The root-rhizosphere interface of Populus is the nexus of a variety of associations between bacteria, fungi, and the host plant and an ideal model for studying interactions between plants and microorganisms. However, such studies have generally been confined to greenhouse and plantation systems. Here we analyze microbial communities from the root endophytic and rhizospheric habitats of Populus deltoides in mature natural trees from both upland and bottomland sites in central Tennessee. Community profiling utilized 454 pyrosequencing with separate primers targeting the V4 region for bacterial 16S rRNA and the D1/D2 region for fungal 28S rRNA genes. Rhizosphere bacteria were dominated by Acidobacteria (31%) and Alphaproteobacteria (30%), whereas most endophytes were from the Gammaproteobacteria (54%) as well as Alphaproteobacteria (23%). A single Pseudomonas-like operational taxonomic unit (OTU) accounted for 34% of endophytic bacterial sequences. Endophytic bacterial richness was also highly variable and 10-fold lower than in rhizosphere samples originating from the same roots. Fungal rhizosphere and endophyte samples had approximately equal amounts of the Pezizomycotina (40%), while the Agaricomycotina were more abundant in the rhizosphere (34%) than endosphere (17%). Both fungal and bacterial rhizosphere samples were highly clustered compared to the more variable endophyte samples in a UniFrac principal coordinates analysis, regardless of upland or bottomland site origin. Hierarchical clustering of OTU relative abundance patterns also showed that the most abundant bacterial and fungal OTUs tended to be dominant in either the endophyte or rhizosphere samples but not both.Together, these findings demonstrate that root endophytic communities are distinct assemblages rather than opportunistic subsets of the rhizosphere.Populus is considered the model organism for the study of woody perennials (46) and represents the first tree genome to be fully sequenced (47). Populus has also received attention in bioenergy research for the production of cellulose-derived biofuels (47). Populus can be grown on land not suitable for food production and increase carbon sequestration, thus minimizing the competition between food and fuel production and reducing the carbon debt incurred through land use changes (39). Populus may also provide an ideal model for understanding a variety of plant-microbe interactions (45). Populus and other members of the Salicaceae are capable of establishing associations with both arbuscular mycorrhizal (AM) and ectomycorrhizal (ECM) fungi (14) that may result in unique interactions between these fungi, as well as other endophytic and rhizospheric organisms and the host. Populus bacterial rhizosphere and endophytic constituents have received some attention due to their potential role in phytoremediation of industrial chemicals (33) and heavy metals (4), as well as plant growth-promoting bacteria (PGPB), which benefit plants by providing fixed nitrogen and/or aiding resistance to infection by pathogens ...

We show that the mouse macrophage-restricted F4/80 protein is not required for the development and distribution of tissue macrophages but is involved in the generation of antigen-specific efferent regulatory T (T reg) cells that suppress antigen-specific immunity. In the in vivo anterior chamber (a.c.)–associated immune deviation (ACAID) model of peripheral tolerance, a.c. inoculation of antigen into F4/80−/− mice was unable to induce efferent T reg cells and suppress delayed-type hypersensitivity (DTH) responses. Moreover, the use of anti-F4/80 mAb and F4/80−/− APCs in an in vitro ACAID model showed that all APC cells in the culture must be able to express F4/80 protein if efferent T reg cells were to be generated. In a low-dose oral tolerance model, WT but not F4/80−/− mice generated an efferent CD8+ T reg cell population that suppressed an antigen-specific DTH response. Peripheral tolerance was restored in F4/80−/− mice by adoptive transfer of F4/80+ APCs in both peripheral tolerance models, indicating a central role for the F4/80 molecule in the generation of efferent CD8+ T reg cells.

The mammalian Nell1 gene encodes a protein kinase C-beta1 (PKC-beta1) binding protein that belongs to a new class of cell-signaling molecules controlling cell growth and differentiation. Over-expression of Nell1 in the developing cranial sutures in both human and mouse induces craniosynostosis, the premature fusion of the growing cranial bone fronts. Here, we report the generation, positional cloning and characterization of Nell1(6R), a recessive, neonatal-lethal point mutation in the mouse Nell1 gene, induced by N-ethyl-N-nitrosourea. Nell1(6R) has a T-->A base change that converts a codon for cysteine into a premature stop codon [Cys(502)Ter], resulting in severe truncation of the predicted protein product and marked reduction in steady-state levels of the transcript. In addition to the expected alteration of cranial morphology, Nell1(6R) mutants manifest skeletal defects in the vertebral column and ribcage, revealing a hitherto undefined role for Nell1 in signal transduction in endochondral ossification. Real-time quantitative reverse transcription-PCR assays of 219 genes showed an association between the loss of Nell1 function and reduced expression of genes for extracellular matrix (ECM) proteins critical for chondrogenesis and osteogenesis. Several affected genes are involved in the human cartilage disorder Ehlers-Danlos Syndrome and other disorders associated with spinal curvature anomalies. Nell1(6R) mutant mice are a new tool for elucidating basic mechanisms in osteoblast and chrondrocyte differentiation in the developing skull and vertebral column and understanding how perturbations in the production of ECM proteins can lead to anomalies in these structures.

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Background: Analysis of an allelic series of point mutations in a gene, generated by N-ethyl-N-nitrosourea (ENU) mutagenesis, is a valuable method for discovering the full scope of its biological function. Here we present an efficient gene-driven approach for identifying ENU-induced point mutations in any gene in C57BL/6J mice. The advantage of such an approach is that it allows one to select any gene of interest in the mouse genome and to go directly from DNA sequence to mutant mice.