No abstract

Legume GRAS (GAI, RGA, SCR)-type transcription factors NODULATION SIGNALING PATHWAY1 (NSP1) and NSP2 are essential for rhizobium Nod factor-induced nodulation. Both proteins are considered to be Nod factor response factors regulating gene expression after symbiotic signaling. However, legume NSP1 and NSP2 can be functionally replaced by nonlegume orthologs, including rice (Oryza sativa) NSP1 and NSP2, indicating that both proteins are functionally conserved in higher plants. Here, we show that NSP1 and NSP2 are indispensable for strigolactone (SL) biosynthesis in the legume Medicago truncatula and in rice. Mutant nsp1 plants do not produce SLs, whereas in M. truncatula, NSP2 is essential for conversion of orobanchol into didehydro-orobanchol, which is the main SL produced by this species. The disturbed SL biosynthesis in nsp1 nsp2 mutant backgrounds correlates with reduced expression of DWARF27, a gene essential for SL biosynthesis. Rice and M. truncatula represent distinct phylogenetic lineages that split approximately 150 million years ago. Therefore, we conclude that regulation of SL biosynthesis by NSP1 and NSP2 is an ancestral function conserved in higher plants. NSP1 and NSP2 are single-copy genes in legumes, which implies that both proteins fulfill dual regulatory functions to control downstream targets after rhizobium-induced signaling as well as SL biosynthesis in nonsymbiotic conditions.

The promise of cell therapy for repair and restoration of damaged tissues or organs relies on administration of large dose of cells whose healing benefits are still limited and sometimes irreproducible due to uncontrollable cell loss and death at lesion sites. Using a large amount of therapeutic cells increases the costs for cell processing and the risks of side effects. Optimal cell delivery strategies are therefore in urgent need to enhance the specificity, efficacy, and reproducibility of cell therapy leading to minimized cell dosage and side effects. Here, we addressed this unmet need by developing injectable 3D microscale cellular niches (microniches) based on biodegradable gelatin microcryogels (GMs). The microniches are constituted by in vitro priming human adipose-derived mesenchymal stem cells (hMSCs) seeded within GMs resulting in tissue-like ensembles with enriched extracellular matrices and enhanced cell-cell interactions. The primed 3D microniches facilitated cell protection from mechanical insults during injection and in vivo cell retention, survival, and ultimate therapeutic functions in treatment of critical limb ischemia (CLI) in mouse models compared with free cell-based therapy. In particular, 3D microniche-based therapy with 10 5 hMSCs realized better ischemic limb salvage than treatment with 10 6 freeinjected hMSCs, the minimum dosage with therapeutic effects for treating CLI in literature. To the best of our knowledge, this is the first convincing demonstration of injectable and primed cell delivery strategy realizing superior therapeutic efficacy for treating CLI with the lowest cell dosage in mouse models. This study offers a widely applicable cell delivery platform technology to boost the healing power of cell regenerative therapy.C ell-based regenerative therapy holds great promise for repair and restoration of damaged tissues or organs with numerous clinical trials and preclinical animal testing reported for treating complex diseases (1). Common route of cell administration for clinical cell therapy is based on either systematic administration (e.g., i.v. infusion), relying on cells homing to the lesion sites (2), or direct injection of cells into the damaged tissues (3). However, therapeutic benefits of the administered cells are still limited and sometimes irreproducible due to cell loss and cell death (4). Taking cell therapy for ischemic heart diseases as an example, only ∼5% of mesenchymal stem cells (MSCs) survived after being transplanted into an infarcted porcine heart (5). Mechanical damage during injection, high rate of cell loss and leakage to surrounding tissues, cell death due to lack of appropriate cell-cell and cell-matrix interactions in the ischemic and inflammatory lesion tissues could all contribute to poor cell retention, survival, functionality, and reproducibility of the treatment (6, 7).A rational solution to enhance the therapeutic efficacy and reproducibility of cell therapy is to administer a large dose of cells to ensure sufficient number of functional cells ...

Biomimetic cell-membrane-camouflaged nanoparticles with desirable features have been widely used for various biomedical applications. However, the current research focuses on single cell membrane coating and using multiple cell membranes for nanoparticle functionalization is still challenging. In this work, platelet (PLT) and leukocyte (WBC) membranes are fused, PLT-WBC hybrid membranes are coated onto magnetic beads, and then their surface is modified with specific antibodies. The resulting PLT-WBC hybrid membrane-coated immunomagnetic beads (HM-IMBs) inherit enhanced cancer cell binding ability from PLTs and reduce homologous WBC interaction from WBCs, and are further used for highly efficient and highly specific isolation of circulating tumor cells (CTCs). By using spiked blood samples, it is found that, compared with commercial IMBs, the cell separation efficiency of HM-IMBs is improved to 91.77% from 66.68% and the cell purity is improved to 96.98% from 66.53%. Furthermore, by using the HM-IMBs, highly pure CTCs are successfully identified in 19 out of 20 clinical blood samples collected from breast cancer patients. Finally, the robustness of HM-IMBs is verified in downstream CTC analysis such as the detection of PIK3CA gene mutations. It is anticipated that this novel hybrid membrane coating strategy will open new possibilities for overcoming the limitations of current theranostic platforms.

BackgroundStrigolactones are a class of plant hormones whose biosynthesis is activated in response to phosphate starvation. This involves several enzymes, including the carotenoid cleavage dioxygenases 7 (CCD7) and CCD8 and the carotenoid isomerase DWARF27 (D27). D27 expression is known to be responsive to phosphate starvation. In Medicago truncatula and rice (Oryza sativa) this transcriptional response requires the GRAS-type proteins NSP1 and NSP2; both proteins are essential for rhizobium induced root nodule formation in legumes. In line with this, we questioned whether MtNSP1-MtNSP2 dependent MtD27 regulation is co-opted in rhizobium symbiosis.ResultsWe provide evidence that MtD27 is involved in strigolactone biosynthesis in M. truncatula roots upon phosphate stress. Spatiotemporal expression studies revealed that this gene is also highly expressed in nodule primordia and subsequently becomes restricted to the meristem and distal infection zone of a mature nodules. A similar expression pattern was found for MtCCD7 and MtCCD8. Rhizobium lipo-chitooligosaccharide (LCO) application experiments revealed that of these genes MtD27 is most responsive in an MtNSP1 and MtNSP2 dependent manner. Symbiotic expression of MtD27 requires components of the symbiosis signaling pathway; including MtDMI1, MtDMI2, MtDMI3/MtCCaMK and in part MtERN1. This in contrast to MtD27 expression upon phosphate starvation, which only requires MtNSP1 and MtNSP2.ConclusionOur data show that the phosphate-starvation responsive strigolactone biosynthesis gene MtD27 is also rapidly induced by rhizobium LCO signals in an MtNSP1 and MtNSP2-dependent manner. Additionally, we show that MtD27 is co-expressed with MtCCD7 and MtCCD8 in nodule primordia and in the infection zone of mature nodules.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-015-0651-x) contains supplementary material, which is available to authorized users.

In the current work, a green and recyclable FeCl3-catalyzed deep eutectic solvent system (F-DES) was invented to fabricate cellulose nanocrystals (CNCs) with a high yield and excellent thermal stability. It was found that the optimum composition of the FeCl3-catalyzed deep eutectic solvent was composed of oxalic acid dihydrate (Oxd), choline chloride (ChCl), and FeCl3·6H2O in a mass ratio of 4:1:0.2 (corresponding to the molar ratio of 4.43:1:0.1). Results showed that CNCs with a diameter range of 5–20 nm and length of 50–300 nm could be isolated from bleached eucalyptus kraft pulp (BEKP) at a high yield (over 90% based on the cellulose content in BEKP) by a one-step F-DES treatment under mild conditions (80 °C, 6 h). The resultant CNCs showed a much higher thermal stability (onset thermal degradation temperature was over 310 °C) than the traditional sulfuric acid hydrolyzed ones and also exhibited superior dispersion stability in water due to the introduction of carboxyl groups on the surface of CNCs by esterification. In addition, the separated F-DES could be directly reused to produce CNCs at least three times. Intriguingly, all the components of the reused F-DES could be separated by a simple separation process with few pollutants releasing into the environment. Therefore, the F-DES process could be a green and economically feasible method for the preparation of thermally stable and dispersible CNCs.

In the past few years, cellulose nanomaterials obtained from lignocellulose have attracted extensive attention as functional nanomaterials with excellent properties and great application potentials in a variety of high-tech fields....

The phoPR operon encodes a response regulator, PhoP, and a histidine kinase, PhoR, which activate or repress genes of the Bacillus subtilis Pho regulon in response to an extracellular phosphate deficiency. Induction of phoPR upon phosphate starvation required activity of both PhoP and PhoR, suggesting autoregulation of the operon, a suggestion that is supported here by PhoP footprinting on the phoPR promoter. Primer extension analyses, using RNA from JH642 or isogenic sigE or sigB mutants isolated at different stages of growth and/or under different growth conditions, suggested that expression of the phoPR operon represents the sum of five promoters, each responding to a specific growth phase and environmental controls. The temporal expression of the phoPR promoters was investigated using in vitro transcription assays with RNA polymerase holoenzyme isolated at different stages of Pho induction, from JH642 or isogenic sigE or sigB mutants. In vitro transcription studies using reconstituted E A , E B , and E E holoenzymes identified P A4 and P A3 as E A promoters and P E2 as an E E promoter. Phosphorylated PhoP (PhoPϳP) enhanced transcription from each of these promoters. E B was sufficient for in vitro transcription of the P B1 promoter. P 5 was active only in a sigB mutant strain. These studies are the first to report a role for PhoPϳP in activation of promoters that also have activity in the absence of Pho regulon induction and an activation role for PhoPϳP at an E E promoter. Information concerning P B1 and P 5 creates a basis for further exploration of the regulatory coordination or overlap of the PhoPR and SigB regulons during phosphate starvation.Inorganic phosphate (P i ) is the limiting nutrient for biological growth in the soil, the natural habitat of Bacillus subtilis. To thrive in this environment where P i levels are often 2 to 3 orders of magnitude lower than levels of other required ions (29), B. subtilis has evolved complex regulatory systems for utilization of this limiting nutrient. At least three global regulatory systems are responsible for changes in gene expression upon phosphate deprivation. One set of genes is controlled either positively or negatively by the PhoP-PhoR two-component regulators, genes referred to as the Pho regulon genes (for review, see reference 12). Other genes that are induced upon phosphate limitation are dependent on SigB (1), an alternative stress sigma factor. A third class of genes is expressed under phosphate-limiting growth conditions that are independent of either SigB or PhoP-PhoR (1). The regulatory coordination between these three sets of genes is unclear, although up-regulation of certain Pho regulon genes has been reported in a sigB mutant strain (12, 33).Pho regulon genes are the most extensively studied set of phosphate-regulated genes in B. subtilis. Identification of genes of known function that are directly regulated by PhoP-PhoR provides insight into one strategy B. subtilis may use to deal with conditions of limiting phosphate. A high-affinity P i tra...