Constitutive expression of IL-22 in the human intervertebral disc and its reduction by exposure to pro-inflammatory cytokines in vitro

The importance of cytokines in disc degeneration is well recognized. Little is known about IL-22 expression in the human intervertebral disc. We investigated IL-22 immuno-localization in disc tissue, and molecular expression and production of IL-22 by annulus cells cultured in three-dimensional (3D) culture. We examined human disc tissue using immunohistochemistry and we cultured isolated annulus cells in 3D to analyze IL-22 expression and production, and its receptor, IL-22R, in conditioned media. Ingenuity pathway analysis (IPA) also was used to identify significant gene expression networks within the molecular data. IL-22 and IL-22R were immunolocalized in many cells in the human outer and inner annulus; fewer cells exhibited localization in the nucleus. Three-dimensional culture of annulus cells demonstrated production of IL-22 in conditioned media; exposure to IL-1ß or TNF-α significantly reduced IL-22 levels. Significant decreases also were identified in conditioned media assayed for IL-22R in TNF-α treated cells. IPA analysis showed that IL-22 ranked among the top canonical pathways. We found constitutive expression and production of IL-22 and IL-22R in the disc, which expands our understanding of the effect of pro-inflammatory cytokines on IL-22 expression and production. Three-dimensional cultured annulus cells exposed to IL-1ß or TNF produced significantly lower levels of IL-22 into their conditioned media compared to levels produced by control cells. Our findings have clinical relevance because of the elevated pro-inflammatory milieu within the degenerating human disc.     

Global changes in soil stocks of carbon,nitrogen, phosphorus,and sulphur as influenced by long‐term agricultural production

Quantifying changes in stocks of C, N, P, and S in agricultural soils is important not only for managing these soils sustainably as required to feed a growing human population, but for C and N, they are also important for understanding fluxes of greenhouse gases from the soil environment. In a global meta‐analysis, 102 studies were examined to investigate changes in soil stocks of organic C, total N, total P, and total S associated with long‐term land‐use changes. Conversion of native vegetation to cropping resulted in substantial losses of C (?1.6 kg m^?2, ?43%), N (?0.15 kg m^?2, ?42%), P (?0.029 kg m^?2, ?27%), and S (?0.015 kg m^?2, ?33%). The subsequent conversion of conventional cropping systems to no‐till, organic agriculture, or organic amendment systems subsequently increased stocks, but the magnitude of this increase (average of +0.47 kg m^?2 for C and +0.051 kg m^?2 for N) was small relative to the initial decrease. We also examined the conversion of native vegetation to pasture, with changes in C (?11%), N (+4.1%), and P (+25%) generally being modest relative to changes caused by conversion to cropping. The C:N ratio remained relatively constant irrespective of changes in land use, whilst in contrast, the C:S ratio decreased by 21% in soils converted to cropping – this suggesting that biochemical mineralization is of importance for S. The data presented here will assist in the assessment of different agricultural production systems on soil stocks of C, N, P, and S – this information assisting not only in quantifying the effects of existing agricultural production on these stocks, but also allowing for informed decision‐making regarding the potential effects of future land‐use changes.     

Glutathione reductase functions as vanadate(V) reductase

The oxidation of NADPH by vanadate(V) in the presence of glutathione reductase showed typical enzymatic kinetics. The oxidation was inhibited by N-ethylmaleimide, a glutathione reductase inhibitor. Superoxide dismutase had no significant effect on the oxidation, indicating noninvolvement of the superoxide radical. The vanadate(V) reduction was found to be a one-electron transfer process. These results suggest a new pathway for vanadate(V) metabolism and a new function of glutathione reductase.     

The SecY complex forms a channel capable of ionic discrimination

Protein translocation across the bacterial membrane occurs at the SecY complex or channel. The resting SecY channel is impermeable to small molecules owing to a plug domain that creates a seal. Here, we report that a channel loosely sealed, or with a plug locked open, does not, however, lead to general membrane permeability. Instead, strong selectivity towards small monovalent anions, especially chloride, is observed. Mutations in the pore ring‐structure increase both the translocation activity of the channel and its ionic conductance, however the selectivity is maintained. The same ionic specificity also occurs at the onset of protein translocation and across the archaeal SecY complex. Thus, the ion‐conducting characteristic of the channel seems to be conserved as a normal consequence of protein translocation. We propose that the pore ring‐structure forms a selectivity filter, allowing cells to tolerate channels with imperfect plugs.     

Enzyme classification with peptide programs: a comparative study

Background  

Efficient and accurate prediction of protein function from sequence is one of the standing problems in Biology. The generalised use of sequence alignments for inferring function promotes the propagation of errors, and there are limits to its applicability. Several machine learning methods have been applied to predict protein function, but they lose much of the information encoded by protein sequences because they need to transform them to obtain data of fixed length.     

Unigene derived microsatellite markers for the cereal genomes

Unigene derived microsatellite (UGMS) markers have the advantage of assaying variation in the expressed component of the genome with unique identity and positions. We characterized the microsatellite motifs present in the unigenes of five cereal species namely, rice, wheat, maize, Sorghum and barley and compared with those in Arabidopsis. The overall UGMS frequency in the five cereal species was 1/7.6 kb. The maximum UGMS frequency was in rice (1/3.6 kb) and the lowest in wheat (1/10.6 kb). GC-rich trinucleotide repeat motifs coding for alanine followed by arginine and the dinucleotide repeat motif GA were found to be abundant UGMS classes across all the five cereal species. Primers could be designed for 95% (wheat and barley) to 97% (rice) of the identified microsatellites. The proportion and frequency of occurrence of long hypervariable class I (≥20 nucleotides) and potentially variable class II (12–20 nucleotides) UGMS across five cereal species were characterized. The class I UGMS markers were physically mapped in silico on to the finished rice genome and bin-mapped in wheat. Comparative mapping based on class I UGMS markers in rice and wheat revealed syntenic relationships between the two genomes. High degree of conservation and cross-transferability of the class I UGMS markers were evident among the five cereal species, which was validated experimentally. The class I UGMS-conserved orthologous set (COS) markers identified in this study would be useful for understanding the evolution of genes and genomes in cereals. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Thermodynamics and stability of the PAAD/DAPIN/PYRIN domain of IFI-16

The PAAD domain is a conserved domain recently identified in more than 35 human proteins that are involved in apoptosis and inflammatory signaling pathways. Structural studies have confirmed that this domain belongs to the death domain superfamily which includes PAAD/CARD/DED/DD families. Recently, the 3D structures determined by NMR of NALP1 and ASC PAAD domain, members of the PAAD family, have shown that it is composed of a 6 helix bundle as with other death domain family members. However, helix-3 in the solved structures is unordered in solution. In this study we compare the thermodynamic, folding and stability properties of different members of the PAAD and CARD families and investigate structural conformational changes induced by the helix inducers trifluoroethanol and SDS on the PAAD domain of IFI16 and on the CARD domain of RAIDD. We show that inside the PAAD and CARD families, members have similar thermodynamic properties, however, the DeltaG of folding for PAAD and CARD members are, respectively, -1.4 and -5.5 kcal mol(-1). This difference is attributed to less alpha helical content for PAAD due to the unfolding of helix-3 that lowers bonded energy and increases disorder when compared to CARD members. Despite identical fold between PAAD and CARD families but limited sequence identity, there are striking differences in the thermodynamics of both families.