Intrinsically Disordered Regions in the Transcription Factor MYC:MAX Modulate DNA Binding via Intramolecular Interactions.

The basic helix-loop-helix leucine zipper (bHLH-LZ) transcription factor (TF) MYC is in large part an intrinsically disordered oncoprotein. In complex with its obligate heterodimerization partner MAX, MYC preferentially binds E-Box DNA sequences (CANNTG). At promoters containing these sequence motifs, MYC controls fundamental cellular processes such as cell cycle progression, metabolism, and apoptosis. A vast network of proteins in turn regulates MYC function via intermolecular interactions. In this work, we establish another layer of MYC regulation by intramolecular interactions. We used nuclear magnetic resonance (NMR) spectroscopy to identify and map multiple binding sites for the C-terminal MYC:MAX DNA-binding domain (DBD) on the intrinsically disordered regions (IDRs) in the MYC N-terminus. We find that these binding events in trans are driven by electrostatic attraction, that they have distinct affinities, and that they are competitive with DNA binding. Thereby, we observe the strongest effects for the N-terminal MYC box 0 (Mb0), a conserved motif involved in MYC transactivation and target gene induction. We prepared recombinant full-length MYC:MAX complex and demonstrate that the interactions identified in this work are also relevant in cis, i.e., as intramolecular interactions. These findings are supported by surface plasmon resonance (SPR) experiments, which revealed that intramolecular IDR:DBD interactions in MYC decelerate the association of MYC:MAX complexes to DNA. Our work offers new insights into how bHLH-LZ TFs are regulated by intramolecular interactions, which open up new possibilities for drug discovery.

The Disordered MAX N-terminus Modulates DNA Binding of the Transcription Factor MYC:MAX.

The intrinsically disordered protein MYC belongs to the family of basic helix-loop-helix leucine zipper (bHLH-LZ) transcription factors (TFs). In complex with its cognate binding partner MAX, MYC preferentially binds to E-Box promotor sequences where it controls fundamental cellular processes such as cell cycle progression, metabolism, and apoptosis. Intramolecular regulation of MYC:MAX has not yet been investigated in detail. In this work, we use Nuclear Magnetic Resonance (NMR) spectroscopy to identify and map interactions between the disordered MAX N-terminus and the MYC:MAX DNA binding domain (DBD). We find that this binding event is mainly driven by electrostatic interactions and that it is competitive with DNA binding. Using NMR spectroscopy and Surface Plasmon Resonance (SPR), we demonstrate that the MAX N-terminus serves to accelerate DNA binding kinetics of MYC:MAX and MAX:MAX dimers, while it simultaneously provides specificity for E-Box DNA. We also establish that these effects are further enhanced by Casein Kinase 2-mediated phosphorylation of two serine residues in the MAX N-terminus. Our work provides new insights how bHLH-LZ TFs are regulated by intramolecular interactions between disordered regions and the folded DNA binding domain.

Unveiling Chemical Cues of Insect-Tree and Insect-Insect Interactions for the Eucalyptus Weevil and Its Egg Parasitoid by Multidimensional Gas Chromatographic Methods.

Multidimensional gas chromatography is, presently, an established and powerful analytical tool, due to higher resolving power than the classical 1D chromatographic approaches. Applied to multiple areas, it allows to isolate, detect and identify a larger number of compounds present in complex matrices, even in trace amounts. Research was conducted to determine which compounds, emitted by host plants of the eucalyptus weevil, Gonipterus platensis, might mediate host selection behavior. The identification of a pheromone blend of G. platensis is presented, revealing to be more attractive to weevils of both sexes, than the individual compounds. The volatile organic compounds (VOCs) were collected by headspace solid phase microextraction (HS-SPME), MonoTrapTM disks, and simultaneous distillation-extraction (SDE). Combining one dimensional (1D) and two-dimensional (2D) chromatographic systems-comprehensive and heart-cut two-dimensional gas chromatography (GC×GC and H/C-MD-GC, respectively) with mass spectrometry (MS) and electroantennographic (EAD) detection, enabled the selection and identification of pertinent semiochemicals which were detected by the insect antennal olfactory system. The behavioral effect of a selected blend of compounds was assessed in a two-arm olfactometer with ten parallel walking chambers, coupled to video tracking and data analysis software. An active blend, composed by cis and trans-verbenol, verbenene, myrtenol and trans-pinocarveol was achieved.

Collective Dissipative Molecule Formation in a Cavity.

We propose a mechanism to realize high-yield molecular formation from ultracold atoms. Atom pairs are continuously excited by a laser, and a collective decay into the molecular ground state is induced by a coupling to a lossy cavity mode. Using a combination of analytical and numerical techniques, we demonstrate that the molecular yield can be improved by simply increasing the number of atoms, and can overcome efficiencies of state-of-the-art association schemes. We discuss realistic experimental setups for diatomic polar and nonpolar molecules, opening up collective light matter interactions as a tool for quantum state engineering, enhanced molecule formation, collective dynamics, and cavity mediated chemistry.

Methyl TROSY spectroscopy: A versatile NMR approach to study challenging biological systems.

A major goal in structural biology is to unravel how molecular machines function in detail. To that end, solution-state NMR spectroscopy is ideally suited as it is able to study biological assemblies in a near natural environment. Based on methyl TROSY methods, it is now possible to record high-quality data on complexes that are far over 100 kDa in molecular weight. In this review, we discuss the theoretical background of methyl TROSY spectroscopy, the information that can be extracted from methyl TROSY spectra and approaches that can be used to assign methyl resonances in large complexes. In addition, we touch upon insights that have been obtained for a number of challenging biological systems, including the 20S proteasome, the RNA exosome, molecular chaperones and G-protein-coupled receptors. We anticipate that methyl TROSY methods will be increasingly important in modern structural biology approaches, where information regarding static structures is complemented with insights into conformational changes and dynamic intermolecular interactions.

Atomic-level insight into mRNA processing bodies by combining solid and solution-state NMR spectroscopy.

Liquid-liquid phase separation is increasingly recognized as a process involved in cellular organization. Thus far, a detailed structural characterization of this intrinsically heterogeneous process has been challenging. Here we combine solid- and solution-state NMR spectroscopy to obtain atomic-level insights into the assembly and maturation of cytoplasmic processing bodies that contain mRNA as well as enzymes involved in mRNA degradation. In detail, we have studied the enhancer of decapping 3 (Edc3) protein that is a central hub for processing body formation in yeast. Our results reveal that Edc3 domains exhibit diverse levels of structural organization and dynamics after liquid-liquid phase separation. In addition, we find that interactions between the different Edc3 domains and between Edc3 and RNA in solution are largely preserved in the condensed protein state, allowing processing bodies to rapidly form and dissociate upon small alterations in the cellular environment.

EAG response and behavioral orientation of Dastarcus helophoroides (Fairmaire) (Coleoptera: Bothrideridae) to synthetic host-associated volatiles.

Dastarcus helophoroides Fairmaire (Coleoptera: Bothrideridae) is an effective predatory beetle of larvae and pupae of several cerambycid beetles including Monochamus alternatus and Anoplophora glabripennis. Electroantennography (EAG) and a dynamic two-choice olfactometer were respectively used to measure the antennal and behavioral responses of both sexes to selected volatile compounds. Female and male D. helophoroides exhibited similar EAG and behavioral responses. Significant dose-dependent EAG responses in both sexes were elicited by nonanal, octanal, cis-3-hexenol, 3-carene, (R)-(+)-α-pinene, (S)-(-)-α-pinene, (R)-(+)-limonene and (S)-(-)-limonene. Female and male beetles were repelled at high concentration by cis-3-hexenol and (S)-(-)-limonene, respectively. Both sexes of D. helophoroides were significantly attracted to nonanal, cis-3-hexenol, 3-carene and (R)-(+)-limonene even at low concentrations. These compounds might be used either individually or in mixtures for developing biological control methods to attract this predatory beetle into forest stands threatened by cerambycid beetles.

Cavity-Enhanced Transport of Charge.

We theoretically investigate charge transport through electronic bands of a mesoscopic one-dimensional system, where interband transitions are coupled to a confined cavity mode, initially prepared close to its vacuum. This coupling leads to light-matter hybridization where the dressed fermionic bands interact via absorption and emission of dressed cavity photons. Using a self-consistent nonequilibrium Green's function method, we compute electronic transmissions and cavity photon spectra and demonstrate how light-matter coupling can lead to an enhancement of charge conductivity in the steady state. We find that depending on cavity loss rate, electronic bandwidth, and coupling strength, the dynamics involves either an individual or a collective response of Bloch states, and we explain how this affects the current enhancement. We show that the charge conductivity enhancement can reach orders of magnitudes under experimentally relevant conditions.

[Psychiatric Emergencies in the Preclinical Emergency Medicine Service in Ulm, Germany in 2000 and 2010, and Practical Consequences]. / Psychiatrische Notfälle im Notarzt- und Rettungsdienst in Ulm in den Jahren 2000 und 2010 und Konsequenzen für die Praxis.

Background Psychiatric emergencies (PE) in preclinical emergency medical services are about 5 - 10 % of all emergencies and represent often a source of difficulties in handling for the non-psychiatric professional helpers that deal with them. Studies informing about quantitative and qualitative changes of PEs in preclinical emergency medicine in Germany are scarce. Methods Therefore, we conducted a retrospective cross-sectional study of PE in a preclinical emergency medical service based on the protocols of the emergency ambulance of the Section for Emergency Medicine at the University Hospital Ulm comparing the years 2000 and 2010. Results We observed a significant increase of PEs from 8.8 % in the year 2000 (n = 285, from a total of n = 3227) to 10.3 % in 2010 (n = 454, from a total of n = 4425). In both years intoxications were the most common PE [2000: n = 116 (44.4 %); 2010: n = 171 (37.7 %)], followed by suicide-related behavior [2000: n = 59 (22.6 %); 2010: n = 78 (17.2 %)] and acute anxiety disorders [2000: n = 37 (13 %); 2010: n = 105 (23.1 %)]. The mentioned three conditions accounted for about 80 % of all PE. Most frequently PE occurred at the weekend and with the highest density in the evening and at night (18 - 24 h) in both years. Patients with PE were predominantly men, but the rate of women causing PE increased between 2000 and 2010. Discussion/Conclusion This study provides preliminary data on current trends in PEs in preclinical emergency medicine in Germany and has implications for improving the medical care provided.

A synergistic network of interactions promotes the formation of in vitro processing bodies and protects mRNA against decapping.

Cellular liquid-liquid phase separation (LLPS) results in the formation of dynamic granules that play an important role in many biological processes. On a molecular level, the clustering of proteins into a confined space results from an indefinite network of intermolecular interactions. Here, we introduce and exploit a novel high-throughput bottom-up approach to study how the interactions between RNA, the Dcp1:Dcp2 mRNA decapping complex and the scaffolding proteins Edc3 and Pdc1 result in the formation of processing bodies. We find that the LLPS boundaries are close to physiological concentrations upon inclusion of multiple proteins and RNA. Within in vitro processing bodies the RNA is protected against endonucleolytic cleavage and the mRNA decapping activity is reduced, which argues for a role of processing bodies in temporary mRNA storage. Interestingly, the intrinsically disordered region (IDR) in the Edc3 protein emerges as a central hub for interactions with both RNA and mRNA decapping factors. In addition, the Edc3 IDR plays a role in the formation of irreversible protein aggregates that are potentially detrimental for cellular homeostasis. In summary, our data reveal insights into the mechanisms that lead to cellular LLPS and into the way this influences enzymatic activity.

The Rrp4-exosome complex recruits and channels substrate RNA by a unique mechanism.

The exosome is a large molecular machine involved in RNA degradation and processing. Here we address how the trimeric Rrp4 cap enhances the activity of the archaeal enzyme complex. Using methyl-TROSY NMR methods we identified a 50-Å long RNA binding path on each Rrp4 protomer. We show that the Rrp4 cap can thus simultaneously recruit three substrates, one of which is degraded in the core while the others are positioned for subsequent degradation rounds. The local interaction energy between the substrate and the Rrp4-exosome increases from the periphery of the complex toward the active sites. Notably, the intrinsic interaction strength between the cap and the substrate is weakened as soon as substrates enter the catalytic barrel, which provides a means to reduce friction during substrate movements toward the active sites. Our data thus reveal a sophisticated exosome-substrate interaction mechanism that enables efficient RNA degradation.

Emission of Volatile Compounds from Apple Plants Infested with Pandemis heparana Larvae, Antennal Response of Conspecific Adults, and Preliminary Field Trial.

This study investigated the volatile emission from apple (Malus x domestica Borkh., cv. Golden Delicious) foliage that was either intact, mechanically-damaged, or exposed to larval feeding by Pandemis heparana (Denis and Schiffermüller) (Lepidoptera: Tortricidae). Volatiles were collected by closed-loop-stripping-analysis and characterized by gas chromatography-mass spectrometry in three time periods: after 1 h and again 24 and 48 h later. Volatiles for all treatments also were monitored continuously over a 72-h period by the use of proton transfer reaction - time of flight-mass spectrometry (PTR-ToF-MS). In addition, the volatile samples were analyzed by gas chromatography-electroantennographic detection (GC-EAD) using male and female antennae of P. heparana. Twelve compounds were detected from intact foliage compared with 23 from mechanically-damaged, and 30 from P. heparana-infested foliage. Interestingly, six compounds were released only by P. heparana-infested foliage. The emission dynamics of many compounds measured by PTR-ToF-MS showed striking differences according to the timing of herbivory and the circadian cycle. For example, the emission of green leaf volatiles began shortly after the start of herbivory, and increased over time independently from the light-dark cycle. Conversely, the emission of terpenes and aromatic compounds showed a several-hour delay in response to herbivory, and followed a diurnal rhythm. Methanol was the only identified volatile showing a nocturnal rhythm. Consistent GC-EAD responses were found for sixteen compounds, including five aromatic ones. A field trial in Sweden demonstrated that benzyl alcohol, 2-phenylethanol, phenylacetonitrile, and indole lures placed in traps were not attractive to Pandemis spp. adults, but 2-phenylethanol and phenylacetonitrile when used in combination with acetic acid were attractive to both sexes.

Dissipation-Assisted Prethermalization in Long-Range Interacting Atomic Ensembles.

We theoretically characterize the semiclassical dynamics of an ensemble of atoms after a sudden quench across a driven-dissipative second-order phase transition. The atoms are driven by a laser and interact via conservative and dissipative long-range forces mediated by the photons of a single-mode cavity. These forces can cool the motion and, above a threshold value of the laser intensity, induce spatial ordering. We show that the relaxation dynamics following the quench exhibits a long prethermalizing behavior which is first dominated by coherent long-range forces and then by their interplay with dissipation. Remarkably, dissipation-assisted prethermalization is orders of magnitude longer than prethermalization due to the coherent dynamics. We show that it is associated with the creation of momentum-position correlations, which remain nonzero for even longer times than mean-field predicts. This implies that cavity cooling of an atomic ensemble into the self-organized phase can require longer time scales than the typical experimental duration. In general, these results demonstrate that noise and dissipation can substantially slow down the onset of thermalization in long-range interacting many-body systems.

A Model for Carrier-Mediated Biological Signal Transduction Based on Equilibrium Ligand Binding Theory.

Different variants of a mathematical model for carrier-mediated signal transduction are introduced with focus on the odor dose-electrophysiological response curve of insect olfaction. The latter offers a unique opportunity to observe experimentally the effect of an alteration in the carrier molecule composition on the signal molecule-dependent response curve. Our work highlights the role of involved carrier molecules, which have largely been ignored in mathematical models for response curves in the past. The resulting model explains how the involvement of more than one carrier molecule in signal molecule transport can cause dose-response curves as observed in experiments, without the need of more than one receptor per neuron. In particular, the model has the following features: (1) An extended sensitivity range of neuronal response is implemented by a system consisting of only one receptor but several carrier molecules with different affinities for the signal molecule. (2) Given that the sensitivity range is extended by the involvement of different carrier molecules, the model implies that a strong difference in the expression levels of the carrier molecules is absolutely essential for wide range responses. (3) Complex changes in dose-response curves which can be observed when the expression levels of carrier molecules are altered experimentally can be explained by interactions between different carrier molecules. The principles we demonstrate here for electrophysiological responses can also be applied to any other carrier-mediated biological signal transduction process. The presented concept provides a framework for modeling and statistical analysis of signal transduction processes if sufficient information on the underlying biology is available.

Observation and Quantification of Mating Behavior in the Pinewood Nematode, Bursaphelenchus xylophilus.

A method for observing and quantifying the mating behavior of the pinewood nematode, Bursaphelenchus xylophilus, was established under a stereomicroscope. To improve the mating efficiency of B. Xylophilus and to increase the chances of mating observation, virgin adults were cultured and used for the investigation. Eggs were obtained by keeping the nematodes in water and allowing the females to lay eggs for 10 min. The second-stage juveniles (J2) were synchronized by incubating the eggs for 24 h at 25 °C in the dark, and the early J4 were obtained by culturing the J2 with grey mold, Botrytis cinerea, for another 52 h. At this time point, most J4 nematodes could be clearly distinguished as being male or female using their genital morphology. The male and female J4 were collected and cultured separately in two different Petri dishes for 24 h to get virgin adult nematodes. A virgin male and a virgin female were paired in a drop of water in the well of a concave slide. The mating behavior was filmed with a video recorder under a stereomicroscope. The whole period of the mating process was 82.8 ±3.91 min (mean ±SE) and could be divided into 4 different phases: searching, contacting, copulating, and lingering. The mean minutes of duration were 21.8 ± 2.0, 28.0 ± 1.9, 25.8 ± 0.7 and 7.2 ± 0.5, respectively. Eleven sub-behaviors were described: cruising, approaching, encountering, touching, hooping, locating, attaching, ejaculating, separating, quiescence, and roaming. Interestingly, obvious intra-sexual competition was observed when one female was grouped with 3 males or one male with 3 females. This protocol is useful and valuable, not only in investigating the mating behavior of B. xylophilus, but also in acting as a reference for ethological studies of other nematodes.

Electronic Olfactory Sensor Based on A.†mellifera Odorant-Binding Protein†14 on a Reduced Graphene Oxide Field-Effect Transistor.

An olfactory biosensor based on a reduced graphene oxide (rGO) field-effect transistor (FET), functionalized by the odorant-binding protein 14 (OBP14) from the honey bee (Apis mellifera) has been designed for the in situ and real-time monitoring of a broad spectrum of odorants in aqueous solutions known to be attractants for bees. The electrical measurements of the binding of all tested odorants are shown to follow the Langmuir model for ligand-receptor interactions. The results demonstrate that OBP14 is able to bind odorants even after immobilization on rGO and can discriminate between ligands binding within a range of dissociation constants from K(d)=4 µM to K(d)=3.3 mM. The strongest ligands, such as homovanillic acid, eugenol, and methyl vanillate all contain a hydroxy group which is apparently important for the strong interaction with the protein.

Bortezomib-containing regimens are effective in multiple myeloma--results of a non-interventional phase IV study.

BACKGROUND/AIMS: The incorporation of bortezomib into the chemotherapeutic regimens for non-transplant patients with multiple myeloma resulted in improved outcomes in controlled studies. This prospective, non-interventional study assessed the effectiveness and safety of bortezomib-containing regimens in daily practice. METHODS: Patients with untreated or relapsed multiple myeloma not eligible for high-dose chemotherapy followed by autologous stem cell transplantation and who were scheduled for bortezomib mono- or combination therapy or melphalan-prednisone (MP) alone were included in this study. Dosage and treatment decisions were at the discretion of the physicians. RESULTS: 353 patients received bortezomib-containing therapies and 37 patients MP alone. Overall response rates at treatment end were 65.9% for bortezomib-containing therapies and 50.0% for MP. Partial or complete remissions considered best responses were achieved in 82.6% (first line) and 63.8% (second or later line) of the bortezomib-treated patients. The median duration of response to bortezomib-containing therapies was 18.2 months in 109 first-line and 11.3 months in 110 second- or later-line patients. Adverse drug reactions of any grade were reported during the treatment phase in 79.6% (bortezomib) and 70.3% (MP) of treated patients. CONCLUSION: Bortezomib-containing therapies were effective in patients with multiple myeloma in a real-life setting. The increasingly individualized treatment regimens of multiple myeloma require standardized assessments of response in daily practice.

Volatile emission of decomposing pig carcasses (Sus scrofa domesticus L.) as an indicator for the postmortem interval.

This study aimed at correlating selected carcass borne volatile organic compounds (VOCs) with the postmortem interval (PMI). Selected volatiles should 1st be reliably emitted during vertebrate decay, 2nd be emitted at high concentrations, and 3rd show a reproducible quantitative dynamic during the decaying process. Four pigs (Sus scrofa domesticus L.) were placed in a deciduous forest in different seasons and volatiles emitted during the decaying process were sampled. Seventeen compounds were identified and quantified by GC-MS. Electrophysiological experiments on the antenna of female Calliphora vicina and additional data of Dermestes maculans were used as an evolutionary tuned information filter to evaluate the 1st criterion. The relative quantitative emission of hexanal, nonanal, dimethyl disulfide, dimethyl trisulfide, 1-butanol, and phenol were correlated with the PMI, and the observed stages of decay and the limitations of this model were discussed.

Tissue-specific transcriptomics, chromosomal localization, and phylogeny of chemosensory and odorant binding proteins from the red flour beetle Tribolium castaneum reveal subgroup specificities for olfaction or more general functions.

BACKGROUND: Chemoreception is based on the senses of smell and taste that are crucial for animals to find new food sources, shelter, and mates. The initial step in olfaction involves the translocation of odorants from the periphery through the aqueous lymph of the olfactory sensilla to the odorant receptors most likely by chemosensory proteins (CSPs) or odorant binding proteins (OBPs). RESULTS: To better understand the roles of CSPs and OBPs in a coleopteran pest species, the red flour beetle Tribolium castaneum (Coleoptera, Tenebrionidae), we performed transcriptome analyses of male and female antennae, heads, mouthparts, legs, and bodies, which revealed that all 20 CSPs and 49 of the 50 previously annotated OBPs are transcribed. Only six of the 20 CSP are significantly transcriptionally enriched in the main chemosensory tissues (antenna and/or mouthparts), whereas of the OBPs all eight members of the antenna binding proteins II (ABPII) subgroup, 18 of the 20 classic OBP subgroup, the C + OBP, and only five of the 21 C-OBPs show increased chemosensory tissue expression. By MALDI-TOF-TOF MS protein fingerprinting, we confirmed three CSPs, four ABPIIs, three classic OBPs, and four C-OBPs in the antennae. CONCLUSIONS: Most of the classic OBPs and all ABPIIs are likely involved in chemoreception. A few are also present in other tissues such as odoriferous glands and testes and may be involved in release or transfer of chemical signals. The majority of the CSPs as well as the C-OBPs are not enriched in antennae or mouthparts, suggesting a more general role in the transport of hydrophobic molecules.

Prethermalization of atoms due to photon-mediated long-range interactions.

Atoms can spontaneously form spatially ordered structures in optical resonators when they are transversally driven by lasers. This occurs when the laser intensity exceeds a threshold value and results from the mechanical forces on the atoms associated with superradiant scattering into the cavity mode. We treat the atomic motion semiclassically and show that, while the onset of spatial ordering depends on the intracavity-photon number, the stationary momentum distribution is a Gaussian function whose width is determined by the rate of photon losses. Above threshold, the dynamics is characterized by two time scales: after a violent relaxation, the system slowly reaches the stationary state over time scales exceeding the cavity lifetime by several orders of magnitude. In this transient regime the atomic momenta form non-Gaussian metastable distributions, which emerge from the interplay between the long-range dispersive and dissipative mechanical forces of light. We argue that the dynamics of self-organization of atoms in cavities offers a test bed for studying the statistical mechanics of long-range interacting systems.