Cutting edge: impaired MHC class I expression in mice deficient for Nlrc5/class I transactivator

MHC class I and class II are crucial for the adaptive immune system. Although regulation of MHC class II expression by CIITA has long been recognized, the mechanism of MHC class I transactivation has been largely unknown until the recent discovery of NLRC5/class I transactivator. In this study, we show using Nlrc5-deficient mice that NLRC5 is required for both constitutive and inducible MHC class I expression. Loss of Nlrc5 resulted in severe reduction in the expression of MHC class I and related genes such as β(2)-microglobulin, Tap1, or Lmp2, but did not affect MHC class II levels. IFN-γ stimulation could not overcome the impaired MHC class I expression in Nlrc5-deficient cells. Upon infection with Listeria monocyogenes, Nlrc5-deficient mice displayed impaired CD8(+) T cell activation, accompanied with increased bacterial loads. These findings illustrate critical roles of NLRC5/class I transactivator in MHC class I gene regulation and host defense by CD8(+) T cell responses.     

SMALL ACIDIC PROTEIN1 Acts with RUB Modification Components, the COP9 Signalosome, and AXR1 to Regulate Growth and Development of Arabidopsis

Previously, a dysfunction of the SMALL ACIDIC PROTEIN1 (SMAP1) gene was identified as the cause of the anti-auxin resistant1 (aar1) mutant of Arabidopsis (Arabidopsis thaliana). SMAP1 is involved in the response pathway of synthetic auxin, 2,4-dichlorophenoxyacetic acid, and functions upstream of the auxin/indole-3-acetic acid protein degradation step in auxin signaling. However, the exact mechanism by which SMAP1 functions in auxin signaling remains unknown. Here, we demonstrate that SMAP1 is required for normal plant growth and development and the root response to indole-3-acetic acid or methyl jasmonate in the auxin resistant1 (axr1) mutation background. Deletion analysis and green fluorescent protein/glutathione S-transferase pull-down assays showed that SMAP1 physically interacts with the CONSTITUTIVE PHOTOMORPHOGENIC9 SIGNALOSOME (CSN) via the SMAP1 F/D region. The extremely dwarf phenotype of the aar1-1 csn5a-1 double mutant confirms the functional role of SMAP1 in plant growth and development under limiting CSN functionality. Our findings suggest that SMAP1 is involved in the auxin response and possibly in other cullin-RING ubiquitin ligase-regulated signaling processes via its interaction with components associated with RELATED TO UBIQUITIN modification.     

CD16 regulates TRIF-dependent TLR4 response in human monocytes and their subsets

Blood monocytes recognize Gram-negative bacteria through the TLR4, which signal via MyD88- and TRIF-dependent pathway to trigger an immune-inflammatory response. However, a dysregulated inflammatory response by these cells often leads to severe pathologies such as sepsis. We investigated the role of CD16 in the regulation of human monocyte response to Gram-negative endotoxin and sepsis. Blood monocytes from sepsis patients demonstrated an upregulation of several TRIF-dependent genes as well as a selective expansion of CD16-expressing (CD16(+)) monocytes. Gene expression and biochemical studies revealed CD16 to regulate the TRIF-dependent TLR4 pathway in monocytes by activating Syk, IFN regulatory factor 3, and STAT1, which resulted in enhanced expression of IFNB, CCL5, and CXCL10. CD16 also upregulated the expression of IL-1R-associated kinase M and IL-1 receptor antagonist, which are negative regulators of the MyD88-dependent pathway. CD16 overexpression or small interfering RNA knockdown in monocytes confirmed the above findings. Interestingly, these results were mirrored in the CD16(+) monocyte subset isolated from sepsis patients, providing an in vivo confirmation to our findings. Collectively, the results from the current study demonstrate CD16 as a key regulator of the TRIF-dependent TLR4 pathway in human monocytes and their CD16-expressing subset, with implications in sepsis.     

Temporal changes in species diversity and composition in abandoned fields in a <Emphasis Type="Italic">trans</Emphasis>-Himalayan landscape,Nepal

Secondary succession is an increasing phenomenon due to global changes in agriculture policies and practices. The empirical findings are biased towards the temperate zone. Abandonment of agriculture fields is less frequent in the subtropical and tropical zones where agriculture areas are, in general, expanding. But there are exceptions; a rapid rate of abandonment of agricultural fields have taken place in the arid trans-Himalayan region, due to today’s globalization of economy. We analysed agriculture fields that were abandoned between 1950 and 2003 in a large u-valley in central Nepal (3400 m a.s.l.). The potential forest vegetation is dominated by Pinus wallichina and shrubs of junipers and cotoneaster species. We tested the intermediate richness hypothesis in relation to vegetation cover, soil development and whether old-field succession is convergent or divergent with species data from 242 1 m² plots in 5 age-classes. The main species compositional turnover expressed by Detrended Correspondence Analyses (DCA) correlated, as expected, with time after abandonment. Fields that were abandoned a long time ago are closer to forest at the periphery of the agricultural landscape. Moisture of the soil significantly increased with age of abandonment, but total vegetation cover and pH were negatively related to age. Beta diversity expressed in DCA SD-units showed an increasing trend with age of abandonment, supporting the divergence pattern in old-field succession. The reason why the succession is not converging may be due to browsing by domestic animals that prevent a closed canopy of pines and juniper to develop. There was a significant hump-shaped pattern in species richness along the temporal gradient, which agrees with the intermediate species-richness hypothesis. There was a rapid increase in species richness in plots close to the villages that were used for haymaking which increased the seed input significantly.     

Characterization of AtlL, a bifunctional autolysin of Staphylococcus lugdunensis with N-acetylglucosaminidase and N-acetylmuramoyl-l-alanine amidase activities

The nucleotide sequence of atlL , a gene encoding a putative Staphylococcus lugdunensis peptidoglycan hydrolase, was determined using degenerate consensus PCR and genome walking. This 3837-bp gene encodes a protein, AtlL, that appears as a putative bifunctional autolysin with a 29-amino acid putative signal peptide and two enzymatic putative centres ( N -acetylmuramoyl- l -alanine amidase and N -acetylglucosaminidase) interconnected with three imperfect repeated sequences displaying glycine–tryptophan motifs. In order to determine whether both lytic domains were functional, and verify their exact enzymatic activities, gene fragments harbouring both putative domains, AM ( N -acetylmuramoyl- l -alanine amidase enzymatic centre plus two repeated sequences) and GL ( N -acetylglucosaminidase enzymatic centre plus one repeated sequence), were isolated, subcloned, and expressed in Escherichia coli . Purified recombinant AM and GL protein truncations exhibited cell wall lytic activity in zymograms performed with cell walls of Micrococcus lysodeikticus, Bacillus subtilis , and S. lugdunensis. AtlL is expressed during the whole growth, with an overexpression in the early-exponential stage. Liquid chromatography-mass spectrometry analysis of muropeptides generated by digestion of B. subtilis cell walls demonstrated the hydrolytic bond specificities and confirmed both of the acetyl domains' activities as predicted by sequence homology data. AtlL is the first autolysin described in S. lugdunensis , with a bifunctional enzymatic activity involved in peptidoglycan hydrolysis.     

Exploiting BAC-end sequences for the mining,characterization and utility of new short sequences repeat (SSR) markers in Citrus

The aim of this study was to develop a large set of microsatellite markers based on publicly available BAC-end sequences (BESs), and to evaluate their transferability, discriminating capacity of genotypes and mapping ability in Citrus. A set of 1,281 simple sequence repeat (SSR) markers were developed from the 46,339 Citrus clementina BAC-end sequences (BES), of them 20.67% contained SSR longer than 20 bp, corresponding to roughly one perfect SSR per 2.04 kb. The most abundant motifs were di-nucleotide (16.82%) repeats. Among all repeat motifs (TA/AT)n is the most abundant (8.38%), followed by (AG/CT)n (4.51%). Most of the BES-SSR are located in the non-coding region, but 1.3% of BES-SSRs were found to be associated with transposable element (TE). A total of 400 novel SSR primer pairs were synthesized and their transferability and polymorphism tested on a set of 16 Citrus and Citrus relative’s species. Among these 333 (83.25%) were successfully amplified and 260 (65.00%) showed cross-species transferability with Poncirus trifoliata and Fortunella sp. These cross-species transferable markers could be useful for cultivar identification, for genomic study of Citrus, Poncirus and Fortunella sp. Utility of the developed SSR marker was demonstrated by identifying a set of 118 markers each for construction of linkage map of Citrus reticulata and Poncirus trifoliata. Genetic diversity and phylogenetic relationship among 40 Citrus and its related species were conducted with the aid of 25 randomly selected SSR primer pairs and results revealed that citrus genomic SSRs are superior to genic SSR for genetic diversity and germplasm characterization of Citrus spp.     

Therapeutic applications of lysostaphin against Staphylococcus aureus

Staphylococcus aureus, an opportunistic pathogen, causes diverse community and nosocomial-acquired human infections, including folliculitis, impetigo, sepsis, septic arthritis, endocarditis, osteomyelitis, implant-associated biofilm infections and contagious mastitis in cattle. In recent days, both methicillin-sensitive and methicillin-resistant S. aureus infections have increased. Highly effective anti-staphylococcal agents are urgently required. Lysostaphin is a 27 kDa zinc metallo antimicrobial lytic enzyme that is produced by Staphylococcus simulans biovar staphylolyticus and was first discovered in the 1960s. Lysostaphin is highly active against S. aureus strains irrespective of their drug-resistant patterns with a minimum inhibitory concentration of ranges between 0·001 and 0·064 μg ml⁻¹. Lysostaphin has activity against both dividing and non-dividing S. aureus cells; and can seep through the extracellular matrix to kill the biofilm embedded S. aureus. In spite of having excellent anti-staphylococcal activity, its clinical application is hindered because of its immunogenicity and reduced bio-availability. Extensive research with lysostaphin lead to the development of several engineered lysostaphin derivatives with reduced immunogenicity and increased serum half-life. Therapeutic efficacy of both native and engineered lysostaphin derivatives was studied by several research groups. This review provides an overview of the therapeutic applications of native and engineered lysostaphin derivatives developed to eradicate S. aureus infections.     

Dietary Mannan-oligosaccharides potentiate the beneficial effects of Bifidobacterium bifidum in broiler chicken

This study investigated the effects of dietary Bifidobacterium bifidum (BFD) and mannan-oligosaccharide (MOS), as a synbiotic, on the production performance, gut microbiology, serum biochemistry, antioxidant profile and health indices of broiler chicken. Six dietary treatments were T1 (negative control), T2 (positive control-20 mg antibiotic BMD kg⁻¹ diet; BMD: bacitracin methylene disalicylate), T3 (0·1% MOS + 10⁶ CFU BFD per g feed), T4 (0·1% MOS + 10⁷ CFU BFD per g feed), T5 (0·2% MOS + 10⁶ CFU BFD per g feed) and T6 (0·2% MOS + 10⁷ CFU BFD per g feed). Significantly (P < 0·01) better growth performance and efficiency was observed in birds supplemented with 0·2% MOS along with 10⁶ CFU BFD per g of feed compared to BMD and control birds. Supplementation with 0·2% MOS along with either 10⁶ or 10⁷ CFU BFD per g feed reduced (P < 0·01) the gut coliform, Escherichia coli, total plate count, and Clostridium perfringens count and increased the Lactobacillus and Bifidobacterium count. Significantly (P < 0·01) higher serum and liver antioxidant enzyme pool, serum HDL cholesterol and lower serum glucose, triglyceride, total cholesterol, cardiac risk ratio, atherogenic coefficient and atherogenic index of plasma were observed in birds supplemented with 0·2% MOS along with 10⁶ CFU BFD per g of feed compared to control or BMD supplemented birds. Better production performance, gut microbial composition, serum biochemistry, antioxidant profile and health indices were depicted by broiler chicken supplemented with 0·2% MOS and 10⁶ CFU BFD per g of feed.