Lysophosphatidic acid,human osteoblast formation,maturation and the role of 1α,25-Dihydroxyvitamin D3 (calcitriol)

The simplest signalling lipid Lysophosphatidic acid (LPA) elicits pleiotropic actions upon most mammalian cell types. Although LPA has an established role in many biological processes, particularly wound healing and cancer, the function of LPA for human osteoblast (hOB) biology is still unravelling. Early studies, identified in this review, gave a reliable indication that LPA, via binding to one of several transmembrane receptors, stimulated multiple intracellular signalling networks coupled to changes in cell growth, fibronectin binding, maturation and survival. The majority of studies exploring the actions of LPA on hOB responses have done so using the lipid in isolation. Our own research has focussed on the co-operation of LPA with the active vitamin D3 metabolite, 1α25,dihydroxycholecalciferol (calcitriol), in light of a serendipitous discovery that calcitriol, in a serum-free culture setting, was unable to promote hOB maturation. We subsequently learnt that the serum-borne factor co-operating with calcitriol to enhance hOB differentiation was LPA bound to the albumin fraction of whole serum. Recent studies from our laboratory have identified that LPA and calcitriol are a potent pairing for securing hOB formation from their stem cell progeny. Greater understanding of the ability of LPA to influence, for example, hOB growth, maturation and survival could be advantageous in developing novel strategies aimed at improving skeletal tissue repair and regeneration. Herein this review provides an insight into the diversity of studies exploring the actions of a small lipid on a major cell type key to bone tissue health and homeostasis. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.     

The desire for variety: Italian wall lizard (Podarcis siculus) populations introduced to the United States via the pet trade are derived from multiple native-range sources

Tests of invasion success often require comparisons between introduced and native populations, but determining the native-range sources for introduced populations can be difficult. Molecular markers can help clarify the geographic extent of native-range sources, helping to identify which populations are appropriate for comparative studies. The Italian Wall Lizard (Podarcis siculus) was introduced multiple times to the United States with extant populations in California, Kansas, New Jersey, and New York. We used phylogeographic analysis of mtDNA sequences (cytb gene) for individuals sampled from these introduced populations and across the native range to identify the number of independent introductions and the location of the source populations. Haplotypes sampled from introduced populations were nested within three geographically distinct, well-supported clades that together encompassed a large portion of the native range. Combining these phylogeographic results with documentation of the introductions revealed putative sources: California individuals are derived from Sicily; Kansas and New York populations are from Tuscany near Florence; and the New Jersey population is likely from the Adriatic coastal region, but a more specific locality is not possible. The pet trade dominates the invasion pathway for P. siculus introductions to the US. The genetically and geographically diverse sampling of its native range may be driven by the desire for phenotypic variety in the pet trade, a hypothesis that needs future testing.     

Temporal dynamics of plant community regeneration sources during tallgrass prairie restoration

Ecological restoration aims to augment and steer the composition and contribution of propagules for community regeneration in degraded environments. We quantified patterns in the abundance, richness, and diversity of seed and bud banks across an 11-year chronosequence of restored prairies and in prairie remnants to elucidate the degree to which the germinable seed bank, emerged seedlings, belowground buds, and emerged ramets were related to community regeneration. There were no directional patterns in the abundance, richness, or diversity of the germinable seed bank across the chronosequence. Emerged seedling abundance of sown species decreased during restoration. Richness and diversity of all emerged seedlings and non-sown emerged seedling species decreased across the chronosequence. Conversely, abundance and richness of belowground buds increased with restoration age and belowground bud diversity of sown species increased across the chronosequence. Numbers of emerged ramets also increased across the chronosequence and was driven primarily by the number of graminoid ramets. There were no temporal changes in abundance and richness of sown and non-sown emerged ramets, but diversity of sown emerged ramets increased across the chronosequence. This study demonstrates that after initial seeding, plant community structure in restored prairies increasingly reflects the composition of the bud bank.     

Characterization of the Late Endosomal ESCRT Machinery in Trypanosoma brucei

The multivesicular body (MVB) is a specialized Rab7+ late endosome (LE) containing multiple intralumenal vesicles that function in targeting ubiquitinylated cell surface proteins to the lysosome for degradation. African trypanosomes lack a morphologically well‐defined MVB, but contain orthologs of the ESCRT (Endosomal Sorting Complex Required for Transport) machinery that mediates MVB formation. We investigate the role of TbVps23, an early ESCRT component, and TbVps4, the terminal ESCRT ATPase, in lysosomal trafficking in bloodstream form trypanosomes. Both localize to the TbRab7+ LE and RNAi silencing of each rapidly blocks growth. TbVps4 silencing results in approximately threefold accumulation of TbVps23 at the LE, consistent with blocking terminal ESCRT disassembly. Trafficking of endocytic and biosynthetic cargo, but not default lysosomal reporters, is also negatively affected. Others reported that TbVps23 mediates ubiquitin‐dependent lysosomal degradation of invariant surface glycoproteins (ISG65) (Leung et al., Traffic 2008;9:1698–1716). In contrast, we find that TbVps23 ablation does not affect ISG65 turnover, while TbVps4 silencing markedly enhances lysosomal degradation. We propose several models to accommodate these results, including that the ESCRT machinery actually retrieves ISG65 from the LE to earlier endocytic compartments, and in its absence ISG65 traffics more efficiently to the lysosome. Overall, these results confirm that the ESCRT machinery is essential in Trypanosoma brucei and plays important and novel role(s) in LE function in trypanosomes .     

Bacterial pathogen phytosensing in transgenic tobacco and Arabidopsis plants

Plants are subject to attack by a wide range of phytopathogens. Current pathogen detection methods and technologies are largely constrained to those occurring post‐symptomatically. Recent efforts were made to generate plant sentinels (phytosensors) that can be used for sensing and reporting pathogen contamination in crops. Engineered phytosensors indicating the presence of plant pathogens as early‐warning sentinels potentially have tremendous utility as wide‐area detectors. We previously showed that synthetic promoters containing pathogen and/or defence signalling inducible cis‐acting regulatory elements (RE) fused to a fluorescent protein (FP) reporter could detect phytopathogenic bacteria in a transient phytosensing system. Here, we further advanced this phytosensing system by developing stable transgenic tobacco and Arabidopsis plants containing candidate constructs. The inducibility of each synthetic promoter was examined in response to biotic (bacterial pathogens) or chemical (plant signal molecules salicylic acid, ethylene and methyl jasmonate) treatments using stably transgenic plants. The treated plants were visualized using epifluorescence microscopy and quantified using spectrofluorometry for FP synthesis upon induction. Time‐course analyses of FP synthesis showed that both transgenic tobacco and Arabidopsis plants were capable to respond in predictable ways to pathogen and chemical treatments. These results provide insights into the potential applications of transgenic plants as phytosensors and the implementation of emerging technologies for monitoring plant disease outbreaks in agricultural fields.     

Comprehensive genotyping of the USA national maize inbred seed bank

Background

Genotyping by sequencing, a new low-cost, high-throughput sequencing technology was used to genotype 2,815 maize inbred accessions, preserved mostly at the National Plant Germplasm System in the USA. The collection includes inbred lines from breeding programs all over the world.

Results

The method produced 681,257 single-nucleotide polymorphism (SNP) markers distributed across the entire genome, with the ability to detect rare alleles at high confidence levels. More than half of the SNPs in the collection are rare. Although most rare alleles have been incorporated into public temperate breeding programs, only a modest amount of the available diversity is present in the commercial germplasm. Analysis of genetic distances shows population stratification, including a small number of large clusters centered on key lines. Nevertheless, an average fixation index of 0.06 indicates moderate differentiation between the three major maize subpopulations. Linkage disequilibrium (LD) decays very rapidly, but the extent of LD is highly dependent on the particular group of germplasm and region of the genome. The utility of these data for performing genome-wide association studies was tested with two simply inherited traits and one complex trait. We identified trait associations at SNPs very close to known candidate genes for kernel color, sweet corn, and flowering time; however, results suggest that more SNPs are needed to better explore the genetic architecture of complex traits.

Conclusions

The genotypic information described here allows this publicly available panel to be exploited by researchers facing the challenges of sustainable agriculture through better knowledge of the nature of genetic diversity.     

Biogeography of the ecosystems of the healthy human body

Background

Characterizing the biogeography of the microbiome of healthy humans is essential for understanding microbial associated diseases. Previous studies mainly focused on a single body habitat from a limited set of subjects. Here, we analyzed one of the largest microbiome datasets to date and generated a biogeographical map that annotates the biodiversity, spatial relationships, and temporal stability of 22 habitats from 279 healthy humans.

Results

We identified 929 genera from more than 24 million 16S rRNA gene sequences of 22 habitats, and we provide a baseline of inter-subject variation for healthy adults. The oral habitat has the most stable microbiota with the highest alpha diversity, while the skin and vaginal microbiota are less stable and show lower alpha diversity. The level of biodiversity in one habitat is independent of the biodiversity of other habitats in the same individual. The abundances of a given genus at a body site in which it dominates do not correlate with the abundances at body sites where it is not dominant. Additionally, we observed the human microbiota exhibit both cosmopolitan and endemic features. Finally, comparing datasets of different projects revealed a project-based clustering pattern, emphasizing the significance of standardization of metagenomic studies.

Conclusions

The data presented here extend the definition of the human microbiome by providing a more complete and accurate picture of human microbiome biogeography, addressing questions best answered by a large dataset of subjects and body sites that are deeply sampled by sequencing.     

Caloric restriction augments radiation efficacy in breast cancer

Dietary modification such as caloric restriction (CR) has been shown to decrease tumor initiation and progression. We sought to determine if nutrient restriction could be used as a novel therapeutic intervention to enhance cytotoxic therapies such as radiation (IR) and alter the molecular profile of triple-negative breast cancer (TNBC), which displays a poor prognosis. In two murine models of TNBC, significant tumor regression is noted with IR or diet modification, and a greater regression is observed combining diet modification with IR. Two methods of diet modification were compared, and it was found that a daily 30% reduction in total calories provided more significant tumor regression than alternate day feeding. At the molecular level, tumors treated with CR and IR showed less proliferation and more apoptosis. cDNA array analysis demonstrated the IGF-1R pathway plays a key role in achieving this physiologic response, and multiple members of the IGF-1R pathway including IGF-1R, IRS, PIK3ca and mTOR were found to be downregulated. The innovative use of CR as a novel therapeutic option has the potential to change the biology of tumors and enhance the opportunity for clinical benefit in the treatment of patients with TNBC.     

Involvement of miRNAs in the Differentiation of Human Glioblastoma Multiforme Stem-Like Cells

Glioblastoma multiforme (GBM)-initiating cells (GICs) represent a tumor subpopulation with neural stem cell-like properties that is responsible for the development, progression and therapeutic resistance of human GBM. We have recently shown that blockade of NFκB pathway promotes terminal differentiation and senescence of GICs both in vitro and in vivo, indicating that induction of differentiation may be a potential therapeutic strategy for GBM. MicroRNAs have been implicated in the pathogenesis of GBM, but a high-throughput analysis of their role in GIC differentiation has not been reported. We have established human GIC cell lines that can be efficiently differentiated into cells expressing astrocytic and neuronal lineage markers. Using this in vitro system, a microarray-based high-throughput analysis to determine global expression changes of microRNAs during differentiation of GICs was performed. A number of changes in the levels of microRNAs were detected in differentiating GICs, including over-expression of hsa-miR-21, hsa-miR-29a, hsa-miR-29b, hsa-miR-221 and hsa-miR-222, and down-regulation of hsa-miR-93 and hsa-miR-106a. Functional studies showed that miR-21 over-expression in GICs induced comparable cell differentiation features and targeted SPRY1 mRNA, which encodes for a negative regulator of neural stem-cell differentiation. In addition, miR-221 and miR-222 inhibition in differentiated cells restored the expression of stem cell markers while reducing differentiation markers. Finally, miR-29a and miR-29b targeted MCL1 mRNA in GICs and increased apoptosis. Our study uncovers the microRNA dynamic expression changes occurring during differentiation of GICs, and identifies miR-21 and miR-221/222 as key regulators of this process.