Nuclear proteome of virus-infected and healthy potato leaves

BackgroundInfection of plants by viruses interferes with expression and subcellular localization of plant proteins. Potyviruses comprise the largest and most economically damaging group of plant-infecting RNA viruses. In virus-infected cells, at least two potyviral proteins localize to nucleus but reasons remain partly unknown.ResultsIn this study, we examined changes in the nuclear proteome of leaf cells from a diploid potato line (Solanum tuberosum L.) after infection with potato virus A (PVA; genus Potyvirus; Potyviridae) and compared the data with that acquired for healthy leaves. Gel-free liquid chromatography-coupled to tandem mass spectrometry was used to identify 807 nuclear proteins in the potato line v2-108; of these proteins, 370 were detected in at least two samples of healthy leaves. A total of 313 proteins were common in at least two samples of healthy and PVA-infected leaves; of these proteins, 8 showed differential accumulation. Sixteen proteins were detected exclusively in the samples from PVA-infected leaves, whereas other 16 proteins were unique to healthy leaves. The protein Dnajc14 was only detected in healthy leaves, whereas different ribosomal proteins, ribosome-biogenesis proteins, and RNA splicing-related proteins were over-represented in the nuclei of PVA-infected leaves. Two virus-encoded proteins were identified in the samples of PVA-infected leaves.ConclusionsOur results show that PVA infection alters especially ribosomes and splicing-related proteins in the nucleus of potato leaves. The data increase our understanding of potyvirus infection and the role of nucleus in infection. To our knowledge, this is the first study of the nuclear proteome of potato leaves and one of the few studies of changes occurring in nuclear proteomes in response to plant virus infection.Peer reviewe

Thoracic high resolution CT using the modified VetMousetrap (TM) device is a feasible method for diagnosing canine idiopathic pulmonary fibrosis in awake West Highland White Terriers

Canine idiopathic pulmonary fibrosis is a chronic, progressive interstitial lung disease particularly prevalent in West Highland White Terriers. In the present prospective pilot study, we evaluated the feasibility of modified VetMousetrap (TM) device in high resolution CT to detect idiopathic pulmonary fibrosis in West Highland White Terriers. Twelve awake West Highland White Terriers with canine idiopathic pulmonary fibrosis and 24 clinically healthy West Highland White Terriers were scanned using a helical dual slice scanner utilizing VetMousetrap (TM) device without or with minimal chemical restraint with butorphanol. Three evaluators blindly assessed the images for image quality and the presence of canine idiopathic pulmonary fibrosis related imaging findings such as ground glass opacity and reticular opacities. Additionally, the attenuation of the lung was quantified with ImageJ software using histogram analysis of density over the lung fields. Computed tomography was successfully completed and motion artifact ranked in statistical analysis barely noticeable to mild in all dogs. The agreement between imaging findings and clinical status was very good with overall kappa value 0.91 and percentage of agreement of 94%. There was also very good intraobserver (kappa(range) = 0.79-0.91) and interobserver agreement (kappa = 0.94). Moderate to severe ground glass opacity was present in all affected dogs. In the ImageJ analysis, a significant difference in lung attenuation between the study groups was observed. We conclude that modified VetMousetrap (TM) device is applicable in diagnosing canine idiopathic pulmonary fibrosis in awake West Highland White Terriers avoiding anesthetic risk in these often severely hypoxic patients.Peer reviewe

Protocol: Optimised methodology for isolation of nuclei from leaves of species in the Solanaceae and Rosaceae families

Peer reviewe

Viral RNase3 Co-Localizes and Interacts with the Antiviral Defense Protein SGS3 in Plant Cells

Sweet potato chlorotic stunt virus (SPCSV; family Closteroviridae) encodes a Class 1 RNase III endoribonuclease (RNase3) that suppresses post-transcriptional RNA interference (RNAi) and eliminates antiviral defense in sweetpotato plants (Ipomoea batatas). For RNAi suppression, RNase3 cleaves double-stranded small interfering RNAs (ds-siRNA) and long dsRNA to fragments that are too short to be utilized in RNAi. However, RNase3 can suppress only RNAi induced by sense RNA. Sense-mediated RNAi involves host suppressor of gene silencing 3 (SGS3) and RNA-dependent RNA polymerase 6 (RDR6). In this study, subcellular localization and host interactions of RNase3 were studied in plant cells. RNase3 was found to interact with SGS3 of sweetpotato and Arabidopsis thaliana when expressed in leaves, and it localized to SGS3/RDR6 bodies in the cytoplasm of leaf cells and protoplasts. RNase3 was also detected in the nucleus. Co-expression of RNase3 and SGS3 in leaf tissue enhanced the suppression of RNAi, as compared with expression of RNase3 alone. These results suggest additional mechanisms needed for efficient RNase3-mediated suppression of RNAi and provide new information about the subcellular context and phase of the RNAi pathway in which RNase3 realizes RNAi suppression.Peer reviewe

Pathogenic seedborne viruses are rare but Phaseolus vulgaris endornaviruses are common in bean varieties grown in Nicaragua and Tanzania

Common bean (Phaseolus vulgaris) is an annual grain legume that was domesticated in Mesoamerica (Central America) and the Andes. It is currently grown widely also on other continents including Africa. We surveyed seedborne viruses in new common bean varieties introduced to Nicaragua (Central America) and in landraces and improved varieties grown in Tanzania (eastern Africa). Bean seeds, harvested from Nicaragua and Tanzania, were grown in insect-controlled greenhouse or screenhouse, respectively, to obtain leaf material for virus testing. Equal amounts of total RNA from different samples were pooled (30-36 samples per pool), and small RNAs were deep-sequenced (Illumina). Assembly of the reads (21-24 nt) to contiguous sequences and searches for homologous viral sequences in data-bases revealed Phaseolus vulgaris endornavirus 1 (PvEV-1) and PvEV-2 in the bean varieties in Nicaragua and Tanzania. These viruses are not known to cause symptoms in common bean and are considered non-pathogenic. The small-RNA reads from each pool of samples were mapped to the previously characterized complete PvEV-1 and PvEV-2 sequences (genome lengths ca. 14 kb and 15 kb, respectively). Coverage of the viral genomes was 87.9-99.9%, depending on the pool. Coverage per nucleotide ranged from 5 to 471, confirming virus identification. PvEV-1 and PvEV-2 are known to occur in Phaseolus spp. in Central America, but there is little previous information about their occurrence in Nicaragua, and no information about occurrence in Africa. Aside from Cowpea mild mosaic virus detected in bean plants grown from been seeds harvested from one region in Tanzania, no other pathogenic seedborne viruses were detected. The low incidence of infections caused by pathogenic viruses transmitted via bean seeds may be attributable to new, virus-resistant CB varieties released by breeding programs in Nicaragua and Tanzania.Peer reviewe

Matrix metalloproteinase-2,-7, and-9 activities in dogs with idiopathic pulmonary fibrosis compared to healthy dogs and dogs with other respiratory diseases

Background Canine idiopathic pulmonary fibrosis (CIPF) is a chronic, interstitial lung disease that mainly affects West Highland white terriers (WHWTs) and is characterized by excessive deposition of extracellular matrix (ECM) in the lung. Matrix metalloproteinases (MMPs) participate in remodeling of ECM. Objectives To compare metalloproteinase-2, -7 and -9 activities in blood or bronchoalveolar lavage fluid (BALF) samples or both of CIPF WHWTs with healthy WHWTs, healthy dogs of other breeds, and dogs with other lung diseases and determine if these MMPs could be used as diagnostic and prognostic markers for CIPF. Animals Forty-four CIPF WHWTs, 24 dogs with chronic bronchitis (CB), 17 with eosinophilic bronchopneumopathy (EBP), 10 with bacterial pneumonia, 39 healthy WHWTs, and 35 healthy dogs of other breeds. Methods Cross-sectional observational study. Pro-MMP and active MMP activities were analyzed by zymography. Results In serum, significantly higher (P <.01) pro-MMP-7 activities were observed in CIPF WHWTs compared to healthy dogs of other breeds, dogs with CB and dogs with EBP. In BALF of CIPF WHWTs, both pro-MMP-9 and pro-MMP-2 activities were significantly higher (P <.01) compared to healthy WHWTs, but these differences were not detected in plasma. The CIPF WHWTs had significantly higher (P <.05) activities of pro-MMP-9 compared to dogs with CB and of pro-MMP-2 compared to dogs with CB and EBP. No statistically significant prognostic factors were observed in CIPF WHWTs. Conclusions and clinical relevance Serum MMP-7 and BALF MMP-2 and -9 potentially may be useful diagnostic markers but not prognostic markers for CIPF.Peer reviewe