Analyses of pea necrotic yellow dwarf virus-encoded proteins.

Pea necrotic yellow dwarf virus (PNYDV) is a multipartite, circular, single-stranded DNA plant virus. PNYDV encodes eight proteins and the function of three of which remains unknown-U1, U2, and U4. PNYDV proteins cellular localization was analyzed by GFP tagging and bimolecular fluorescence complementation (BiFC) studies. The interactions of all eight PNYDV proteins were tested pairwise in planta (36 combinations in total). Seven interactions were identified and two (M-Rep with CP and MP with U4) were characterized further. MP and U4 complexes appeared as vesicle-like spots and were localized at the nuclear envelope and cell periphery. These vesicle-like spots were associated with the endoplasmatic reticulum. In addition, a nuclear localization signal (NLS) was mapped for U1, and a mutated U1 with NLS disrupted localized at plasmodesmata and therefore might also have a role in movement. Taken together, this study provides evidence for previously undescribed nanovirus protein-protein interactions and their cellular localization with novel findings not only for those proteins with unknown function, but also for characterized proteins such as the CP.

Plastidial thioredoxin z interacts with two fructokinase-like proteins in a thiol-dependent manner: evidence for an essential role in chloroplast development in Arabidopsis and Nicotiana benthamiana.

Here, we characterize a plastidial thioredoxin (TRX) isoform from Arabidopsis thaliana that defines a previously unknown branch of plastidial TRXs lying between x- and y-type TRXs and thus was named TRX z. An Arabidopsis knockout mutant of TRX z had a severe albino phenotype and was inhibited in chloroplast development. Quantitative real-time RT-PCR analysis of the mutant suggested that the expressions of genes that depend on a plastid-encoded RNA polymerase (PEP) were specifically decreased. Similar results were obtained upon virus-induced gene silencing (VIGS) of the TRX z ortholog in Nicotiana benthamiana. We found that two fructokinase-like proteins (FLN1 and FLN2), members of the pfkB-carbohydrate kinase family, were potential TRX z target proteins and identified conserved Cys residues mediating the FLN-TRX z interaction. VIGS in N. benthamiana and inducible RNA interference in Arabidopsis of FLNs also led to a repression of PEP-dependent gene transcription. Remarkably, recombinant FLNs displayed no detectable sugar-phosphorylating activity, and amino acid substitutions within the predicted active site imply that the FLNs have acquired a new function, which might be regulatory rather than metabolic. We were able to show that the FLN2 redox state changes in vivo during light/dark transitions and that this change is mediated by TRX z. Taken together, our data strongly suggest an important role for TRX z and both FLNs in the regulation of PEP-dependent transcription in chloroplasts.

Risk factors associated with necrotizing enterocolitis in preterm infants: A case-control study.

BACKGROUND: Necrotizing enterocolitis (NEC) is the most common life-threatening gastrointestinal emergency in prematurity. The pathophysiology is multifactorial and remains incompletely understood. Early diagnosis and treatment could reduce the risk of mortality and morbidity. We aimed to identify factors associated with NEC in preterm newborns. METHOD: This case-control study included all preterm newborns presenting with NEC and managed between January 1, 2009 and December 31, 2018 in the neonatal intensive care unit of Nancy. For each case, two controls were matched according to three criteria: gestational age (WG), date of birth, and mode of delivery. Antenatal, peripartum, and postnatal risk factors prior to NEC were analyzed. RESULTS: A total of 292 infants were involved in the study, 113 of whom had NEC. Mean gestational age for newborns with NEC was 29 WG, and mean birth weight, 1340 g. Only early-onset infection was identified as a significant risk factor for NEC (15% vs. 6.6% for infection p<0.04, and 28.3% vs. 16.4% p<0.02 for infection and sepsis, NEC vs. controls, respectively). Late-onset feeding and initial continuous enteral feeding were significantly associated with the occurrence of more severe NEC (p<0.02 and p = 0.03, respectively). CONCLUSION: The results of this study are consistent with intestinal dysbiosis being a risk factor for NEC. Early-onset infection was found to be a significant risk factor. Enteral feeding practice may also be associated with NEC.

Outcome of Very Premature Newborn Receiving an Early Second Dose of Surfactant for Persistent Respiratory Distress Syndrome.

Background: Infants presenting respiratory distress syndrome (RDS) not responding to surfactant often receive a second instillation. Few studies evaluated the consequences of this second administration. This study aimed at determining the outcome of infants presenting persistent RDS and receiving an early second dose of surfactant. Methods: Infants below 32 weeks' gestation who received a second dose of 100mg/kg of surfactant within the first 72 h of life, were retrospectively involved in this 42 months' study. They were matched to two controls receiving a single dose of 200mg/Kg based upon gender and gestational age. Results: 52/156 infants receiving two doses (Group 2-doses) were significantly more often SGA [22 (42%) vs. 21 (20%) p = 0.04] and outborn [29 (56%) vs. 13 (12%) p = 0.001]. They had received antenatal corticos teroid therapy less often [26 (50%) vs. 89 (86%) p = 0.001] and presented more severe RDS based upon FiO2 level, oxygenation index and radiography. Group 2-doses survival was lower (65.4% vs. 79.6 % p < 0.1) but surviving infants did not have different morbidity than controls. Discussion: Premature newborn receiving a second dose of surfactant had adverse antenatal characteristics, presented more severe RDS and only partially responded to the first dose. Outcomes of surviving infants who received 2 doses of surfactant were comparable to others.

A case with coincidental diagnosis of primary central nervous system lymphoma and lymph node sarcoidosis.

Primary central nervous system lymphoma (PCNSL) is rare. Clinical and histological differential diagnosis of systemic lymphoma and sarcoidosis continues to be a challenge. The first case report in the German and English literature of PCNSL and synchronous sarcoidosis is presented. Synchronous mediastinal lymphadenopathy suggestive of non-Hodgkin's lymphoma (NHL) or sarcoidosis was noted. Both conditions require alternative therapeutic and prognostic considerations to PCNSL. A regime of intrathecal and adjuvant systemic chemotherapy led to transient clinical improvement prior to the patient's demise through overwhelming sepsis and multiorgan failure. Post mortem findings confirmed synchronous PCNSL with mediastinal lymph node sarcoidosis.

The stress granule component G3BP is a novel interaction partner for the nuclear shuttle proteins of the nanovirus pea necrotic yellow dwarf virus and geminivirus abutilon mosaic virus.

Stress granules (SGs) are structures within cells that regulate gene expression during stress response, e.g. viral infection. In mammalian cells assembly of SGs is dependent on the Ras-GAP SH3-domain-binding protein (G3BP). The C-terminal domain of the viral nonstructural protein 3 (nsP3) of Semliki Forest virus (SFV) forms a complex with mammalian G3BP and sequesters it into viral RNA replication complexes in a manner that inhibits the formation of SGs. The binding domain of nsP3 to HsG3BP was mapped to two tandem 'FGDF' repeat motifs close to the C-terminus of the viral proteins. It was speculated that plant viruses employ a similar strategy to inhibit SG function. This study identifies an Arabidopsis thaliana NTF2-RRM domain-containing protein as a G3BP-like protein (AtG3BP), which localizes to plant SGs. Moreover, the nuclear shuttle protein (NSP) of the begomovirus abutilon mosaic virus (AbMV), which harbors a 'FVSF'-motif at its C-terminal end, interacts with the AtG3BP-like protein, as does the 'FNGSF'-motif containing NSP of pea necrotic yellow dwarf virus (PNYDV), a member of the Nanoviridae family. We therefore propose that SG formation upon stress is conserved between mammalian and plant cells and that plant viruses may follow a similar strategy to inhibit plant SG function as it has been shown for their mammalian counterparts.

Begomoviral Movement Protein Effects in Human and Plant Cells: Towards New Potential Interaction Partners.

Geminiviral single-stranded circular DNA genomes replicate in nuclei so that the progeny DNA has to cross both the nuclear envelope and the plasmodesmata for systemic spread within plant tissues. For intra- and intercellular transport, two proteins are required: a nuclear shuttle protein (NSP) and a movement protein (MP). New characteristics of ectopically produced Abutilon mosaic virus (AbMV) MP (MPAbMV), either authentically expressed or fused to a yellow fluorescent protein or epitope tags, respectively, were determined by localization studies in mammalian cell lines in comparison to plant cells. Wild-type MPAbMV and the distinct MPAbMV: reporter protein fusions appeared as curled threads throughout mammalian cells. Co-staining with cytoskeleton markers for actin, intermediate filaments, or microtubules identified these threads as re-organized microtubules. These were, however, not stabilized by the viral MP, as demonstrated by nocodazole treatment. The MP of a related bipartite New World begomovirus, Cleome leaf crumple virus (ClLCrV), resulted in the same intensified microtubule bundling, whereas that of a nanovirus did not. The C-terminal section of MPAbMV, i.e., the protein's oligomerization domain, was dispensable for the effect. However, MP expression in plant cells did not affect the microtubules network. Since plant epidermal cells are quiescent whilst mammalian cells are proliferating, the replication-associated protein RepAbMV protein was then co-expressed with MPAbMV to induce cell progression into S-phase, thereby inducing distinct microtubule bundling without MP recruitment to the newly formed threads. Co-immunoprecipitation of MPAbMV in the presence of RepAbMV, followed by mass spectrometry identified potential novel MPAbMV-host interaction partners: the peptidyl-prolyl cis-trans isomerase NIMA-interacting 4 (Pin4) and stomatal cytokinesis defective 2 (SCD2) proteins. Possible roles of these putative interaction partners in the begomoviral life cycle and cytoskeletal association modes are discussed.

No influence on disease progression of non-HLA susceptibility genes in MS.

Recently, several non-HLA loci have been shown to be convincingly associated with Multiple Sclerosis (MS) susceptibility, assumingly indicating important pathways in the pathogenesis. A genotype influence on disease outcome measures by these genes would support a role of these pathways in ongoing tissue damage. Here, however, we report a consistent dissociation between causation and progression for five non-HLA genotypes (IL7R, IL2RA, CLEC16A, CD226 and SH2B3) in 1776 Scandinavian MS patients.

HLA-DRB115 and cerebrospinal-fluid-specific oligoclonal immunoglobulin G bands lower age at attainment of important disease milestones in multiple sclerosis.

Carriage of HLA-DRB1*15 is the most important genetic risk factor in multiple sclerosis (MS), while CSF-specific oligoclonal immunoglobulin G bands (OCB) constitute the most sensitive biochemical marker for diagnosing MS. We demonstrated in an earlier study the interdependence of HLA-DRB1 genotype and OCB status; the effect of these phenotypic features on MS prognosis remains controversial, however. We investigated by survival analysis the impact of each variable on age at two important MS milestones: onset of clinical symptoms and an Expanded Disability Status Scale (EDSS) score of 6.0. Both carriage of HLA-DRB1*15 and the presence of OCB hastened attainment of EDSS 6.0.

Genetic variation in the KIF1B locus influences susceptibility to multiple sclerosis.

The few loci associated with multiple sclerosis (MS) are all related to immune function. We report a GWA study identifying a new locus replicated in 2,679 cases and 3,125 controls. An rs10492972[C] variant located in the KIF1B gene was associated with MS with an odds ratio of 1.35 (P = 2.5 x 10(-10)). KIF1B is a neuronally expressed gene plausibly implicated in the irreversible axonal loss characterizing MS in the long term.

HFA1 encoding an organelle-specific acetyl-CoA carboxylase controls mitochondrial fatty acid synthesis in Saccharomyces cerevisiae.

The Saccharomyces cerevisiae gene, HFA1, encodes a >250-kDa protein, which is required for mitochondrial function. Hfa1p exhibits 72% overall sequence similarity (54% identity) to ACC1-encoded yeast cytoplasmic acetyl-CoA carboxylase. Nevertheless, HFA1 and ACC1 functions are not overlapping because mutants of the two genes have different phenotypes and do not complement each other. Whereas ACC1 is involved in cytoplasmic fatty acid synthesis, the phenotype of hfa1Delta disruptants resembles that of mitochondrial fatty-acid synthase mutants. They fail to grow on lactate or glycerol, and the mitochondrial cofactor, lipoic acid, is reduced to <10% of its normal cellular concentration. Other than Acc1p, the N-terminal sequence of Hfa1p comprises a canonical mitochondrial targeting signal together with a matrix protease cleavage site. Accordingly, the HFA1-encoded protein was specifically assigned by Western blotting of appropriate cell fractions to the mitochondrial compartment. Removal of the mitochondrial targeting sequence abolished the competence of HFA1 DNA to complement hfal null mutants. Conversely and in contrast to the intact HFA1 sequence, the signal sequence-free HFA1 gene complemented the mutational loss of cytoplasmic acetyl-CoA carboxylase. Expression of HFA1 under the control of the ACC1 promoter restored cellular ACC activity in ACC1-defective yeast mutants to wild type levels. From this finding, it is concluded that HFA1 encodes a specific mitochondrial acetyl-CoA carboxylase providing malonyl-CoA for intraorganellar fatty acid and, in particular, lipoic acid synthesis.