The repeat region of the circumsporozoite protein is an elastic linear spring with a functional role in Plasmodium sporozoite motility

The circumsporozoite protein (CSP) forms a dense coat on the surface of the sporozoite, the infective stage of the malaria parasite. The central repeat region of CSP is a critical component of the only licensed malaria vaccine yet little is known about its structure or function. We found that sporozoite mutants with severely truncated or scrambled repeats have impaired motility due to altered adhesion site formation and dynamics, suggesting that the CSP repeats provide a cohesive environment in which adhesion sites can form. We hypothesized that biophysical properties of the repeats are important in this role and interrogated this using single-molecule fluorescence-force spectroscopy. We show that the repeats are a stiff, linear spring with elastic properties, dependent upon length and lost when the repeats are scrambled. These data are the first evidence that the CSP repeat region serves a functional role during infection and motility, likely mediated through its biophysical properties.

Structural insights into integrin α5β1 opening by fibronectin ligand

Integrin α5β1 is a major fibronectin receptor critical for cell migration. Upon complex formation, fibronectin and α5β1 undergo conformational changes. While this is key for cell-tissue connections, its mechanism is unknown. Here, we report cryo–electron microscopy structures of native human α5β1 with fibronectin to 3.1-angstrom resolution, and in its resting state to 4.6-angstrom resolution. The α5β1-fibronectin complex revealed simultaneous interactions at the arginine-glycine-aspartate loop, the synergy site, and a newly identified binding site proximal to adjacent to metal ion–dependent adhesion site, inducing the translocation of helix α1 to secure integrin opening. Resting α5β1 adopts an incompletely bent conformation, challenging the model of integrin sharp bending inhibiting ligand binding. Our biochemical and structural analyses showed that affinity of α5β1 for fibronectin is increased with manganese ions (Mn2+) while adopting the half-bent conformation, indicating that ligand-binding affinity does not depend on conformation, and α5β1 opening is induced by ligand-binding.

Hamstring autograft and anatomical footprint evaluation for anterior talofibular ligament reconstruction: Cadaveric study

Purpose: The aim of the study was to evaluate whether or not there was any incompatibility between two-strand hamstring tendons taken from the same knee and the ATFL and it was the determination of suitable footprint points in the fibula and talus for anatomical ATFL reconstruction. Methods: 16 fresh frozen cadaver specimens were dissected to gracilis and semitendinosus tendons and the anterior talofibular ligament. The origins, insertions, distances from osseous landmarks of fibular talus of ATFL were determined. The diameters of gracilis, semitendinosus and ATFL were calculated. There was a moderate correlation between body height and the distance between the distal of inferior lateral malleolus and the fibular adhesion site of ATFL (r: 36.5 p: 0.036). There was a weak correlation between body height and the distance between the apex of the lateral talar process and the talus adhesion site of ATFL in a single bundle (r: 28.4 p: 0.002). There was no correlation between the distance from proximal and distal adhesion side of ATFL and body height in the double bundle (p: 0.241). Results: There was no significant relationship between ATFL diameter and gracilis, semitendinosus and both hamstring in women. A significant relationship at 80.5% was determined between the ATFL and the gracilis diameter in man. A significant relationship at 92.6% was determined between the ATFL and the semitendinosus diameter in man. Conclusion: It was determined that there is not compatibility between the gracilis tendons, the semitendinosus tendon and ATFL in women. It should be supported by biomechanical and clinical studies whether this incompatibility has a clinical effect or not.

Revealing the polarity of actin filaments by cryo-electron tomography

SummaryThe actin cytoskeleton plays a fundamental role in numerous cellular processes, such as cell motility, cytokinesis, and adhesion to the extracellular matrix. Revealing the polarity of individual actin filaments in cells, would foster an unprecedented understanding of cytoskeletal processes and their associated mechanical forces. Cryo-electron tomography provides the means for high-resolution structural imaging of cells. However, the low signal-to-noise ratio of cryo-tomograms obscures the high frequencies and therefore the polarity of actin filaments cannot be directly measured. Here, we developed an approach that enables to determine the polarity of actin filaments in cellular cryo-tomograms. We applied it to reveal the actin polarity distribution in focal adhesions, and show a linear relation between actin polarity and distance from the apical boundary of the adhesion site.

VARIATION AND SELECTION IN AXON NAVIGATION THROUGH MICROTUBULE-DEPENDENT STEPWISE GROWTH CONE ADVANCE

ABSTRACTMyosin II (MII) activity is required for elongating mammalian sensory axons to change speed and direction in response to Nerve Growth Factor (NGF) and laminin-1 (LN). NGF signaling induces faster outgrowth on LN through regulation of actomyosin restraint of microtubule advance into the growth cone periphery. It remains unclear whether growth cone turning on LN works through the same mechanism and, if it does, how the mechanism produces directed advance. Using a novel method for substrate patterning, we tested how directed advance occurs on LN by creating a gap immediately in front of a growth cone advancing on a narrow LN path. The growth cone stopped until an actin-rich protrusion extended over the gap, adhered to LN, and became stabilized. Stepwise advance over the gap was triggered by microtubule +tip entry up to the adhesion site of the protrusion and was independent of traction force pulling. We found that the probability of microtubule entry is regulated at the level of the individual protrusion and is sensitive to the rate of microtubule polymerization and the rate of rearward actin flow as controlled by adhesion-cytoskeletal coupling and MII. We conclude that growth cone navigation is an iterative process of variation and selection. Growth cones extend leading edge actin-rich protrusions that adhere transiently (variation). Microtubule entry up to an adhesion site stabilizes a protrusion (selection) leading to engorgement, consolidation, protrusive activity distal to the adhesion site, and stepwise growth cone advance. The orientation of the protrusion determines the direction of advance.

Adhesion force spectroscopy with nanostructured colloidal probes reveals nanotopography-dependent early mechanotransductive interactions at the cell membrane level

The study shows, by exploiting a novel adhesion force spectroscopy approach, that microenvironmental nanotopography impacts strongly on integrin-mediated cellular mechanosensing, by influencing adhesion site force loading dynamics.

Heavy chain-1 of inter-α-inhibitor has an integrin-like structure with immune regulatory activities

Inter-α-inhibitor (IαI) is a proteoglycan essential for mammalian reproduction that also plays a less well-characterised role in inflammation. IαI is composed of 2 homologous ‘heavy chains’ (HC1 and HC2) covalently attached to chondroitin sulphate on the bikunin core protein. Prior to ovulation HCs are transferred onto the polysaccharide hyaluronan (HA), thereby stabilising a matrix that is required for fertilisation. Here we show that human HC1 has a structure similar to integrin β-chains and contains a functional MIDAS (metal ion-dependent adhesion site) motif that can mediate self-association of heavy chains, providing a mechanism for matrix crosslinking. Surprisingly, its interaction with RGD-containing integrin ligands, such as vitronectin and the latency-associated peptides of TGFβ, occurs in a MIDAS/cation-independent manner. However, HC1 utilises its MIDAS motif to bind to, and inhibit the cleavage of, complement C3, thus identifying it as a novel regulator of innate immunity through inhibition of the alternative pathway C3 convertase.AbbreviationsADPs, atomic displacement parameter; AUC, analytical ultracentrifugation; CMG2, capillary morphogenesis protein-2; COC, cumulus-oocyte complex; CS, chondroitin sulphate; FB, complement factor B; FnIII; fibronectin type III; HA, hyaluronan; HC, heavy chain; HC•HA, covalent complex of HC with HA; IαI, inter-α-inhibitor; ITGA, integrin α-chain; ITGB, integrin β-chain; LAP, latency associated peptide; LLC, large latent complex; LTBP, latent TGFβ binding protein; MIDAS, metal ion-dependent adhesion site; PαI, pre-α-inhibitor; PTX3, pentraxin-3; rHC1, recombinant HC1; SAXS, small-angle X-ray scattering; SHAP, serum-derived HA binding protein; SLC, small latent complex; TEM8, tumour endothelial marker-8; TGFβ, transforming factor β; TSG-6, tumour necrosis factor-stimulated gene-6; TSG-6•HC, covalent complex of TSG-6 and HC; vWFA domain, von Willebrand Factor A domain.