Propagating Waves of Directionality and Coordination Orchestrate Collective Cell Migration

The ability of cells to coordinately migrate in groups is crucial to enable them to travel long distances during embryonic development, wound healing and tumorigenesis, but the fundamental mechanisms underlying intercellular coordination during collective cell migration remain elusive despite considerable research efforts. A novel analytical framework is introduced here to explicitly detect and quantify cell clusters that move coordinately in a monolayer. The analysis combines and associates vast amount of spatiotemporal data across multiple experiments into transparent quantitative measures to report the emergence of new modes of organized behavior during collective migration of tumor and epithelial cells in wound healing assays. First, we discovered the emergence of a wave of coordinated migration propagating backward from the wound front, which reflects formation of clusters of coordinately migrating cells that are generated further away from the wound edge and disintegrate close to the advancing front. This wave emerges in both normal and tumor cells, and is amplified by Met activation with hepatocyte growth factor/scatter factor. Second, Met activation was found to induce coinciding waves of cellular acceleration and stretching, which in turn trigger the emergence of a backward propagating wave of directional migration with about an hour phase lag. Assessments of the relations between the waves revealed that amplified coordinated migration is associated with the emergence of directional migration. Taken together, our data and simplified modeling-based assessments suggest that increased velocity leads to enhanced coordination: higher motility arises due to acceleration and stretching that seems to increase directionality by temporarily diminishing the velocity components orthogonal to the direction defined by the monolayer geometry. Spatial and temporal accumulation of directionality thus defines coordination. The findings offer new insight and suggest a basic cellular mechanism for long-term cell guidance and intercellular communication during collective cell migration.     

In vivo debonding strength and enamel damage in two orthodontic debonding methods

Bracket debonding strength related to diverse debonding methods and enamel damage has not been assessed in vivo. The study hypothetized a direct relationship between these three parameters. Debonding strength was measured clinically in the wings method and base method on 50 patients in a split mouth method using a calibrated debonding plier. Brackets from 30 of these patients were scanned in SEM and EDAX for adhesive remnant index and enamel calcium remnants. Base method debonding force was significantly greater than wings method (163.5+/-68.7 N, 106.1+/-66.2 N, respectively, p < 0.001). A positive adhesive remnant index score was found in both methods (68.7%, 66.7%, respectively). Debonding strength vs. adhesive remnant index or calcium index scores were not correlated. However, the latter two were significantly correlated (0.524 < R < 0.895, p < 0.031). Half of the debonding failures developed at the adhesive enamel interface. The results warnts the potential of enamel damage during debonding.     

Non‐canonical roles of tRNAs and tRNA mimics in bacterial cell biology

Transfer RNAs (tRNAs) are the macromolecules that transfer activated amino acids from aminoacyl‐tRNA synthetases to the ribosome, where they are used for the mRNA guided synthesis of proteins. Transfer RNAs are ancient molecules, perhaps even predating the existence of the translation machinery. Albeit old, these molecules are tremendously conserved, a characteristic that is well illustrated by the fact that some bacterial tRNAs are efficient and specific substrates of eukaryotic aminoacyl‐tRNA synthetases and ribosomes. Considering their ancient origin and high structural conservation, it is not surprising that tRNAs have been hijacked during evolution for functions outside of translation. These roles beyond translation include synthetic, regulatory and information functions within the cell. Here we provide an overview of the non‐canonical roles of tRNAs and their mimics in bacteria, and discuss some of the common themes that arise when comparing these different functions.     

Crowding alone cannot account for cosolute effect on amyloid aggregation

Amyloid fiber formation is a specific form of protein aggregation, often resulting from the misfolding of native proteins. Aimed at modeling the crowded environment of the cell, recent experiments showed a reduction in fibrillation halftimes for amyloid-forming peptides in the presence of cosolutes that are preferentially excluded from proteins and peptides. The effect of excluded cosolutes has previously been attributed to the large volume excluded by such inert cellular solutes, sometimes termed "macromolecular crowding". Here, we studied a model peptide that can fold to a stable monomeric β-hairpin conformation, but under certain solution conditions aggregates in the form of amyloid fibrils. Using Circular Dichroism spectroscopy (CD), we found that, in the presence of polyols and polyethylene glycols acting as excluded cosolutes, the monomeric β-hairpin conformation was stabilized with respect to the unfolded state. Stabilization free energy was linear with cosolute concentration, and grew with molecular volume, as would also be predicted by crowding models. After initiating the aggregation process with a pH jump, fibrillation in the presence and absence of cosolutes was followed by ThT fluorescence, transmission electron microscopy, and CD spectroscopy. Polyols (glycerol and sorbitol) increased the lag time for fibril formation and elevated the amount of aggregated peptide at equilibrium, in a cosolute size and concentration dependent manner. However, fibrillation rates remained almost unaffected by a wide range of molecular weights of soluble polyethylene glycols. Our results highlight the importance of other forces beyond the excluded volume interactions responsible for crowding that may contribute to the cosolute effects acting on amyloid formation.     

Mechanisms of Salinity Control in Sea Bass

Sea bass can regulate the concentration of Na⁺, K⁺, and Cl^-, among other ions, in their blood, skin, gills, and kidney. Therefore, the salinity of the water does not have a great influence on their metabolism, and sea bass can live in both sea and freshwater in accordance with the salt concentration. Most salinity control occurs in the gills, primarily through the control of chloride cells present there. The concentration of ions in the blood is controlled by the cotransporter Na⁺ / K⁺ / 2Cl^- (NKCC) in the chloride cell, and the subunits of Na⁺ / K⁺ ATPase (NKA) function to maintain homeostasis. The expression of NKA is regulated by subunits of the protein FXYD, allowing the sea bass to survive in compliance with the salinity. In this way, it is possible for sea bass to live in sea and freshwater by controlling the salinity of its body using functions of various channels, proteins, and genes present in the chloride cells of sea bass. In this study, we investigated recent studies of salt control mechanisms in sea bass and their application.     

Diagnosis of rotavirus, adenovirus, and herpesvirus infections by immune electron microscopy using a serum-in-agar diffusion method

The sensitivity of immune electron microscopy (IEM) for the detection and identification of bovine rotavirus, infectious bovine rhinotracheitis virus (IBR), and canine adenovirus has been studied by using the serum-in-agar (SIA) method in which a specific antiserum has been incorporated in agar.     

Variations in Lipid and Fatty Acid Content in Relation to Acetyl CoA Carboxylase in the Marine Prymnesiophyte Isochrysis galbana

Lipid and fatty acid content was determined in Isochrysis galbanacultures grown under various environmental conditions in steadystate continuous cultures. Lipid and fatty acid accumulationwas observed under severe nitrogen limiting conditions. Therate of in situ lipid synthesis, determined from ¹⁴C bicarbonateincorporation into lipid fraction, decreased under nitrogenlimited growth (µ<0.72day^–1), concomitant witha reduction in the in vitro activity of the enzyme acetyl CoAcarboxylase. Cellular lipid and fatty acid content remainedfairly constant over a wide range of irradiance levels. Therate of lipid synthesis, however, increased as irra-diance levelwas elevated to a maximal value at a light intensity of 150µmolquanta m^–2s^–1 and slightly decreased at a higherphoton flux. The in vitro activity of ACCase roughly followedthe same pattern of lipid synthesis in response to light intensity.The relative abundance of acetyl CoA carboxylase significantlydecreased in nitrogen limited cultures grown at low dilutionrates (µ<0.72day^–1). A fairly good linear correlationwas measured between the cellular content of ACCase and theenzyme activity in cultures grown under nitrogen limiting conditions.Furthermore, in nitrogen limited cultures, the cellular fattyacid content was linearly related to the cell capacity to producemalonyl CoA, the end product of ACCase and the building blockin fatty acid synthesis. (Received November 20, 1990; Accepted January 17, 1991)     

Optimized small‐molecule pull‐downs define MLBP1 as an acyl‐lipid‐binding protein

Abscisic acid (ABA) receptors belong to the START domain superfamily, which encompasses ligand‐binding proteins present in all kingdoms of life. START domain proteins contain a central binding pocket that, depending on the protein, can couple ligand binding to catalytic, transport or signaling functions. In Arabidopsis, the best characterized START domain proteins are the 14 PYR/PYL/RCAR ABA receptors, while the other members of the superfamily do not have assigned ligands. To address this, we used affinity purification of biotinylated proteins expressed transiently in Nicotiana benthamiana coupled to untargeted LC‐MS to identify candidate binding ligands. We optimized this method using ABA–PYL interactions and show that ABA co‐purifies with wild‐type PYL5 but not a binding site mutant. The K_d of PYL5 for ABA is 1.1 μm , which suggests that the method has sufficient sensitivity for many ligand–protein interactions. Using this method, we surveyed a set of 37 START domain‐related proteins, which resulted in the identification of ligands that co‐purified with MLBP1 (At4G01883) or MLP165 (At1G35260). Metabolite identification and the use of authentic standards revealed that MLBP1 binds to monolinolenin, which we confirmed using recombinant MLBP1. Monolinolenin also co‐purified with MLBP1 purified from transgenic Arabidopsis, demonstrating that the interaction occurs in a native context. Thus, deployment of this relatively simple method allowed us to define a protein–metabolite interaction and better understand protein–ligand interactions in plants.