A horizontally transferred cyanase gene in the spider mite Tetranychus urticae is involved in cyanate metabolism and is differentially expressed upon host plant change

The genome of the phytophagous two-spotted spider mite Tetranychus urticae was recently sequenced, representing the first complete chelicerate genome, but also the first genome of a highly polyphagous agricultural pest. Genome analysis revealed the presence of an unexpected high number of cases of putative horizontal gene transfers, including a gene that encodes a cyanase or cyanate lyase. In this study we show by recombinant expression that the T. urticae cyanase remained functionally active after horizontal gene transfer and has a high affinity for cyanate. Cyanases were also detected in other plant parasitic spider mites species such as Tetranychus evansi and Panonychus citri, suggesting that an ancient gene transfer occurred before the diversification within the Tetranychidae family. To investigate the potential role of cyanase in the evolution of plant parasitic spider mites, we studied cyanase expression patterns in T. urticae in relation to host plant range and cyanogenesis, a common plant defense mechanism. Spider mites can alter cyanase expression levels after transfer to several new host plants, including the cyanogenic Phaseolus lunatus. However, the role of cyanase is probably not restricted to cyanide response, but likely to the plant nutritional quality as a whole. We finally discuss potential interactions between cyanase activity and pyrimidine and amino acid synthesis.     

Microdamage evaluation in human trabecular bone based on nonlinear ultrasound vibro-modulation (NUVM)

The primary aim of this work is to investigate the potential of nonlinear ultrasound for microdamage detection in human bone. Microdamage evaluation in human bone is of great importance, because it is considered a significant parameter for characterizing fracture risk. Experiments employing nonlinear acoustic vibro-modulation were carried out in human femoral trabecular specimens removed during surgery. A frequency mixing (inter-modulation) was observed between an ultrasound wave, propagating in the bone, and a low-frequency vibration applied directly to the bone specimens. The appearance of side frequencies, which are related to the vibrational excitation, around the fundamental ultrasound frequency manifests the modulation nonlinear phenomenon. Instead of inducing microdamage by mechanical fatigue loading, specimens with different degree of osteoporosis were used. The experiments demonstrated that osteoporotic bone exhibits stronger nonlinearity compared to healthy bone presenting significant increase of the modulation amplitude with increasing degree of osteoporosis. The obtained results indicate that, in contrast to conventional hysteretic nonlinearity, dissipative acoustic nonlinearity can be of significance in the generation of nonlinear modulation effects. In the proposed technique the size and the shape of samples are not crucial compared to nonlinear resonant ultrasound spectroscopy (NRUS). Furthermore, the method is sensitive to the presence of microdamage, non-invasive, easy to implement and most important, it can be proved valuable tool for in vivo bone damage characterization.     

Hierarchical protein targeting and secretion is controlled by an affinity switch in the type III secretion system of enteropathogenic Escherichia coli

Type III secretion (T3S), a protein export pathway common to Gram‐negative pathogens, comprises a trans‐envelope syringe, the injectisome, with a cytoplasm‐facing translocase channel. Exported substrates are chaperone‐delivered to the translocase, EscV in enteropathogenic Escherichia coli, and cross it in strict hierarchical manner, for example, first “translocators”, then “effectors”. We dissected T3S substrate targeting and hierarchical switching by reconstituting them in vitro using inverted inner membrane vesicles. EscV recruits and conformationally activates the tightly membrane‐associated pseudo‐effector SepL and its chaperone SepD. This renders SepL a high‐affinity receptor for translocator/chaperone pairs, recognizing specific chaperone signals. In a second, SepD‐coupled step, translocators docked on SepL become secreted. During translocator secretion, SepL/SepD suppress effector/chaperone binding to EscV and prevent premature effector secretion. Disengagement of the SepL/SepD switch directs EscV to dedicated effector export. These findings advance molecular understanding of T3S and reveal a novel mechanism for hierarchical trafficking regulation in protein secretion channels.     

Structural instability tuning as a regulatory mechanism in protein-protein interactions

Protein-protein interactions mediate a vast number of cellular processes. Here, we present a regulatory mechanism in protein-protein interactions mediated by finely tuned structural instability and coupled with molecular mimicry. We show that a set of type III secretion (TTS) autoinhibited homodimeric chaperones adopt a molten globule-like state that transiently exposes the substrate binding site as a means to become rapidly poised for binding to their cognate protein substrates. Packing defects at the homodimeric interface stimulate binding, whereas correction of these defects results in less labile chaperones that give rise to nonfunctional biological systems. The protein substrates use structural mimicry to offset the weak spots in the chaperones and to counteract their autoinhibitory conformation. This regulatory mechanism of protein activity is evolutionarily conserved among several TSS systems and presents a lucid example of functional advantage conferred upon a biological system by finely tuned structural instability.     

Erv1 mediates the Mia40-dependent protein import pathway and provides a functional link to the respiratory chain by shuttling electrons to cytochrome c

Unlike matrix-targeted or inner membrane proteins, those that are targeted to the mitochondrial intermembrane space (IMS) do not require ATP or the inner membrane electrochemical potential. Their import is mediated primarily by the essential IMS protein Mia40/Tim40. Here, we show that the mitochondrial flavin adenine dinucleotide (FAD)-linked sulfhydryl oxidase Erv1 (essential for respiration and vegetative growth 1) plays a central role in the biogenesis of small, cysteine proteins of the IMS that are import substrates for Mia40. In a temperature-sensitive strain of Erv1, steady-state levels of small translocases of the inner membrane (Tims) are specifically affected when cells are grown at the non-permissive temperature. Furthermore, mitochondria isolated from the erv1-ts show a specific import and assembly defect for the small Tims but not in any other protein import pathway. Erv1 does not directly oxidise the small Tims, as thiol trapping assays show that the small Tims can still be oxidised in erv1-ts cells grown at the non-permissive temperature and in isolated mitochondria from this strain. Moreover, addition of pure Erv1 into erv1-ts mitochondria lacking the endogenous protein restores import and assembly of the small Tims only to an extent, arguing for a cascade of interactions with Erv1 rather than for a direct interaction of Erv1 with the small Tims. Cytochrome c (cyt c) is the in vivo oxidase for Erv1, as yeast cells mutated in cyt c cannot grow under anaerobic conditions. Therefore, Erv1 functionally links the Mia40-dependent import pathway to the Mia40-independent cyt c import pathway transferring electrons from the incoming precursors to cyt c as an acceptor. In this context, the protein import process is linked to the respiratory chain via the communication of Erv1 with cyt c.