Nondegradative role of Atg5-Atg12/ Atg16L1 autophagy protein complex in antiviral activity of interferon gamma

Host resistance to viral infection requires type I (α/β) and II (γ) interferon (IFN) production. Another important defense mechanism is the degradative activity of macroautophagy (herein autophagy), mediated by the coordinated action of evolutionarily conserved autophagy proteins (Atg). We show that the Atg5-Atg12/Atg16L1 protein complex, whose prior known function is in autophagosome formation, is required for IFNγ-mediated host defense against murine norovirus (MNV) infection. Importantly, the direct antiviral activity of IFNγ against MNV in macrophages required Atg5-Atg12, Atg7, and Atg16L1, but not induction of autophagy, the degradative activity of lysosomal proteases, fusion of autophagosomes and lysosomes, or the Atg8-processing protein Atg4B. IFNγ, via Atg5-Atg12/Atg16L1, inhibited formation of the membranous cytoplasmic MNV replication complex, where Atg16L1 localized. Thus, the Atg5-Atg12/Atg16L1 complex performs a pivotal, nondegradative role in IFNγ-mediated antiviral defense, establishing that multicellular organisms have evolved to use portions of the autophagy pathway machinery in a cassette-like fashion for host defense.     

The stability and aggregation of ovine prion protein associated with classical and atypical scrapie correlates with the ease of unwinding of helix-2

Susceptibility to scrapie disease in sheep, the archetypal prion disease, correlates with polymorphisms within the ovine PrP (prion-related protein) gene. The VRQ (Val136Arg154Gln171) and AL141RQ (Ala136Leu141Arg154Gln171) allelic variants are associated with classical scrapie, whereas the ARR (Ala136Arg154Arg171), AF141RQ (Ala136Phe141Arg154Gln171) and AHQ (Ala136His154Gln171) allelic variants are associated with atypical scrapie. Recent studies have suggested that there are differences in the stability of PrPSc (abnormal disease-specific conformation of PrP) associated with these different forms of scrapie. To address which structural features of ovine PrP may contribute to this difference, in the present study we have investigated the conformational stability and susceptibility to aggregation of allelic variants of ovine PrP associated with classical or atypical scrapie. We find that the melting temperature of ovine recombinant VRQ and AL141RQ PrP is higher than that of AF141RQ, AHQ and ARR. In addition, monoclonal-antibody studies show that the region around helix-1 of VRQ and AL141RQ is less accessible compared with other ovine PrP allelic variants. Furthermore, the extent of both the structural change to copper-ion-treatment and denaturant-induced aggregation was reduced in PrP associated with atypical scrapie compared with PrP associated with classical scrapie. Through the use of molecular dynamics simulations we have found that these biochemical and biophysical properties of ovine PrP correlate with the ease of unwinding of helix-2 and a concurrent conformational change of the helix-2-helix-3 loop. These results reveal significant differences in the overall stability and potential for aggregation of different allelic variants of ovine PrP and consequently have implications for the differences in stability of PrPSc associated with classical and atypical scrapie.     

Definition of common carotid wall thickness affects risk classification in relation to degree of internal carotid artery stenosis: the Plaque At RISK (PARISK) study

Background

Mean or maximal intima-media thickness (IMT) is commonly used as surrogate endpoint in intervention studies. However, the effect of normalization by surrounding or median IMT or by diameter is unknown. In addition, it is unclear whether IMT inhomogeneity is a useful predictor beyond common wall parameters like maximal wall thickness, either absolute or normalized to IMT or lumen size. We investigated the interrelationship of common carotid artery (CCA) thickness parameters and their association with the ipsilateral internal carotid artery (ICA) stenosis degree.

Methods

CCA thickness parameters were extracted by edge detection applied to ultrasound B-mode recordings of 240 patients. Degree of ICA stenosis was determined from CT angiography.

Results

Normalization of maximal CCA wall thickness to median IMT leads to large variations. Higher CCA thickness parameter values are associated with a higher degree of ipsilateral ICA stenosis (p?<?0.001), though IMT inhomogeneity does not provide extra information. When the ratio of wall thickness and diameter instead of absolute maximal wall thickness is used as risk marker for having moderate ipsilateral ICA stenosis (>50%), 55 arteries (15%) are reclassified to another risk category.

Conclusions

It is more reasonable to normalize maximal wall thickness to end-diastolic diameter rather than to IMT, affecting risk classification and suggesting modification of the Mannheim criteria.

Trial registration

Clinical trials.gov NCT01208025.

     

<i>Streptomyces</i> PRIO41 as plant growth promoter of jalapeño pepper plants and as biocontrol agent of <i>Fusarium</i>

Chili pepper is one of the main crops of economic importance in Mexico, and Fusarium wilting is a disease that limits its production. In addition, the inappropriate use of agrochemicals in farming activities generate environmental and health problems. Therefore, in this study the effectiveness of Streptomyces sp PRIO41 was evaluated as a (1) biocontrol agent of Fusarium spp and (2) plant growth promoter bacteria. Assays of pathogenicity and virulence of Fusarium spp. in jalapeño pepper seeds, and interactions of these pathogens with Streptomyces PRIO41 were evaluated under two nutritional conditions. In the greenhouse, the effectiveness of Streptomyces sp. PRIO41 was determined as a (1) biocontrol of Fusarium, and (2) plant growth promoter of wilt of pepper plants. The results showed that all fungal isolates caused symptoms in pepper seeds and seedlings with different degrees of virulence. Interactions in vitro showed that Streptomyces showed the most effective range of virulence against Fusarium isolates in the poor medium (37.6%-100%), with fungicidal effects in some cases. In the greenhouse, Streptomyces PRIO41 reduced Fusarium wilting up to a 40%, and positively affected all vegetative growth parameters, particularly plant height, leaf area, root length, and leaf and root dry biomasses. This study showed the potential of Streptomyces PRIO41 as a biocontrol agent of Fusarium spp., and as a biofertilizer of pepper plants.     

A covariotide model explains apparent phylogenetic structure of oxygenic photosynthetic lineages

The aims of the work were (1) to develop statistical tests to identify whether substitution takes place under a covariotide model in sequences used for phylogenetic inference and (2) to determine the influence of covariotide substitution on phylogenetic trees inferred for photosynthetic and other organisms. (Covariotide and covarion models are ones in which sites that are variable in some parts of the underlying tree are invariable in others and vice versa.) Two tests were developed. The first was a contingency test, and the second was an inequality test comparing the expected number of variable sites in two groups with the observed number. Application of these tests to 16S rDNA and tufA sequences from a range of nonphotosynthetic prokaryotes and oxygenic photosynthetic prokaryotes and eukaryotes suggests the occurrence of a covariotide mechanism. The degree of support for partitioning of taxa in reconstructed trees involving these organisms was determined in the presence or absence of sites showing particular substitution patterns. This analysis showed that the support for splits between (1) photosynthetic eukaryotes and prokaryotes and (2) photosynthetic and nonphotosynthetic organisms could be accounted for by patterns arising from covariotide substitution. We show that the additional problem of compositional bias in sequence data needs to be considered in the context of patterns of covariotide/covarion substitution. We argue that while covariotide or covarion substitution may give rise to phylogenetically informative patterns in sequence data, this may not always be so.      

Myocardial dysfunction in rheumatoid arthritis: epidemiology and pathogenesis

Data from population- and clinic-based epidemiologic studies of rheumatoid arthritis patients suggest that individuals with rheumatoid arthritis are at risk for developing clinically evident congestive heart failure. Many established risk factors for congestive heart failure are over-represented in rheumatoid arthritis and likely account for some of the increased risk observed. In particular, data from animal models of cytokine-induced congestive heart failure have implicated the same inflammatory cytokines produced in abundance by rheumatoid synovium as the driving force behind maladaptive processes in the myocardium leading to congestive heart failure. At present, however, the direct effects of inflammatory cytokines (and rheumatoid arthritis therapies) on the myocardia of rheumatoid arthritis patients are incompletely understood.     

In silico single strand melting curve: a new approach to identify nucleic acid polymorphisms in Totiviridae

Background

The PCR technique and its variations have been increasingly used in the clinical laboratory and recent advances in this field generated new higher resolution techniques based on nucleic acid denaturation dynamics. The principle of these new molecular tools is based on the comparison of melting profiles, after denaturation of a DNA double strand. Until now, the secondary structure of single-stranded nucleic acids has not been exploited to develop identification systems based on PCR. To test the potential of single-strand RNA denaturation as a new alternative to detect specific nucleic acid variations, sequences from viruses of the Totiviridae family were compared using a new in silico melting curve approach. This family comprises double-stranded RNA virus, with a genome constituted by two ORFs, ORF1 and ORF2, which encodes the capsid/RNA binding proteins and an RNA-dependent RNA polymerase (RdRp), respectively.

Results

A phylogenetic tree based on RdRp amino acid sequences was constructed, and eight monophyletic groups were defined. Alignments of RdRp RNA sequences from each group were screened to identify RNA regions with conserved secondary structure. One region in the second half of ORF2 was identified and individually modeled using the RNAfold tool. Afterwards, each DNA or RNA sequence was denatured in silico using the softwares MELTSIM and RNAheat that generate melting curves considering the denaturation of a double stranded DNA and single stranded RNA, respectively. The same groups identified in the RdRp phylogenetic tree were retrieved by a clustering analysis of the melting curves data obtained from RNAheat. Moreover, the same approach was used to successfully discriminate different variants of Trichomonas vaginalis virus, which was not possible by the visual comparison of the double stranded melting curves generated by MELTSIM.

Conclusion

In silico analysis indicate that ssRNA melting curves are more informative than dsDNA melting curves. Furthermore, conserved RNA structures may be determined from analysis of individuals that are phylogenetically related, and these regions may be used to support the reconstitution of their phylogenetic groups. These findings are a robust basis for the development of in vitro systems to ssRNA melting curves detection.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2105-15-243) contains supplementary material, which is available to authorized users.