Studies on fluid dynamics of the flow field and gas transfer in orbitally shaken tubes

Orbitally shaken cylindrical bioreactors [OrbShake bioreactors (OSRs)] without an impeller or sparger are increasingly being used for the suspension cultivation of mammalian cells. Among small volume OSRs, 50‐mL tubes with a ventilated cap (OSR50), originally derived from standard laboratory centrifuge tubes with a conical bottom, have found many applications including high‐throughput screening for the optimization of cell cultivation conditions. To better understand the fluid dynamics and gas transfer rates at the liquid surface in OSR50, we established a three‐dimensional simulation model of the unsteady liquid forms (waves) in this vessel. The studies verified that the operating conditions have a large effect on the interfacial surface. The volumetric mass transfer coefficient (k_La) was determined experimentally and from simulations under various working conditions. We also determined the liquid‐phase mass transfer coefficient (k_L) and the specific interfacial area (a) under different conditions to demonstrate that the value of a affected the gas transfer rate more than did the value of k_L. High oxygen transfer rates, sufficient for supporting the high‐density culture of mammalian cells, were found. Finally, the average axial velocity of the liquid was identified to be an important parameter for maintaining cells in suspension. Overall these studies provide valuable insights into the preferable operating conditions for the OSR50, such as those needed for cell cultures requiring high oxygen levels. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:192–200, 2017     

Identification of lectin receptors for conserved SARS‐CoV‐2 glycosylation sites

New SARS‐CoV‐2 variants are continuously emerging with critical implications for therapies or vaccinations. The 22 N‐glycan sites of Spike remain highly conserved among SARS‐CoV‐2 variants, opening an avenue for robust therapeutic intervention. Here we used a comprehensive library of mammalian carbohydrate‐binding proteins (lectins) to probe critical sugar residues on the full‐length trimeric Spike and the receptor binding domain (RBD) of SARS‐CoV‐2. Two lectins, Clec4g and CD209c, were identified to strongly bind to Spike. Clec4g and CD209c binding to Spike was dissected and visualized in real time and at single‐molecule resolution using atomic force microscopy. 3D modelling showed that both lectins can bind to a glycan within the RBD‐ACE2 interface and thus interferes with Spike binding to cell surfaces. Importantly, Clec4g and CD209c significantly reduced SARS‐CoV‐2 infections. These data report the first extensive map and 3D structural modelling of lectin‐Spike interactions and uncovers candidate receptors involved in Spike binding and SARS‐CoV‐2 infections. The capacity of CLEC4G and mCD209c lectins to block SARS‐CoV‐2 viral entry holds promise for pan‐variant therapeutic interventions.     

The NALCN ion channel is activated by M3 muscarinic receptors in a pancreatic β-cell line

A previously uncharacterized putative ion channel, NALCN (sodium leak channel, non-selective), has been recently shown to be responsible for the tetrodotoxin (TTX)-resistant sodium leak current implicated in the regulation of neuronal excitability. Here, we show that NALCN encodes a current that is activated by M3 muscarinic receptors (M3R) in a pancreatic β-cell line. This current is primarily permeant to sodium ions, independent of intracellular calcium stores and G proteins but dependent on Src activation, and resistant to TTX. The current is recapitulated by co-expression of NALCN and M3R in human embryonic kidney-293 cells and in Xenopus oocytes. We also show that NALCN and M3R belong to the same protein complex, involving the intracellular I–II loop of NALCN and the intracellular i3 loop of M3R. Taken together, our data show the molecular basis of a muscarinic-activated inward sodium current that is independent of G-protein activation, and provide new insights into the properties of NALCN channels.     

Influence of figure skating skates on vertical jumping performance

The purpose of this study was to investigate the influence of wearing figure skating skates on vertical jump performance and interjoint co-ordinations described in terms of sequencing and timing of joint rotations. Ten national to international figure skaters were filmed while performing a squat jump (SJ) on a force platform. Three experimental conditions were successively realized: barefoot (BF), lifting a 1.5 kg weight (LW) corresponding to the skates' mass, attached on the distal extremity of each leg and wearing skates (SK). Jump height, angular kinematics as well as joints kinetics were calculated. Relative to the SJ height reached in the BF condition, SJ performance was significantly decreased by 2.1 and 5.5 cm in the LW and SK conditions, respectively. The restriction of ankle amplitude imposed by wearing skates was found to significantly limit the knee joint amplitude while the hip angular motion was not affected. Neither the skates' mass nor the limited ankle angular motion modified the proximo-distal organization of joint co-ordination observed when jumping barefoot. However, with plantar flexion restriction, the delay between hip and knee extensions increased while it was reduced between knee and ankle extensions. Work output at the knee and ankle joints were significantly lowered when wearing skates. The decrease of work at the knee was shown to result from an early flexing moment causing a premature deceleration of the knee and from a reduction of knee amplitude. Taken together, these results show a minimization of the participation of the knee when plantar flexion is limited. It was proposed that constraining the distal joint causes a reorganization of interjoint co-ordinations and a redistribution of the energy produced by knee extensors to the hip and ankle joints.     

Desulfamplus magnetovallimortis gen. nov., sp. nov., a magnetotactic bacterium from a brackish desert spring able to biomineralize greigite and magnetite,that represents a novel lineage in the Desulfobacteraceae

A magnetotactic bacterium, designated strain BW-1^T, was isolated from a brackish spring in Death Valley National Park (California, USA) and cultivated in axenic culture. The Gram-negative cells of strain BW-1^T are relatively large and rod-shaped and possess a single polar flagellum (monotrichous). This strain is the first magnetotactic bacterium isolated in axenic culture capable of producing greigite and/or magnetite nanocrystals aligned in one or more chains per cell. Strain BW-1^T is an obligate anaerobe that grows chemoorganoheterotrophically while reducing sulfate as a terminal electron acceptor. Optimal growth occurred at pH 7.0 and 28 °C with fumarate as electron donor and carbon source. Based on its genome sequence, the G + C content is 40.72 mol %. Phylogenomic and phylogenetic analyses indicate that strain BW-1^T belongs to the Desulfobacteraceae family within the Deltaproteobacteria class. Based on average amino acid identity, strain BW-1^T can be considered as a novel species of a new genus, for which the name Desulfamplus magnetovallimortis is proposed. The type strain of D. magnetovallimortis is BW-1^T (JCM 18010^T–DSM 103535^T).     

SGO1 but not SGO2 is required for maintenance of centromere cohesion in Arabidopsis thaliana meiosis

Shugoshin is a protein conserved in eukaryotes and protects sister chromatid cohesion at centromeres in meiosis. In our study, we identified the homologs of SGO1 and SGO2 in Arabidopsis thaliana. We show that AtSGO1 is necessary for the maintenance of centromere cohesion in meiosis I since atsgo1 mutants display premature separation of sister chromatids starting from anaphase I. Furthermore, we show that the localization of the specific centromeric cohesin AtSYN1 is not affected in atsgo1, suggesting that SGO1 centromere cohesion maintenance is not mediated by protection of SYN1 from cleavage. Finally, we show that AtSGO2 is dispensable for both meiotic and mitotic cell progression in Arabidopsis.