Multiple TPR motifs characterize the Fanconi anemia FANCG protein

The genome protection pathway that is defective in patients with Fanconi anemia (FA) is controlled by at least eight genes, including BRCA2. A key step in the pathway involves the monoubiquitylation of FANCD2, which critically depends on a multi-subunit nuclear 'core complex' of at least six FANC proteins (FANCA, -C, -E, -F, -G, and -L). Except for FANCL, which has WD40 repeats and a RING finger domain, no significant domain structure has so far been recognized in any of the core complex proteins. By using a homology search strategy comparing the human FANCG protein sequence with its ortholog sequences in Oryzias latipes (Japanese rice fish) and Danio rerio (zebrafish) we identified at least seven tetratricopeptide repeat motifs (TPRs) covering a major part of this protein. TPRs are degenerate 34-amino acid repeat motifs which function as scaffolds mediating protein-protein interactions, often found in multiprotein complexes. In four out of five TPR motifs tested (TPR1, -2, -5, and -6), targeted missense mutagenesis disrupting the motifs at the critical position 8 of each TPR caused complete or partial loss of FANCG function. Loss of function was evident from failure of the mutant proteins to complement the cellular FA phenotype in FA-G lymphoblasts, which was correlated with loss of binding to FANCA. Although the TPR4 mutant fully complemented the cells, it showed a reduced interaction with FANCA, suggesting that this TPR may also be of functional importance. The recognition of FANCG as a typical TPR protein predicts this protein to play a key role in the assembly and/or stabilization of the nuclear FA protein core complex.     

EEG Biofeedback Treatment Improves Certain Attention and Somatic Symptoms in Fibromyalgia: A Pilot Study

Fibromyalgia (FMS) is a chronic, painful disorder often associated with measurable deficiencies in attention. Since EEG biofeedback (EEG-BF) has been used successfully to treat attention problems, we reasoned that this modality might be helpful in the treatment of attention problems in FMS. We also speculated that improvement in central nervous system (CNS) function might be accompanied by improvement in FMS somatic symptoms. We studied fifteen FMS patients with attention problems, demonstrated by visual and auditory continuous performance testing (CPT), while completing 40 or more EEG-BF sessions. Training consisted of a “SMR protocol” that augmented 12–15 Hz brainwaves (sensory motor rhythm; SMR), while simultaneously inhibiting 4–7 Hz brainwaves (theta) and 22–30 Hz brainwaves (high beta). Serial measurements of pain, fatigue, psychological distress, morning stiffness, and tenderness were also obtained. Sixty-three FMS patients who received standard medical care, but who did not receive EEG-BF, served as controls. Visual, but not auditory, attention improved significantly (P < 0.008). EEG-BF treated subjects also showed improvement in tenderness, pain and fatigue. Somatic symptoms did not change significantly in controls. Visual attention parameters and certain somatic features of FMS appear to improve with an EEG-BF SMR protocol. EEG-BF training in FMS deserves further study.     

Fire tolerance of a resprouting <Emphasis Type="Italic">Artemisia</Emphasis> (Asteraceae) shrub

In North America, most Artemisia (Asteraceae) shrub species lack the ability to resprout after disturbances that remove aboveground biomass. We studied the response of one of the few resprouting Artemisia shrubs, Artemisia filifolia (sand sagebrush), to the effects of prescribed fires. We collected data on A. filifolia density and structural characteristics (height, canopy area, and canopy volume) in an A. filifolia shrubland in the southern Great Plains of North America. Our study sites included areas that had not been treated with prescribed fire, areas that had been treated with only one prescribed fire within the previous 5 years, and areas that had been treated with two prescribed fires within the previous 10 years. Our data were collected at time periods ranging from ½ to 5 years after the prescribed fires. Density of A. filifolia was not affected by one or two fires. Structural characteristics, although initially altered by prescribed fire, recovered to levels characteristic of unburned areas in 3–4 years after those fires. In contrast to most non-sprouting North American Artemisia shrub species, our research suggested that the resprouting A. filifolia is highly tolerant to the effects of fire.     

Higher number of <Emphasis Type="Italic">Helicobacter pylori</Emphasis> CagA EPIYA C phosphorylation sites increases the risk of gastric cancer,but not duodenal ulcer

Background  

Helicobacter pylori infection is one of the most common infections worldwide and is associated with gastric cancer and peptic ulcer. Bacterial virulence factors such as CagA have been shown to increase the risk of both diseases. Studies have suggested a causal role for CagA EPIYA polymorphisms in gastric carcinogenesis, and it has been shown to be geographically diverse. We studied associations between H. pylori CagA EPIYA patterns and gastric cancer and duodenal ulcer, in an ethnically admixed Western population from Brazil. CagA EPIYA was determined by PCR and confirmed by sequencing. A total of 436 patients were included, being 188 with gastric cancer, 112 with duodenal ulcer and 136 with gastritis.     

Macromolecular composition of phloem exudate from white lupin (Lupinus albus L.)

Background  

Members of the legume genus Lupinus exude phloem 'spontaneously' from incisions made to the vasculature. This feature was exploited to document macromolecules present in exudate of white lupin (Lupinus albus [L.] cv Kiev mutant), in particular to identify proteins and RNA molecules, including microRNA (miRNA).     

Low CCR7-mediated migration of human monocyte derived dendritic cells in response to human respiratory syncytial virus and human metapneumovirus

Human respiratory syncytial virus (HRSV) and, to a lesser extent, human metapneumovirus (HMPV) and human parainfluenza virus type 3 (HPIV3), can re-infect symptomatically throughout life without significant antigenic change, suggestive of incomplete or short-lived immunity. In contrast, re-infection by influenza A virus (IAV) largely depends on antigenic change, suggestive of more complete immunity. Antigen presentation by dendritic cells (DC) is critical in initiating the adaptive immune response. Antigen uptake by DC induces maturational changes that include decreased expression of the chemokine receptors CCR1, CCR2, and CCR5 that maintain DC residence in peripheral tissues, and increased expression of CCR7 that mediates the migration of antigen-bearing DC to lymphatic tissue. We stimulated human monocyte-derived DC (MDDC) with virus and found that, in contrast to HPIV3 and IAV, HMPV and HRSV did not efficiently decrease CCR1, 2, and 5 expression, and did not efficiently increase CCR7 expression. Consistent with the differences in CCR7 mRNA and protein expression, MDDC stimulated with HRSV or HMPV migrated less efficiently to the CCR7 ligand CCL19 than did IAV-stimulated MDDC. Using GFP-expressing recombinant virus, we showed that the subpopulation of MDDC that was robustly infected with HRSV was particularly inefficient in chemokine receptor modulation. HMPV- or HRSV-stimulated MDDC responded to secondary stimulation with bacterial lipopolysaccharide or with a cocktail of proinflammatory cytokines by increasing CCR7 and decreasing CCR1, 2 and 5 expression, and by more efficient migration to CCL19, suggesting that HMPV and HRSV suboptimally stimulate rather than irreversibly inhibit MDDC migration. This also suggests that the low concentration of proinflammatory cytokines released from HRSV- and HMPV-stimulated MDDC is partly responsible for the low CCR7-mediated migration. We propose that inefficient migration of HRSV- and HMPV-stimulated DC to lymphatic tissue contributes to reduced adaptive responses to these viruses.     

Optimization of cell lysis and protein digestion protocols for the analysis of HeLa S3 cells by LC-MS/MS

In order to maximize the number of proteins identified from Hela S3 cell lysate we tested various cell lysis, protein precipitation and digestion protocols. First, we compared three different lysis buffers, two mechanical cell disruption methods and two precipitation methods. Then, we tested six different in-solution digestion protocols, three different in-gel digestion protocols and ten different peptide extraction protocols. The result is a proposal for an optimized protocol to prepare the whole cell lysate samples from HeLa S3 cells.     

Cannabidiol reduces intestinal inflammation through the control of neuroimmune axis

Enteric glial cells (EGC) actively mediate acute and chronic inflammation in the gut; EGC proliferate and release neurotrophins, growth factors, and pro-inflammatory cytokines which, in turn, may amplify the immune response, representing a very important link between the nervous and immune systems in the intestine. Cannabidiol (CBD) is an interesting compound because of its ability to control reactive gliosis in the CNS, without any unwanted psychotropic effects. Therefore the rationale of our study was to investigate the effect of CBD on intestinal biopsies from patients with ulcerative colitis (UC) and from intestinal segments of mice with LPS-induced intestinal inflammation. CBD markedly counteracted reactive enteric gliosis in LPS-mice trough the massive reduction of astroglial signalling neurotrophin S100B. Histological, biochemical and immunohistochemical data demonstrated that S100B decrease was associated with a considerable decrease in mast cell and macrophages in the intestine of LPS-treated mice after CBD treatment. Moreover the treatment of LPS-mice with CBD reduced TNF-α expression and the presence of cleaved caspase-3. Similar results were obtained in ex vivo cultured human derived colonic biopsies. In biopsies of UC patients, both during active inflammation and in remission stimulated with LPS+INF-γ, an increased glial cell activation and intestinal damage were evidenced. CBD reduced the expression of S100B and iNOS proteins in the human biopsies confirming its well documented effect in septic mice. The activity of CBD is, at least partly, mediated via the selective PPAR-gamma receptor pathway. CBD targets enteric reactive gliosis, counteracts the inflammatory environment induced by LPS in mice and in human colonic cultures derived from UC patients. These actions lead to a reduction of intestinal damage mediated by PPARgamma receptor pathway. Our results therefore indicate that CBD indeed unravels a new therapeutic strategy to treat inflammatory bowel diseases.