Chinese hamster ovary (CHO) cells are currently the most widely used host cells for recombinant therapeutic protein (RTP) production. Currently, the RTP yields need to increase further to meet the market needs and reduce costs. In this study, three stabilizing and anti-repressor (SAR) elements from the human genome were selected, including human SAR7, SAR40, and SAR44 elements. SAR elements were cloned upstream of the promoter in the eukaryotic vector, followed by transfection into CHO cells, and were screened under G418 pressure. Flow cytometry was used to detect enhanced green fluorescent protein (eGFP) expression levels. The gene copy numbers and mRNA expression levels were determined through quantitative real-time PCR. Furthermore, the effect of the stronger SAR elements on adalimumab was investigated. The results showed that transgene expression levels in the SAR-containing vectors were higher than that of the control vector, and SAR7 and SAR40 significantly increased and maintained the long-term expression of the transgene in CHO cells. In addition, the transgene expression level increase was related with gene copy numbers and mRNA expression levels. Collectively, SAR elements can enhance the transgene expression and maintain the long-term expression of a transgene in transfected CHO cells, which may be used to increase recombinant protein production in CHO cells.
Significance Stomatal openings in plant leaves admit CO 2 and release water vapor between plant and atmosphere. Environmental stimuli such as darkness or dryness lead to closure, whereas light or low CO 2 stimulate opening. Each stoma is defined by a pair of guard cells, and its pore aperture is controlled by ion channels that regulate the turgor pressure in these cells. SLAC1 anion channels are of central importance here, and environmental factors affect channel activity by controlling SLAC1 phosphorylation. Our studies define multiple sites of regulatory phosphorylation in the context of an atomic structure of the SLAC1 channel, providing a mechanistic understanding of the fine-tuning of channel activity and thereby of stomatal apertures in response to the environment.
Ce:GGAG is promising as scintillators. We present a traveling solvent floating zone method (TSFZ) to stabilize the molten zone. The obtained Ce:GGAG crystals demonstrate a fast decay (64 ns) with higher ratio of fast component (85%).
Aluminum-activated malate transporters (ALMTs) form an anion channel family that plays essential roles in diverse functions in plants. Arabidopsis ALMT12, also named QUAC1 (quick anion channel 1), regulates stomatal closure in response to environmental stimuli. However, the molecular basis of ALMT12/QUAC1 activity remains elusive. Here, we describe the cryo-EM structure of ALMT12/QUAC1 from Glycine max at 3.5-Å resolution. GmALMT12/QUAC1 is a symmetrical dimer, forming a single electropositive T-shaped pore across the membrane. The transmembrane and cytoplasmic domains are assembled into a twisted two-layer architecture, with their associated dimeric interfaces nearly perpendicular. GmALMT12/QUAC1-mediated currents display rapid kinetics of activation/deactivation and a bell-shaped voltage dependency, reminiscent of the rapid (R)-type anion currents. Our structural and functional analyses reveal a domain-twisting mechanism for malate-mediated activation. Together, our study uncovers the molecular basis for a previously uncharacterized class of anion channels and provides insights into the gating and modulation of the ALMT12/QUAC1 anion channel.
Trimeric intracellular cation (TRIC) channels are thought to provide counter-ion currents that facilitate the active release of Ca 2+ from intracellular stores. TRIC activity is controlled by voltage and Ca 2+ modulation, but underlying mechanisms have remained unknown. Here we describe high-resolution crystal structures of vertebrate TRIC-A and TRIC-B channels, both in Ca 2+ -bound and Ca 2+ -free states, and we analyze conductance properties in structure-inspired mutagenesis experiments. The TRIC channels are symmetric trimers, wherein we find a pore in each protomer that is gated by a highly conserved lysine residue. In the resting state, Ca 2+ binding at the luminal surface of TRIC-A, on its threefold axis, stabilizes lysine blockage of the pores. During active Ca 2+ release, luminal Ca 2+ depletion removes inhibition to permit the lysine-bearing and voltage-sensing helix to move in response to consequent membrane hyperpolarization. Diacylglycerol is found at interprotomer interfaces, suggesting a role in metabolic control.
Increasing spikelets differentiation, reducing their degradation and promoting big panicle formation are crucial approaches to increasing grain yield in rice. Straw incorporation and nitrogen (N) application have vital effects on rice yield, but the influence of incorporating wheat straw with N fertilizer ratio on spikelet differentiation, degeneration, and grain yield in rice is not clear. In this experiment, hybrid rice Fengyouxiangzhan (FY) and Yongyou 2640 (YY) were used as materials. Under the condition of total nitrogen application rate of 270 kg ha− 2, the effects of local farmers' fertilizer practice (FP, basal fertilizer: tillering fertilizer: panicle fertilizer = 5: 1: 4) and improving basal fertilizer proportion (IP, basal fertilizer: tillering fertilizer: panicle fertilizer = 7: 1: 2) on the grain yield, branch and spikelet differentiation and degeneration of the above rice varieties were studied under the conditions of non-wheat straw returning (NR) and wheat straw full returning (WR). The 2-year (2020–2021) results in field trials showed that: (1) under NR, compared with FP, although the panicles per unit area were increased, the spikelet number per panicle, filled grains, and grain weight decreased to varying degrees, and finally the grain yield decreased significantly in IP. Under WR, although the filled grains and grain weight were decreased in IP treatment, the total spikelets (panicles per unit area × spikelet number per panicle) was increased significantly, and the grain yield was finally increased significantly compared with FP, with an increase of 6.85 ~ 7.08%. (2) There was a strong positive link between the total spikelets and grain yield, but a substantial negative relationship between the total spikelets and the number of filled grains and grain weight. (3) Straw treatment and nitrogen treatment mainly affected the spikelet number per panicle by affecting the differentiation and degeneration of secondary branches and spikelets. Under WR, IP increased the differentiation number of secondary spikelet in the middle and basal parts of FY by 22.0% and 30.9% with FP, respectively, but also increased the number and rate of secondary spikelet degeneration in the middle and basal parts. Under IP, compared with NR treatment, the differentiated number, degenerated number, surviving number, and degradation rate of the middle and basal parts of the secondary spikelets in the two varieties were increased in WR treatment. Overall, these results demonstrated that under WR, IP was beneficial to increase effective panicle numbers and the spikelet number per panicle. More secondary spikelet differentiation in the middle and basal parts per panicle was a main reason for increasing the spikelet number per panicle to increase grain yield under IP treatment.
Heavy precipitation events occur often over the western Sichuan Basin in summer, near the transition zone between the Sichuan Basin and the steep terrain of the Tibetan Plateau. One such event — a heavy precipitation process that occurred on 18–20 August 2010, with clear nocturnal peaks — is chosen as a case to tentatively explore how the convection associated with convective-scale precipitation is initiated and propagated. By utilizing the vertical momentum equation from the viewpoint of separating perturbation pressure into dynamic and thermal parts, it is demonstrated that the vertical momentum is induced by the imbalance of several forces, including the dynamic/buoyant part of the perturbation pressure gradient force and the buoyancy force, with the latter dominating during the nocturnal-peak period. Although a negative value of the dynamic perturbation pressure gradient force partly offsets the positive buoyant forcing inside the strong updraft, the pattern of vertical motion tendency is largely attributable to its buoyancy because of its larger magnitude. Relative to the buoyancy component, the dynamic part of the vertical perturbation pressure gradient is also examined, revealing a smaller order of magnitude. Thus, it is the thermal effect that should be responsible for the initiation and propagation of convection. As for the convective-scale precipitation, it always presents a trailing morphology relative to the strong leading-side updraft. Furthermore, overlapping strong signals of vertical motion and its tendency point towards strong precipitation in the future.
Exploring the mechanism of cadmium (Cd) accumulation in Cd-safe rice lines is beneficial for ensuring rice safety. D62B, a Cd-safe rice line, accumulates less than 0.2 mg Cd kg-1 in the brown rice due to strong capacity of Cd retention in the roots, and the root cell wall (RCW) polysaccharides play important roles. However, specific underlying mechanism of Cd binding on the polysaccharides is little known. In this study, the role of polysaccharides, especially pectin and hemicellulose 1 (HC1), in RCW of D62B was investigated by adsorption experiments and Fourier Transform Infrared Spectroscopy (FTIR) analysis compared with a common rice line (Luhui17). Cadmium was adsorbed on RCW of two rice lines by a multilayer and inhomogeneous chemisorption way with the force of ion transfer or exchange. Cadmium was adsorbed on RCW rapidly at first stage with the limit of internal and external diffusion, and gradually reached saturation. With the removal of pectin, the Cd adsorption rate, maximum Cd adsorption amount and the shift degree of carboxyl groups in the RCW of D62B sharply decreased, which showed advantages compared with Luhui17. Sequential removal of HC1 further decreased the maximum Cd adsorption amount and the shift degree of hydroxyl groups. The results showed that more available functional groups, especially carboxyl groups in pectin and hydroxyl groups in HC1, contributed to Cd immobilization within the RCW of Cd-safe rice line, thus limiting Cd translocation to the shoot and reducing Cd accumulation in the brown rice.
Abstract The objectives of this study were to investigate the allelopathic effects of sesame extracts of on seed germination of moso bamboo, and to isolate and identify the potential allelochemicals. A factorial design with three organs (root, stem and leaf) and five concentrations (0, 25, 50, 75 and 100 mg mL −1 ) was carried out. Seeds of moso bamboo were soaked in sesame extracts to investigate their germination and growth. The allelochemicals were isolated and identified using the high performance liquid chromatograph (HPLC) system. The germination indices of the same organ decreased with the increase of extract concentrations, while the mean germination time increased, picking at the concentration of 100 mg mL −1 . The radicle length and plumule length decreased, while the ratio of radicle length to plumule length increased. The allelopathy inhibition effect increased with the increase of extract concentrations, and it was significantly higher at the concentration of 100 mg mL −1 than that of 25 mg mL −1 . The synthesis effect increased with the increase of extract concentrations, and it was significantly higher in leaf than root and stem. Chemical analyses identified 9 allelochemicals species (mostly phenolics and alkaloids) in the aqueous extracts. These results indicated that aqueous extracts of sesame caused the delay in seed germination and growth of moso bamboo, and phenolics and alkaloids in the aqueous extracts maybe the major reasons for the observed inhibition effects of sesame.
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