Autonomous self-burying seed carriers for aerial seeding.

Aerial seeding can quickly cover large and physically inaccessible areas1 to improve soil quality and scavenge residual nitrogen in agriculture2, and for postfire reforestation3-5 and wildland restoration6,7. However, it suffers from low germination rates, due to the direct exposure of unburied seeds to harsh sunlight, wind and granivorous birds, as well as undesirable air humidity and temperature1,8,9. Here, inspired by Erodium seeds10-14, we design and fabricate self-drilling seed carriers, turning wood veneer into highly stiff (about 4.9 GPa when dry, and about 1.3 GPa when wet) and hygromorphic bending or coiling actuators with an extremely large bending curvature (1,854 m-1), 45 times larger than the values in the literature15-18. Our three-tailed carrier has an 80% drilling success rate on flat land after two triggering cycles, due to the beneficial resting angle (25°-30°) of its tail anchoring, whereas the natural Erodium seed's success rate is 0%. Our carriers can carry payloads of various sizes and contents including biofertilizers and plant seeds as large as those of whitebark pine, which are about 11 mm in length and about 72 mg. We compare data from experiments and numerical simulation to elucidate the curvature transformation and actuation mechanisms to guide the design and optimization of the seed carriers. Our system will improve the effectiveness of aerial seeding to relieve agricultural and environmental stresses, and has potential applications in energy harvesting, soft robotics and sustainable buildings.

Role of Average Volume Assured Pressure Support Mode (AVAPS) in the Management of Acute Exacerbation of Chronic Obstructive Pulmonary Disease With Type 2 Respiratory Failure.

Background Non-invasive ventilation (NIV) is a well-established approach in the treatment of acute exacerbation of chronic obstructive pulmonary disease (COPD) with type 2 respiratory failure. Average volume-assured pressure support (AVAPS) mode integrates the characteristics of both volume and pressure-controlled modes of NIV. In bilevel positive airway pressure (BiPAP) mode, volume is the dependent variable, whereas in AVAPS mode, pressure is the dependent variable. In this study, we aimed to compare the role of AVAPS mode with BiPAP spontaneous/timed (S/T) mode for the management of patients with acute exacerbation of COPD with type 2 respiratory failure. Methodology A hospital-based comparative and analytical study was carried out on 100 patients with acute exacerbation of COPD with type 2 respiratory failure admitted to respiratory disease hospital, Sardar Patel Medical College, Bikaner (Rajasthan, India). Patients were randomly divided into two groups of 50 patients each. Group A patients were treated with AVAPS mode and group B patients with BiPAP (S/T) mode. Arterial blood gases, average duration of hospital stay, and need for invasive mechanical ventilation were compared between the two groups. Results There was a statistically significant difference in favor of group A in terms of improvement in pH and pCO2 as compared to group B at 6 h (pH, p=0.027; pCO2, p=0.012) and 24 h (pH, p=0.032; pCO2, p=0.013). The duration of hospital stay was found to be lower in group A (p=0.003). However, no significant difference was found in terms of need for invasive mechanical ventilation between both groups (p=0.338). Conclusion Application of AVAPS mode results in more rapid and steady improvement in patients of COPD as compared to BiPAP (S/T) mode. Thus, management through non-invasive ventilation AVAPS mode should be considered in patients with acute exacerbation of COPD with type 2 respiratory failure.

α- and ß-Tubulin Lattice of the Axonemal Microtubule Doublet and Binding Proteins Revealed by Single Particle Cryo-Electron Microscopy and Tomography.

Microtubule doublet (MTD) is the main skeleton of cilia/flagella. Many proteins, such as dyneins and radial spokes, bind to MTD, and generate or regulate force. While the structure of the reconstituted microtubule has been solved at atomic resolution, nature of the axonemal MTD is still unclear. There are a few hypotheses of the lattice arrangement of its α- and ß-tubulins, but it has not been described how dyneins and radial spokes bind to MTD. In this study, we analyzed the three-dimensional structure of Tetrahymena MTD at ∼19 Å resolution by single particle cryo-electron microscopy. To identify α- and ß-tubulins, we combined image analysis of MTD with specific kinesin decoration. This work reveals that α- and ß-tubulins form a B-lattice arrangement in the entire MTD with a seam at the outer junction. We revealed the unique way in which inner arm dyneins, radial spokes, and proteins inside MTD bind and bridge protofilaments.

Cartwheel architecture of Trichonympha basal body.

Centrioles and basal bodies are essential for the formation of cilia, flagella, and centrosomes. They exhibit a characteristic ninefold symmetry imparted by a cartwheel thought to contain rings of SAS-6 proteins. We used cryoelectron tomography to investigate the architecture of the exceptionally long cartwheel of the flagellate Trichonympha. We found that the cartwheel is a stack of central rings that exhibit a vertical periodicity of 8.5 nanometers and is able to accommodate nine SAS-6 homodimers. The spokes that emanate from two such rings associate into a layer, with a vertical periodicity of 17 nanometers on the cartwheel margin. Thus, by using the power of biodiversity, we unveiled the architecture of the cartwheel at the root of the ninefold symmetry of centrioles and basal bodies.

Heterogeneity of dynein structure implies coordinated suppression of dynein motor activity in the axoneme.

Axonemal dyneins provide the driving force for flagellar/ciliary bending. Nucleotide-induced conformational changes of flagellar dynein have been found both in vitro and in situ by electron microscopy, and in situ studies demonstrated the coexistence of at least two conformations in axonemes in the presence of nucleotides (the apo and the nucleotide-bound forms). The distribution of the two forms suggested cooperativity between adjacent dyneins on axonemal microtubule doublets. Although the mechanism of such cooperativity is unknown it might be related to the mechanism of bending. To explore the mechanism by which structural heterogeneity of axonemal dyneins is induced by nucleotides, we used cilia from Tetrahymena thermophila to examine the structure of dyneins in a) the intact axoneme and b) microtubule doublets separated from the axoneme, both with and without additional pure microtubules. We also employed an ATPase assay on these specimens to investigate dynein activity functionally. Dyneins on separated doublets show more activation by nucleotides than those in the intact axoneme, both structurally and in the ATPase assay, and this is especially pronounced when the doublets are coupled with added microtubules, as expected. Paralleling the reduced ATPase activity in the intact axonemes, a lower proportion of these dyneins are in the nucleotide-bound form. This indicates a coordinated suppression of dynein activity in the axoneme, which could be the key for understanding the bending mechanism.

Comparative structural analysis of eukaryotic flagella and cilia from Chlamydomonas, Tetrahymena, and sea urchins.

Although eukaryotic flagella and cilia all share the basic 9+2 microtubule-organization of their internal axonemes, and are capable of generating bending-motion, the waveforms, amplitudes, and velocities of the bending-motions are quite diverse. To explore the structural basis of this functional diversity of flagella and cilia, we here compare the axonemal structure of three different organisms with widely divergent bending-motions by electron cryo-tomography. We reconstruct the 3D structure of the axoneme of Tetrahymena cilia, and compare it with the axoneme of the flagellum of sea urchin sperm, as well as with the axoneme of Chlamydomonas flagella, which we analyzed previously. This comparative structural analysis defines the diversity of molecular architectures in these organisms, and forms the basis for future correlation with their different bending-motions.

Cryoelectron tomography of radial spokes in cilia and flagella.

Radial spokes (RSs) are ubiquitous components in the 9 + 2 axoneme thought to be mechanochemical transducers involved in local control of dynein-driven microtubule sliding. They are composed of >23 polypeptides, whose interactions and placement must be deciphered to understand RS function. In this paper, we show the detailed three-dimensional (3D) structure of RS in situ in Chlamydomonas reinhardtii flagella and Tetrahymena thermophila cilia that we obtained using cryoelectron tomography (cryo-ET). We clarify similarities and differences between the three spoke species, RS1, RS2, and RS3, in T. thermophila and in C. reinhardtii and show that part of RS3 is conserved in C. reinhardtii, which only has two species of complete RSs. By analyzing C. reinhardtii mutants, we identified the specific location of subsets of RS proteins (RSPs). Our 3D reconstructions show a twofold symmetry, suggesting that fully assembled RSs are produced by dimerization. Based on our cryo-ET data, we propose models of subdomain organization within the RS as well as interactions between RSPs and with other axonemal components.

Equilibrium and kinetics of the unfolding and refolding of Escherichia coli Malate Synthase G monitored by circular dichroism and fluorescence spectroscopy.

The equilibrium and kinetics studies of an 82 kDa large monomeric Escherichia coli protein Malate Synthase G (MSG) was investigated by far and near-UV CD, intrinsic tryptophan fluorescence and extrinsic fluorescence spectroscopy. We find that despite of its large size, folding is reversible, in vitro. Equilibrium unfolding process of MSG exhibited three-state transition thus, indicating the presence of at least a stable equilibrium intermediate. Thermodynamic parameters suggest this intermediate resembles the unfolded state. However, the equilibrium intermediate exhibits pronounced secondary structure as measured by far-UV CD, partial tertiary structure as delineated by near-UV CD, compactness (m value) and exposed hydrophobic surface area as assessed by ANS binding, typically depicting a molten globule state. The stopped-flow kinetic data provide clear evidence for the presence of a burst phase during the refolding pathway due to the formation of an early Intermediate, within the dead time of the instrument. Refolding from 4 M to various lower concentrations until 0.4 M of GdnHCl follow biphasic kinetics at lower concentrations of GdnHCl (<0.8 M), whereas monophasic kinetics at concentrations above 1.5 M. Also, rollover in the refolding and unfolding limbs of chevron plot verifies the presence of a fast kinetic intermediate at lower concentration of GdnHCl. Based upon the above observations we hereby propose the folding pathway of a large multi-domain protein Malate Synthase G.

GroEL assisted folding of large polypeptide substrates in Escherichia coli: Present scenario and assignments for the future.

Escherichia coli chaperonins GroEL and GroES are indispensable for survival and growth of the cell since they provide essential assistance to the folding of many newly translated proteins in the cell. Recent studies indicate that a substantial portion of the proteins involved in the host pathways are completely dependent on GroEL-GroES for their folding and hence providing some explanation for why GroEL is essential for cell growth. Many proteins either small-single domain or large multidomains require assistance from GroEL-ES during their lifetime. Proteins of size up to approximately 70kDa can fold via the cis mechanism during GroEL-ES assisted pathway, but other proteins (>70kDa) that cannot be pushed inside the cavity of GroEL-ATP complex upon binding of GroES fold by an evolved mechanism called trans. In recent years, much work has been done on revealing facts about the cis mechanism involving the GroEL assisted folding of small proteins whereas the trans mechanism with larger polypeptide substrates still remains under cover. In order to disentangle the role of chaperonin GroEL-GroES in the folding of large E. coli proteins, this review discusses a number of issues like the range of large polypeptide substrates acted on by GroEL. Do all these substrates need the complete chaperonin system along with ATP for their folding? Does GroEL act as foldase or holdase during the process? We conclude with a discussion of the various queries that need to be resolved in the future for an extensive understanding of the mechanism of GroEL mediated folding of large substrate proteins in E. coli cytosol.