Ionizing radiation, but not stimulation with epidermal growth factor (EGF), triggers EGF receptor (EGFR) import into the nucleus in a probably karyopherin ␣-linked manner. An increase in nuclear EGFR is also observed after treatment with H 2 O 2 , heat, or cisplatin. During this process, the proteins Ku70/80 and the protein phosphatase 1 are transported into the nucleus. As a consequence, an increase in the nuclear kinase activity of DNA-dependent kinase (DNA-PK) and increased formation of the DNA end-binding protein complexes containing DNA-PK, essential for repair of DNA-strand breaks, occurred. Blockade of EGFR import by the anti-EGFR monoclonal antibody C225 abolished EGFR import into the nucleus and radiation-induced activation of DNA-PK, inhibited DNA repair, and increased radiosensitivity of treated cells. Our data implicate a novel function of the EGFR during DNA repair processes.The epidermal growth factor receptor (EGFR) 1 is essential for mediation of both proliferative and survival signals to cells (1). At least five mitogenic growth factors bind to and activate EGFR. In addition to EGF these factors include transforming growth factor-␣, amphiregulin, heparin binding EGF (2), and epiregulin (3). Moreover, in recent years it became apparent that in addition to ligand binding-induced activation of the EGFR, ligand-independent receptor activating processes also exist (4). Activation of the EGFR signaling pathway by ligands has been linked with increased cell proliferation, angiogenesis, and metastasis and decreased apoptosis (5). As a consequence it is assumed that increased EGFR signaling plays an important role in tumorigenesis (6, 7). The mechanisms of ligand-independent EGFR activation and also the importance for the cell fate are not understood so far. But it is noteworthy that the ligand-independent activation always is linked to exposure to genotoxic stress (4,8,9). However, the differences of the observed cell response after ligand-induced EGFR activation, e.g. cell proliferation, and ligandindependent activation, e.g. cell cycle arrest, argue rather for a link to DNA repair processes for ligand-independent activation. This idea was strengthened by the observation that many tumor cells show an increased radiosensitivity after inhibition of EGFR signaling (10). The most detrimental DNA damages after treatment with ionizing radiation are double-strand strands (11), which are preferentially repaired in mammalians by nonhomologous end-joining (12). Looking for a connection between radiation-induced EGFR activation and DNA repair, we found a report stating that there is a physical interaction of EGFR and DNAdependent kinase (DNA-PK) (13) after cell treatment with the EGFR blocking antibody C225. As a consequence, the nuclear DNA-PK protein and activity was reduced, providing an explanation for the radiosensitizing effect of the EGFR blockage. However, this paper did not answer the question of the functional role of the physical interaction between EGFR and DNA-PK. Therefore, we addressed herein the...

BackgroundEarly microbial colonization of the gut reduces the incidence of infectious, inflammatory and autoimmune diseases. Recent population studies reveal that childhood hygiene is a significant risk factor for development of inflammatory bowel disease, thereby reinforcing the hygiene hypothesis and the potential importance of microbial colonization during early life. The extent to which early-life environment impacts on microbial diversity of the adult gut and subsequent immune processes has not been comprehensively investigated thus far. We addressed this important question using the pig as a model to evaluate the impact of early-life environment on microbe/host gut interactions during development.ResultsGenetically-related piglets were housed in either indoor or outdoor environments or in experimental isolators. Analysis of over 3,000 16S rRNA sequences revealed major differences in mucosa-adherent microbial diversity in the ileum of adult pigs attributable to differences in early-life environment. Pigs housed in a natural outdoor environment showed a dominance of Firmicutes, in particular Lactobacillus, whereas animals housed in a hygienic indoor environment had reduced Lactobacillus and higher numbers of potentially pathogenic phylotypes. Our analysis revealed a strong negative correlation between the abundance of Firmicutes and pathogenic bacterial populations in the gut. These differences were exaggerated in animals housed in experimental isolators. Affymetrix microarray technology and Real-time Polymerase Chain Reaction revealed significant gut-specific gene responses also related to early-life environment. Significantly, indoor-housed pigs displayed increased expression of Type 1 interferon genes, Major Histocompatibility Complex class I and several chemokines. Gene Ontology and pathway analysis further confirmed these results.ConclusionEarly-life environment significantly affects both microbial composition of the adult gut and mucosal innate immune function. We observed that a microbiota dominated by lactobacilli may function to maintain mucosal immune homeostasis and limit pathogen colonization.

Background: To elucidate the role of src kinase in caveolin-1 driven internalization and nuclear transport of EGFR linked to regulation of DNA-repair in irradiated cells.

"Roseburia inulinivorans" is an anaerobic polysaccharide-utilizing firmicute bacterium from the human colon that was identified as a producer of butyric acid during growth on glucose, starch, or inulin. R. inulinivorans A2-194 is also able to grow on the host-derived sugar fucose, following a lag period, producing propionate and propanol as additional fermentation products. A shotgun genomic microarray was constructed and used to investigate the switch in gene expression that is involved in changing from glucose to fucose utilization. This revealed a set of genes coding for fucose utilization, propanediol utilization, and the formation of propionate and propanol that are up-regulated during growth on fucose. These include homologues of genes that are implicated in polyhedral body formation in Salmonella enterica. Dehydration of the intermediate 1,2-propanediol involves an enzyme belonging to the new B 12 -independent glycerol dehydratase family, in contrast to S. enterica, which relies on a B 12 -dependent enzyme. A typical gram-positive agr-type quorum-sensing system was also up-regulated in R. inulinivorans during growth on fucose. Despite the lack of genome sequence information for this commensal bacterium, microarray analysis has provided a powerful tool for obtaining new information on its metabolic capabilities.The human colon contains a dense and highly diverse microbial community consisting of over 500 different bacterial species (13,15,20,41). These bacteria are predominantly obligate anaerobes and produce fermentation products that may be beneficial (e.g., butyrate) (35) or detrimental (e.g., hydrogen sulfide) (34) to the epithelial cells lining the human colon. The main butyrate-producing colonic anaerobes belong to Clostridium clusters IV and XIVa (3,20,26) and include cluster XIVa bacteria that have been assigned to the genus Roseburia (11). Studies using fluorescent in situ hybridization have shown that bacteria related to Roseburia (including Eubacterium rectale) can comprise up to 10% of the total bacterial population in human feces (21, 43). All Roseburia spp. produce butyrate, but they differ markedly in their substrate utilization profiles (12,36).Colonic bacteria gain energy from dietary substrates that escape digestion in the upper gastrointestinal tract, including plant cell wall polysaccharides, resistant starch, inulin, and a variety of oligosaccharides. In the distal colon, host-derived secretions and mucin, including the glycoproteins and glycolipids covering the surface of gut epithelial cells, may be significant substrates. L-fucose is frequently present at the terminus of epithelial glycoconjugates (14), and several pathogenic bacteria, including Salmonella enterica serovar Typhimurium LT2 (subsequently referred to as Salmonella serovar Typhimurium LT2) and the commensal bacterium Bacteroides thetaiotaomicron, utilize fucose for growth (see references 1 and 38, respectively). B. fragilis is able to incorporate fucose into its own surface polysaccharides, a mechanism that gives it a c...