In‐depth protein profiling of the postsynaptic density from mouse hippocampus using data‐independent acquisition proteomics

Located at neuronal terminals, the postsynaptic density (PSD) is a highly complex network of cytoskeletal scaffolding and signaling proteins responsible for the transduction and modulation of glutamatergic signaling between neurons. Using ion‐mobility enhanced data‐independent label‐free LC‐MS/MS, we established a reference proteome of crude synaptosomes, synaptic junctions, and PSD derived from mouse hippocampus including TOP3‐based absolute quantification values for identified proteins. The final dataset across all fractions comprised 49 491 peptides corresponding to 4558 protein groups. Of these, 2102 protein groups were identified in highly purified PSD in at least two biological replicates. Identified proteins play pivotal roles in neurological and synaptic processes providing a rich resource for studies on hippocampal PSD function as well as on the pathogenesis of neuropsychiatric disorders. All MS data have been deposited in the ProteomeXchange with identifier PXD000590 ( http://proteomecentral.proteomexchange.org/dataset/PXD000590 ).     

Leaving uncut refuges during meadow harvesting increases the functional diversity of Orthoptera

In the past, insect species richness was high in Central European seminatural grasslands, which were characterized by low‐intensity land use. Currently, however, the hay in most of these grasslands is mechanically harvested, which negatively impacts insect biodiversity. One way to reduce this negative effect is to leave unmown patches as refuges. In the current research we evaluated the short‐term effects of leaving an unmown patch on the taxonomic and functional diversity of the Orthoptera assemblage in a meadow. We found that orthopteran species richness and abundance were significantly reduced by mowing, whether or not a patch was left uncut. In contrast, functional evenness, indicating distribution of species abundances in a niche space, was reduced by mowing only if the plot lacked an uncut refuge. Functional richness, indicating the amount of niche space occupied by species, was elevated if the plot had an uncut refuge. Larger species were negatively affected by mowing, while habitat specialists, mobile species and soil‐ovipositing species benefitted from it. We infer that the presence of an uncut patch increased the diversity of habitats available to orthopterans and maintained even distribution of species among niche space. In summary, leaving an unmown refuge in grasslands could increase the functional diversity of orthopterans, even if it does not preserve taxonomic diversity.     

Towards a coupled paradigm of NH3‐CO2 biosphere–atmosphere exchange modelling

Stomatal conductance, one of the major plant physiological controls within NH₃ biosphere–atmosphere exchange models, is commonly estimated from semi‐empirical multiplicative schemes or simple light‐ and temperature‐response functions. However, due to their inherent parameterization on meteorological proxy variables, instead of a direct measure of stomatal opening, they are unfit for the use in climate change scenarios and of limited value for interpreting field‐scale measurements. Alternatives based on H₂O flux measurements suffer from uncertainties in the partitioning of evapotranspiration at humid sites, as well as a potential decoupling of transpiration from stomatal opening in the presence of hygroscopic particles on leaf surfaces. We argue that these problems may be avoided by directly deriving stomatal conductance from CO₂ fluxes instead. We reanalysed a data set of NH₃ flux measurements based on CO₂‐derived stomatal conductance, confirming the hypothesis that the increasing relevance of stomatal exchange with the onset of vegetation activity caused a rapid decrease of observed NH₃ deposition velocities. Finally, we argue that developing more mechanistic representations of NH₃ biosphere–atmosphere exchange can be of great benefit in many applications. These range from model‐based flux partitioning, over deposition monitoring using low‐cost samplers and inferential modelling, to a direct response of NH₃ exchange to climate change.     

A proof of principle experiment for microbeam radiation therapy at the Munich compact light source

Microbeam radiation therapy (MRT), a preclinical form of spatially fractionated radiotherapy, uses an array of microbeams of hard synchrotron X-ray radiation. Recently, compact synchrotron X-ray sources got more attention as they provide essential prerequisites for the translation of MRT into clinics while overcoming the limited access to synchrotron facilities. At the Munich compact light source (MuCLS), one of these novel compact X-ray facilities, a proof of principle experiment was conducted applying MRT to a xenograft tumor mouse model. First, subcutaneous tumors derived from the established squamous carcinoma cell line FaDu were irradiated at a conventional X-ray tube using broadbeam geometry to determine a suitable dose range for the tumor growth delay. For irradiations at the MuCLS, FaDu tumors were irradiated with broadbeam and microbeam irradiation at integral doses of either 3 Gy or 5 Gy and tumor growth delay was measured. Microbeams had a width of 50 µm and a center-to-center distance of 350 µm with peak doses of either 21 Gy or 35 Gy. A dose rate of up to 5 Gy/min was delivered to the tumor. Both doses and modalities delayed the tumor growth compared to a sham-irradiated tumor. The irradiated area and microbeam pattern were verified by staining of the DNA double-strand break marker γH2AX. This study demonstrates for the first time that MRT can be successfully performed in vivo at compact inverse Compton sources.     

“Into and Out of” the Qinghai‐Tibet Plateau and the Himalayas: Centers of origin and diversification across five clades of Eurasian montane and alpine passerine birds

Encompassing some of the major hotspots of biodiversity on Earth, large mountain systems have long held the attention of evolutionary biologists. The region of the Qinghai‐Tibet Plateau (QTP) is considered a biogeographic source for multiple colonization events into adjacent areas including the northern Palearctic. The faunal exchange between the QTP and adjacent regions could thus represent a one‐way street (“out of” the QTP). However, immigration into the QTP region has so far received only little attention, despite its potential to shape faunal and floral communities of the QTP. In this study, we investigated centers of origin and dispersal routes between the QTP, its forested margins and adjacent regions for five clades of alpine and montane birds of the passerine superfamily Passeroidea. We performed an ancestral area reconstruction using BioGeoBEARS and inferred a time‐calibrated backbone phylogeny for 279 taxa of Passeroidea. The oldest endemic species of the QTP was dated to the early Miocene (ca. 20 Ma). Several additional QTP endemics evolved in the mid to late Miocene (12–7 Ma). The inferred centers of origin and diversification for some of our target clades matched the “out of Tibet hypothesis’ or the “out of Himalayas hypothesis” for others they matched the “into Tibet hypothesis.” Three radiations included multiple independent Pleistocene colonization events to regions as distant as the Western Palearctic and the Nearctic. We conclude that faunal exchange between the QTP and adjacent regions was bidirectional through time, and the QTP region has thus harbored both centers of diversification and centers of immigration.     

1H-nmr study on the preferred conformations of chiral pyridine-dinucleotide models

The synthesis of four chiral NAD⁺ models 1 and their 1,4-dihydro analogs 2 is described. From the temperature dependence of the ¹H-nmr spectra it is concluded that for these compounds two preferred conformations I and II, differing slightly in energy, exist. Both conformations are “folded” with the more or less parallel p-anisyl and pyridine groups mutually gauche, but in I the pyridine group is rotated by about 180° as compared with II, thus leading to a conspicuous difference in orientation of the substituent Z (NH₂CO, C₆H₅NHSO₂, (CH₂)₄NSO₂, or (C₄H₈ON)SO₂) in the pyridine ring toward the anisyl group. The most stable conformation (I) has Z closest to the center of the p-anisyl group. In 360-MHz spectra of the dihydropyridines at low temperature (?10°C), slow interconversion of I and II leads to the observation of an XY pattern for the C-4 methylene protons of the 1,4-dihydropyridine system. The anisochronity in this methylene group is caused mainly by the anisotropy of the neighboring p-anisyl group.     

Flower formation in the short-day plant Kalanchoë by grafting with a long-day and a short-long-day Echeveria

Flower formation was induced in the shortday plant Kalanchoë blossfeldiana Poellnitz under long-day conditions by grafting with flowering shoots of the short-long-day plant Echeveria harmsii (Rose) MacBr. and of the long-day plant Echeveria pulvoliver (E. pulvinata Rose x E. harmsii). Vegetative shoots from induced Echeveria plants failed to cause a flowering response in Kalanchoë. The presence of flowering and vegetative shoots side by side on induced Echeveria plants provides evidence for physiological chimeras in this genus.Abbreviations LD long day(s) - SD short day(s) - LDP long-day plant(s) - LSDP long-short-day plant(s) - SDP short-day plant(s) - SLDP short-long-day plant(s)     

A Tracer Bolus Method for Investigating Glutamine Kinetics in Humans

Glutamine transport between tissues is important for the outcome of critically ill patients. Investigation of glutamine kinetics is, therefore, necessary to understand glutamine metabolism in these patients in order to improve future intervention studies. Endogenous glutamine production can be measured by continuous infusion of a glutamine tracer, which necessitates a minimum measurement time period. In order to reduce this problem, we used and validated a tracer bolus injection method. Furthermore, this method was used to measure the glutamine production in healthy volunteers in the post-absorptive state, with extra alanine and with glutamine supplementation and parenteral nutrition. Healthy volunteers received a bolus injection of [1-¹³C] glutamine, and blood was collected from the radial artery to measure tracer enrichment over 90 minutes. Endogenous rate of appearance (endoR_a) of glutamine was calculated from the enrichment decay curve and corrected for the extra glutamine supplementation. The glutamine endoR_a of healthy volunteers was 6.1±0.9 µmol/kg/min in the post-absorptive state, 6.9±1.0 µmol/kg/min with extra alanyl-glutamine (p = 0.29 versus control), 6.1±0.4 µmol/kg/min with extra alanine only (p = 0.32 versus control), and 7.5±0.9 µmol/kg/min with extra alanyl-glutamine and parenteral nutrition (p = 0.049 versus control). In conclusion, a tracer bolus injection method to measure glutamine endoR_a showed good reproducibility and small variation at baseline as well as during parenteral nutrition. Additionally, we showed that parenteral nutrition including alanyl-glutamine increased glutamine endoR_a in healthy volunteers, which was not attributable to the alanine part of the dipeptide.     

Light-Harvesting Complex Protein LHCBM9 Is Critical for Photosystem II Activity and Hydrogen Production in Chlamydomonas reinhardtii

Photosynthetic organisms developed multiple strategies for balancing light-harvesting versus intracellular energy utilization to survive ever-changing environmental conditions. The light-harvesting complex (LHC) protein family is of paramount importance for this function and can form light-harvesting pigment protein complexes. In this work, we describe detailed analyses of the photosystem II (PSII) LHC protein LHCBM9 of the microalga Chlamydomonas reinhardtii in terms of expression kinetics, localization, and function. In contrast to most LHC members described before, LHCBM9 expression was determined to be very low during standard cell cultivation but strongly increased as a response to specific stress conditions, e.g., when nutrient availability was limited. LHCBM9 was localized as part of PSII supercomplexes but was not found in association with photosystem I complexes. Knockdown cell lines with 50 to 70% reduced amounts of LHCBM9 showed reduced photosynthetic activity upon illumination and severe perturbation of hydrogen production activity. Functional analysis, performed on isolated PSII supercomplexes and recombinant LHCBM9 proteins, demonstrated that presence of LHCBM9 resulted in faster chlorophyll fluorescence decay and reduced production of singlet oxygen, indicating upgraded photoprotection. We conclude that LHCBM9 has a special role within the family of LHCII proteins and serves an important protective function during stress conditions by promoting efficient light energy dissipation and stabilizing PSII supercomplexes.