Clubroot resistance QTL are modulated by nitrogen input in <Emphasis Type="Italic">Brassica napus</Emphasis>

Key message

Nitrogen levels can modulate the effectiveness of clubroot resistance in an isolate- and host-specific manner. While the same QTL were detected under high and low nitrogen, their effects were altered.

Abstract

Clubroot, caused by Plasmodiophora brassicae, is one of the most damaging diseases of oilseed rape and is known to be affected by nitrogen fertilization. However, the genetic factors involved in clubroot resistance have not been characterized under nitrogen-limiting conditions. This study aimed to assess the variability of clubroot resistance under different nitrogen levels and to characterize the impact of nitrogen supply on genetic resistance factors. Linkage analyses and a genome-wide association study were conducted to detect QTL for clubroot resistance and evaluate their sensitivity to nitrogen. The clubroot response of a set of 92 diverse oilseed rape accessions and 108 lines derived from a cross between ‘Darmor-bzh’ (resistant) and ‘Yudal’ (susceptible) was studied in the greenhouse under high- and low-nitrogen conditions, following inoculation with the P. brassicae isolates eH and K92-16. Resistance to each isolate was controlled by a major QTL and a few small-effects QTL. While the same QTL were detected under both high and low nitrogen, their effects were altered. Clubroot resistance to isolate eH, but not K92-16, was greater under a low-N supply versus a high-N supply. New sources of resistance were found among the oilseed rape accessions under both low and high-N conditions. The results are discussed relative to the literature and from a crop improvement perspective.

     

A protein activator for the adenylate cyclase of Bordetella pertussis.

The activity of Bordetella pertussis extracytoplasmic adenylate cyclase is 100-fold higher in organisms grown on blood agar than in those grown in synthetic medium. This increase in activity is due to in vivo activation of the enzyme by a factor present in erythrocytes. Activation also occurs in killed or disrupted organisms. The activator can be separated from heme proteins and has been purified approximately 100-fold from erythrocytes, yielding material of approximately 105,000 daltons. It is sensitive to trypsin and alpha-chymotrypsin and exhibits considerable heat stability. Activation of cyclase in intact B. pertussis organisms exhibits a lag of 3 to 4 min and is not reversed by washing. Response to the activator decreases with increasing purification of the adenylate cyclase and is absent in the pure enzyme. The activation does not appear to be proteolytic and does not appear to change access to the substrate, ATP. The activator has no effect on a number of eukaryotic cyclases. We conclude that this is a new type of activation and that the activator differs from all those previously described.     

Protein translocation into host epithelial cells by infecting enteropathogenic Escherichia coli

Enteropathogenic Escherichia coli (EPEC) causes diarrhoea in young children. EPEC induces the formation of actin pedestal in infected epithelial cells. A type III protein secretion system and several proteins that are secreted by this system, including EspB, are involved in inducing the formation of the actin pedestals. We have demonstrated that contact of EPEC with HeLa cells is associated with the induction of production and secretion of EspB. Shortly after infection, EPEC initiates translocation of EspB, and EspB fused to the CyaA reporter protein (EspB–CyaA), into the host cell. The translocated EspB was distributed between the membrane and the cytoplasm of the host cell. Translocation was strongly promoted by attachment of EPEC to the host cell, and both attachment factors of EPEC, intimin and the bundle-forming pili, were needed for full translocation efficiency. Translocation and secretion of EspB and EspB–CyaA were abolished in mutants deficient in components of the type III protein secretion system, including sepA and sepB mutants. EspB–CyaA was secreted but not translocated by an espB mutant. These results indicate that EspB is both translocated and required for protein translocation by EPEC.     

Epitope mapping of human aromatic L-amino acid decarboxylase

Autoimmune polyendocrine syndrome type I (APS I) is a rare hereditary condition considered a model disease for organ specific autoimmunity. A wide range of autoantibodies targeting antigens present in the affected organs have been identified. Autoantibodies against aromatic L-amino acid decarboxylase (AADC) are present in about 50% of APS I patients. In order to increase our understanding of autoantibody specificity in APS I, the aim of the present study was to localize target regions on AADC recognized by sera from APS I patients. Using several complementing strategies, we have shown that autoantibodies against AADC mainly recognize conformational epitopes. The major antigenic determinants were detected N-terminally to amino acid residue 237. Replacement of amino acids 227-230 (ERDK) with alanine residues reduced the reactivity towards AADC by >80% in all patient sera tested, suggesting that amino acids 227-230 are an important part of an immunodominant epitope.     

Dermal microvascular endothelial cells express the 180-kDa macrophage mannose receptor in situ and in vitro

Expression of the 180-kDa mannose receptor (MR) is mainly found on cells of the macrophage lineage. MR mediates the uptake of micro-organisms and host-derived glycoproteins. We demonstrate that endothelium of the human skin in situ and dermal microvascular endothelial cells (DMEC) in vitro expressed MR at both the protein and mRNA levels. In contrast, HUVEC were consistently negative for MR expression. DMEC internalized dextran as well as Escherichia coli by the way of MR into acidic phagosomes, only a few of which fused with CD63- and lysosomal-associated membrane glycoprotein-2-positive lysosomes. This contrasts with the situation in monocyte-derived dendritic cells, where almost all of the MR-Ag complexes reached CD63- and lysosomal-associated membrane glycoprotein-2-positive compartments, indicating differences in the phagolysosomal fusion rate between DMEC and dendritic cells. In conclusion, DMEC express functional MR, a finding that corroborates a role of skin endothelium in Ag capture/clearing.     

Characterization of Tubulin in Mouse Brain Myelin

Analysis of mouse brain myelin by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that in the high-molecular-weight range it contained, besides the Wolfgram protein doublet, proteins comigrating with actin and with both subunits of tubulin. The occurrence of these alpha and beta subunits was confirmed by peptide mapping in myelin analyzed by two-dimensional electrophoresis. This tubulin did not arise from an artifactual binding of soluble brain tubulin to the myelin fraction: addition of exogenously labeled tubulin to brain homogenates proved that during myelin isolation by the procedure of Norton and Poduslo (1973) the contaminating tubulin was washed out. On the other hand, the distribution of tubulin isoforms in myelin was investigated by isoelectric focusing and compared with the distribution of the 21 isoforms listed for the whole brain soluble tubulin. It was shown that many isoforms were found in myelin (three isoforms for the alpha subunit and nine for the beta subunit), and that some isoforms were represented both in myelin and in soluble tubulin, but in different relative proportions.     

Interdependence of hemagglutinin glycosylation and neuraminidase as regulators of influenza virus growth: a study by reverse genetics

The hemagglutinin (HA) of fowl plague virus A/FPV/Rostock/34 (H7N1) carries two N-linked oligosaccharides attached to Asn123 and Asn149 in close vicinity to the receptor-binding pocket. In previous studies in which HA mutants lacking either one (mutants G1 and G2) or both (mutant G1,2) glycosylation sites had been expressed from a simian virus 40 vector, we showed that these glycans regulate receptor binding affinity (M. Ohuchi, R. Ohuchi, A. Feldmann, and H. D. Klenk, J. Virol. 71:8377-8384, 1997). We have now investigated the effect of these mutations on virus growth using recombinant viruses generated by an RNA polymerase I-based reverse genetics system. Two reassortants of influenza virus strain A/WSN/33 were used as helper viruses to obtain two series of HA mutant viruses differing only in the neuraminidase (NA). Studies using N1 NA viruses revealed that loss of the oligosaccharide from Asn149 (mutant G2) or loss of both oligosaccharides (mutant G1,2) has a pronounced effect on virus growth in MDCK cells. Growth of virus lacking both oligosaccharides from infected cells was retarded, and virus yields in the medium were decreased about 20-fold. Likewise, there was a reduction in plaque size that was distinct with G1,2 and less pronounced with G2. These effects could be attributed to a highly impaired release of mutant progeny viruses from host cells. In contrast, with recombinant viruses containing N2 NA, these restrictions were much less apparent. N1 recombinants showed lower neuraminidase activity than N2 recombinants, indicating that N2 NA is able to partly overrule the high-affinity binding of mutant HA to the receptor. These results demonstrate that N-glycans flanking the receptor-binding site of the HA molecule are potent regulators of influenza virus growth, with the glycan at Asn149 being dominant and that at Asn123 being less effective. In addition, we show here that HA and NA activities need to be highly balanced in order to allow productive influenza virus infection.     

Sensitization of rat alveolar macrophages to enhanced TNF-alpha release by in vivo treatment with dexamethasone.

Treatment of rats with dexamethasone rapidly induced a marked weight loss which occurred within 3 days and persisted for several weeks. The cachectic state was paralleled by increased serum levels of triglycerides, albumin, and protein and a strong reduction of blood mononuclear leukocytes. In lung sections, an increased number of mononuclear giant cells was found but no bacteria, fungi, or Pneumocystis carinii organisms. Quite strikingly, alveolar macrophages from dexamethasone-treated rats, but not from control animals, were highly sensitive to LPS and released large amounts of TNF-alpha ex vivo. Also under in vivo conditions, high TNF-alpha serum concentrations were found in dexamethasone-treated but not control rats when examined 1 1/2 hr after an intravenous LPS injection. These data suggest that the glucocorticoid-induced cachexia of rats may be linked, at least in part, to readily inducible TNF-alpha release from primed macrophages.     

The influenza A virus NS1 protein inhibits activation of Jun N-terminal kinase and AP-1 transcription factors

The influenza A virus nonstructural NS1 protein is known to modulate host cell gene expression and to inhibit double-stranded RNA (dsRNA)-mediated antiviral responses. Here we identify NS1 as the first viral protein that antagonizes virus- and dsRNA-induced activation of the stress response-signaling pathway mediated through Jun N-terminal kinase.