is the only defined constituent of the transmissible agent causing prion diseases. Expression of PrPC in the host organism is necessary for prion replication and for prion neurotoxicity. Understanding prion diseases necessitates detailed structural insights into PrPC and PrPSc. Towards this goal, we have developed a comprehensive collection of monoclonal antibodies denoted POM1 to POM19 and directed against many different epitopes of mouse PrPC. Three epitopes are located within the N-terminal octarepeat region, one is situated within the central unstructured region, and four epitopes are discontinuous within the globular C-proximal domain of PrPC. Some of these antibodies recognize epitopes that are resilient to protease digestion in PrPSc. Other antibodies immunoprecipitate PrPC, but not PrPSc. A third group was found to immunoprecipitate both PrP isoforms. Some of the latter antibodies could be blocked with epitope-mimicking peptides, and incubation with an excess of these peptides allowed for immunochromatography of PrPC and PrPSc. Amino-proximal antibodies were found to react with repetitive PrPC epitopes, thereby vastly increasing their avidity. We have also created functional single-chain miniantibodies from selected POMs, which retained the binding characteristics despite their low molecular mass. The POM collection, thus, represents a unique set of reagents allowing for studies with a variety of techniques, including western blotting, ELISA, immunoprecipitation, conformation-dependent immunoassays, and plasmon surface plasmon resonance-based assays

Alleviating Poverty and Malnutrition in Agro-Biodiversity Hotspots : final technical report (1 March 2011 - 31 August 2014)

The APM project provided a basket of livelihood options, working with local participants to test integrated agricultural systems, production technologies, techniques for crop and animal husbandry, on and off-farm enterprises, improving nutrition literacy, as well as social and institutional innovations. The adoption of integrated practices in cultivation of finger millet in Jeypore for instance, enhanced productivity by 82% and monetary returns by 46%. Women and landless farmers gained access to land for cultivation of short-duration crops, benefiting from cultivation of suitable varieties, consumption of diverse species, and through intercropping across sites. To enhance sustainability, the project anchored all its efforts in community based organizations

Identification of lncRNAs in response to infection by Plasmodiophora brassicae in Brassica napus and development of lncRNA-based SSR markers

Clubroot resistance in spring canola has been introgressed from different Brassica sources; however, molecular mechanism underlying this resistance, especially the involvement of long non-coding RNAs (lncRNAs), yet to be understood. We identified 464 differentially expressed (DE) lncRNAs from the roots of clubroot resistant canola, carrying resistance on chromosome BnaA03, and susceptible canola lines challenged with Plasmodiophora brassicae pathotype 3. Pathway enrichment analysis showed that most of the target genes regulated by these DE lncRNAs belonged to plant-pathogen interaction and hormone signaling, as well as primary and secondary metabolic pathways. Comparative analysis of these lncRNAs with the previously reported 530 DE lncRNAs, identified using resistance located on BnaA08, detected 12 lncRNAs which showed a similar trend of upregulation in both types of resistant lines; these lncRNAs probably play a fundamental role in clubroot resistance. We identified SSR markers within 196 DE lncRNAs. Genotyping of two DH populations carrying resistance on BnaA03 identified a marker capable of detecting the resistance in 98% of the DH lines. To our knowledge, this is the first report of the identification of SSRs within the lncRNAs responsive to P. brassicae infection demonstrating the potential use of the lncRNAs in the breeding of Brassica crops.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Identification and expression analysis of <it>WRKY </it>transcription factor genes in canola <it>(Brassica napus </it>L.) in response to fungal pathogens and hormone treatments

<p>Abstract</p> <p>Background</p> <p>Members of plant WRKY transcription factor families are widely implicated in defense responses and various other physiological processes. For canola (<it>Brassica napus </it>L.), no WRKY genes have been described in detail. Because of the economic importance of this crop, and its evolutionary relationship to <it>Arabidopsis thaliana</it>, we sought to characterize a subset of canola <it>WRKY </it>genes in the context of pathogen and hormone responses.</p> <p>Results</p> <p>In this study, we identified 46 <it>WRKY </it>genes from canola by mining the expressed sequence tag (EST) database and cloned cDNA sequences of 38 <it>BnWRKY</it>s. A phylogenetic tree was constructed using the conserved WRKY domain amino acid sequences, which demonstrated that BnWRKYs can be divided into three major groups. We further compared <it>BnWRKYs </it>to the 72 <it>WRKY </it>genes from <it>Arabidopsis </it>and 91 <it>WRKY </it>from rice, and we identified 46 presumptive orthologs of <it>AtWRKY </it>genes. We examined the subcellular localization of four BnWRKY proteins using green fluorescent protein (GFP) and we observed the fluorescent green signals in the nucleus only.</p> <p>The responses of 16 selected <it>BnWRKY </it>genes to two fungal pathogens, <it>Sclerotinia sclerotiorum </it>and <it>Alternaria brassicae</it>, were analyzed by quantitative real time-PCR (qRT-PCR). Transcript abundance of 13 <it>BnWRKY </it>genes changed significantly following pathogen challenge: transcripts of 10 <it>WRKY</it>s increased in abundance, two <it>WRKY </it>transcripts decreased after infection, and one decreased at 12 h post-infection but increased later on (72 h). We also observed that transcript abundance of 13/16 <it>BnWRKY </it>genes was responsive to one or more hormones, including abscisic acid (ABA), and cytokinin (6-benzylaminopurine, BAP) and the defense signaling molecules jasmonic acid (JA), salicylic acid (SA), and ethylene (ET). We compared these transcript expression patterns to those previously described for presumptive orthologs of these genes in <it>Arabidopsis </it>and rice, and observed both similarities and differences in expression patterns.</p> <p>Conclusion</p> <p>We identified a set of 13 <it>BnWRKY </it>genes from among 16 <it>BnWRKY </it>genes assayed, that are responsive to both fungal pathogens and hormone treatments, suggesting shared signaling mechanisms for these responses. This study suggests that a large number of BnWRKY proteins are involved in the transcriptional regulation of defense-related genes in response to fungal pathogens and hormone stimuli.</p

The crystal structure of an octapeptide repeat of the Prion protein in complex with a fab fragment of the POM2 antibody

Prion diseases are progressive, infectious neurodegenerative disorders caused primarily by the misfolding of the cellular prion protein, PrP(c) , into an insoluble, protease-resistant, aggregated isoform termed PrP(sc) . In native conditions, PrP(c) has a structured C-terminal domain and a highly flexible N-terminal domain. A part of this N-terminal domain consists of 4-5 repeats of an unusual glycine rich, eight amino acids long peptide known as the octapeptide repeat (OR) domain. In this paper, we successfully report the first crystal structure of an octapeptide repeat of PrP(c) bound to the POM2 Fab antibody fragment. The structure was solved at a resolution of 2.3 Å by molecular replacement. While several studies have previously predicted a β-turn like structure of the unbound octapeptide repeats, our structure shows an extended conformation of the octapeptide repeat when bound to a molecule of the POM2 Fab indicating that the bound Fab disrupts any putative native β turn conformation of the octapeptide repeats. Encouraging results from several recent studies have shown that administering small molecule ligands or antibodies targeting the OR domain of PrP result in arresting the progress of peripheral prion infections both in ex vivo and in in vivo models. This makes the structural study of the interactions of POM2 Fab with the OR domain very important as it would help us to design smaller and tighter binding OR ligands