Deletion of BMP receptor type IB decreased bone mass in association with compromised osteoblastic differentiation of bone marrow mesenchymal progenitors

We previously found that disruption of two type I BMP receptors, Bmpr1a and Acvr1, respectively, in an osteoblast-specific manner, increased bone mass in mice. BMPR1B, another BMP type I receptor, is also capable of binding to BMP ligands and transduce BMP signaling. However, little is known about the function of BMPR1B in bone. In this study, we investigated the bone phenotype in Bmpr1b null mice and the impacts of loss of Bmpr1b on osteoblasts and osteoclasts. We found that deletion of Bmpr1b resulted in osteopenia in 8-week-old male mice, and the phenotype was transient and gender specific. The decreased bone mass was neither due to the changes in osteoblastic bone formation activity nor osteoclastic bone resorption activity in vivo. In vitro differentiation of Bmpr1b null osteoclasts was increased but resorption activity was decreased. Calvarial pre-osteoblasts from Bmpr1b mutant showed comparable differentiation capability in vitro, while they showed increased BMP-SMAD signaling in culture. Different from calvarial pre-osteoblasts, Bmpr1b mutant bone marrow mesenchymal progenitors showed compromised differentiation in vitro, which may be a reason for the osteopenic phenotype in the mutant mice. In conclusion, our results suggested that BMPR1B plays distinct roles from BMPR1A and ACVR1 in maintaining bone mass and transducing BMP signaling

Functional Assessment of EnvZ/OmpR Two-Component System in Shewanella oneidensis

EnvZ and OmpR constitute the bacterial two-component signal transduction system known to mediate osmotic stress response in a number of Gram-negative bacteria. In an effort to understand the mechanism through which Shewanella oneidensis senses and responds to environmental osmolarity changes, structure of the ompR-envZ operon was determined with Northern blotting assay and roles of the EnvZ/OmpR two-component system in response to various stresses were investigated with mutational analysis, quantitative reverse transcriptase PCR (qRT-PCR), and phenotype microarrays. Results from the mutational analysis and qRT-PCR suggested that the EnvZ/OmpR system contributed to osmotic stress response of S. oneidensis and very likely engaged a similar strategy employed by E. coli, which involved reciprocal regulation of two major porin coding genes. Additionally, the ompR-envZ system was also found related to cell motility. We further showed that the ompR-envZ dependent regulation of porin genes and motility resided almost completely on ompR and only partially on envZ, indicating additional mechanisms for OmpR phosphorylation. In contrast to E. coli lacking ompR-envZ, however, growth of S. oneidensis did not show a significant dependence on ompR-envZ even under osmotic stress. Further analysis with phenotype microarrays revealed that the S. oneidensis strains lacking a complete ompR-envZ system displayed hypersensitivities to a number of agents, especially in alkaline environment. Taken together, our results suggest that the function of the ompR-envZ system in S. oneidensis, although still connected with osmoregulation, has diverged considerably from that of E. coli. Additional mechanism must exist to support growth of S. oneidensis under osmotic stress

The mid-Miocene Zhangpu biota reveals an outstandingly rich rainforest biome in East Asia

During the Mid-Miocene Climatic Optimum [MMCO, ~14 to 17 million years (Ma) ago], global temperatures were similar to predicted temperatures for the coming century. Limited megathermal paleoclimatic and fossil data are known from this period, despite its potential as an analog for future climate conditions. Here, we report a rich middle Miocene rainforest biome, the Zhangpu biota (~14.7 Ma ago), based on material preserved in amber and associated sedimentary rocks from southeastern China. The record shows that the mid-Miocene rainforest reached at least 24.2°N and was more widespread than previously estimated. Our results not only highlight the role of tropical rainforests acting as evolutionary museums for biodiversity at the generic level but also suggest that the MMCO probably strongly shaped the East Asian biota via the northern expansion of the megathermal rainforest biome. The Zhangpu biota provides an ideal snapshot for biodiversity redistribution during global warming

Transcriptome analysis reveals the molecular mechanisms of the defense response to gray leaf spot disease in maize

Abstract Background Gray leaf spot (GLS), which is caused by the necrotrophic fungi Cercospora zeae-maydis and Cercospora zeina, is one of the most impactful diseases in maize worldwide. The aim of the present study is to identify the resistance genes and understand the molecular mechanisms for GLS resistance. Results Two cultivars, ‘Yayu889’ and ‘Zhenghong532,’ which are distinguished as resistant and susceptible cultivars, respectively, were challenged with the GLS disease and a RNA-seq experiment was conducted on infected plants at 81, 89, 91, and 93 days post planting (dap). Compared with the beginning stage at 81 dap, 4666, 1733, and 1166 differentially expressed genes (DEGs) were identified at 89, 91, and 93 dap, respectively, in ‘Yayu889,’ while relatively fewer, i.e., 4713, 881, and 722 DEGs, were identified in ‘Zhenghong532.’ Multiple pathways involved in the response of maize to GLS, including ‘response to salicylic acid,’ ‘protein phosphorylation,’ ‘oxidation-reduction process,’ and ‘carotenoid biosynthetic process,’ were enriched by combining differential expression analysis and Weighted Gene Co-expression Network Analysis (WGCNA). The expression of 12 candidate resistance proteins in these pathways were quantified by the multiple reaction monitoring (MRM) method. This approach identified two candidate resistance proteins, a calmodulin-like protein and a leucine-rich repeat receptor-like protein kinase with SNPs that were located in QTL regions for GLS resistance. Metabolic analysis showed that, compared with ‘Zhenghong532,’ the amount of salicylic acid (SA) and total carotenoids in ‘Yayu889’ increased, while peroxidase activity decreased during the early infection stages, suggesting that increased levels of SA, carotenoids, and reactive oxygen species (ROS) may enhance the defense response of ‘Yayu889’ to GLS. Conclusion By combining transcriptome and proteome analyses with comparisons of resistance QTL regions, calmodulin-like protein and leucine-rich repeat receptor-like protein kinase were identified as candidate GLS resistance proteins. Moreover, we found that the metabolic pathways for ROS, SA, and carotenoids are especially active in the resistant cultivar. These findings could lead to a better understanding of the GLS resistance mechanisms and facilitate the breeding of GLS-resistant maize cultivars

Cultivar Differences in Root Nitrogen Uptake Ability of Maize Hybrids

Although, considerable differences in root size in response to nitrogen (N) application among crop species and cultivars have been widely reported, there has been limited focus on the differences in root N uptake ability. In this study, two maize (Zea mays L.) hybrids, Zhenghong 311 (ZH 311, N-efficient) and Xianyu 508 (XY 508, N-inefficient), were used to compare differences in root N uptake ability. The two cultivars were grown in field pots Experiment I (Exp. I) and hydroponic cultures Experiment II (Exp. II) supplemented with different concentrations of N fertilizer. In both experiments, the levels of accumulated N were higher in ZH 311 than in XY 508 under low- and high-N supply, and the increment in accumulated N was greater under N deficiency. The maximum N uptake rate (Vm) and average N uptake rate (Va) in Exp. I, the root N kinetic parameter maximum uptake rate (Vmax) per fresh weight (FW) and Vmax per plant in Exp. II, and the root N uptake rate in both experiments were significantly higher for ZH 311 than for XY 508. In contrast, the root-to-shoot N ratio in both experiments and the root N kinetic parameter Michaelis constant (Km) in in Exp. II were markedly higher in XY 508 than in ZH 311, particularly under N-deficient conditions. Higher root N kinetic parameters Vmax per FW and Vmax per plant and lower Km values contributed to higher N affinity and uptake potential, more coordinated N distribution in the root and shoot, and higher root N uptake rates throughout the growth stages, thus enhancing the N accumulation and yield of the N-efficient maize cultivar. We conclude that the N uptake ability of roots in the N-efficient cultivar ZH 311 is significantly greater than that in the N-inefficient cultivar XY 508, and that this advantage is more pronounced under N-deficient conditions. The efficient N acquisition in ZH 311 is due to higher N uptake rate per root FW under optimal N conditions and the comprehensive effects of root size and N uptake rate per root FW under N deficiency

A Crp-Dependent Two-Component System Regulates Nitrate and Nitrite Respiration in <em>Shewanella oneidensis</em>

<div><p>We have previously illustrated the nitrate/nitrite respiratory pathway of <em>Shewanella oneidensis</em>, which is renowned for its remarkable versatility in respiration. Here we investigated the systems regulating the pathway with a reliable approach which enables characterization of mutants impaired in nitrate/nitrite respiration by guaranteeing biomass. The <em>S. oneidensis</em> genome encodes an <em>Escherichia coli</em> NarQ/NarX homolog SO3981 and two <em>E. coli</em> NarP/NarL homologs SO1860 and SO3982. Results of physiological characterization and mutational analyses demonstrated that <em>S. oneidensis</em> possesses a single two-component system (TCS) for regulation of nitrate/nitrite respiration, consisting of the sensor kinase SO3981(NarQ) and the response regulator SO3982(NarP). The TCS directly controls the transcription of <em>nap</em> and <em>nrfA</em> (genes encoding nitrate and nitrite reductases, respectively) but regulates the former less tightly than the latter. Additionally, phosphorylation at residue 57 of SO3982 is essential for its DNA-binding capacity. At the global control level, Crp is found to regulate expression of <em>narQP</em> as well as <em>nap</em> and <em>nrfA</em>. In contrast to NarP-NarQ, Crp is more essential for <em>nap</em> rather than <em>nrfA</em>.</p> </div

Binding analysis of NarP to <i>nap</i> and <i>nrfA</i> promoters.

<p><b>A.</b> EMSA assay with carbamoyl phosphate (lanes 2–6) and NarQ^51–585 (lanes 7–11) treated NarP. NarP^# represents NarP carrying a D57N mutation. Binding assays with NarP^# treated with NarQ^51–585 and ATP are shown in lanes 12–13. All of the binding assays were performed with 2 ng of <i>nrfA</i> upstream fragments in the presence of 2 µg non-specific competitor DNA poly(dI<b>·</b>dC). Lanes 6 and 11 contain 10 µM of unlabeled <i>nrfA</i> upstream fragments as competitor DNA. The concentration of NarP or NarP^# is indicated in the figure (µM). <b>B.</b> The EMSA assay was performed with 2 µM phosphorylated NarP and various amounts of ³³P end-labeled <i>nrfA</i> (−50 to −200 relative to the translation start codon) and <i>nap</i> (−50 to −200) upstream fragments. Non-specific competitor DNA, 2 µg poly(dI·dC), was added in all lanes. <b>C.</b> Western blotting analysis. Upper panel, analysis of NrfA in Δ<i>narP</i>. Cells grown in the presence of nitrite at the indicated time points were assayed. Lower panel, analysis of NarP. Cells grown in the presence of nitrate and/or nitrite at the indicated time points were assayed. In both panels, Δ<i>narP^c</i> represents Δ<i>narP</i> containing pHG102-<i>narP</i> (P<i>_arcA-narP</i>), in which <i>narP</i> is over-expressed. <b>D.</b> qRT-PCR analysis of the <i>narQ-narP</i> operon. The wild type cells grown with nitrate or nitrate aerobically were collected at the indicated time points and assayed. Abundance is given relative to 16S rRNA. Error bars represent the standard deviation (SD) (<i>n</i> = 3).</p

Crp and Fnr in aerobic nitrate/nitrite respiration of <i>S. oneidensis</i>.

<p><b>A.</b> qRT-PCR analysis of <i>crp</i> and <i>fnr</i>. The wild type cells grown in the presence or absence of nitrate or nitrate aerobically were collected at the indicated time points and assayed. Expression level of each gene was presented under three conditions: –, no addition of nitrate or nitrite, NO₃⁻, nitrate added, and NO₂⁻, nitrite added. Abundance is given relative to 16 S rRNA. <b>B.</b> Western blotting analysis of Crp. Upper panel, the wild type cells cultured in the absence of nitrate or presence of nitrate at 4, 8, and 12 h were assayed. Lower panel, the wild type cells cultured in the absence of nitrite or presence of nitrite at 8, 12, and 16 h were assayed. Δ<i>crp</i> was used as the negative control. <b>C.</b> qRT-PCR analysis of <i>nap</i>, <i>nrfA</i>, and <i>narP</i> in cells grown with nitrate. The wild type, Δ<i>crp</i>, and Δ<i>fnr</i> mutant strains were assayed at indicated time points. Abundance is given relative to 16 S rRNA. <b>D.</b> Nitrate/nitrite assay. Cells of tested strains grown aerobically in the presence of nitrate or nitrate were collected at the indicated times. Concentrations of nitrate and nitrite (8 h and after) remaining in cultures were measured. Δ<i>crp^c</i> represented the mutant containing a copy of <i>crp</i> on the complementation plasmid. Solid and dash lines represent cells grown in the presence of nitrate and nitrite, respectively. The wild type and Δ<i>fnr</i> were indistinguishable from each other and thus data for the wild type and Error bars (SD, <i>n</i> = 4) were omitted for clarity.</p