Concurrent Imaging of Synaptic Vesicle Recycling and Calcium Dynamics

Synaptic transmission involves the calcium dependent release of neurotransmitter from synaptic vesicles. Genetically encoded optical probes emitting different wavelengths of fluorescent light in response to neuronal activity offer a powerful approach to understand the spatial and temporal relationship of calcium dynamics to the release of neurotransmitter in defined neuronal populations. To simultaneously image synaptic vesicle recycling and changes in cytosolic calcium, we developed a red-shifted reporter of vesicle recycling based on a vesicular glutamate transporter, VGLUT1-mOrange2 (VGLUT1-mOr2), and a presynaptically localized green calcium indicator, synaptophysin-GCaMP3 (SyGCaMP3) with a large dynamic range. The fluorescence of VGLUT1-mOr2 is quenched by the low pH of synaptic vesicles. Exocytosis upon electrical stimulation exposes the luminal mOr2 to the neutral extracellular pH and relieves fluorescence quenching. Reacidification of the vesicle upon endocytosis again reduces fluorescence intensity. Changes in fluorescence intensity thus monitor synaptic vesicle exo- and endocytosis, as demonstrated previously for the green VGLUT1-pHluorin. To monitor changes in calcium, we fused the synaptic vesicle protein synaptophysin to the recently improved calcium indicator GCaMP3. SyGCaMP3 is targeted to presynaptic varicosities, and exhibits changes in fluorescence in response to electrical stimulation consistent with changes in calcium concentration. Using real time imaging of both reporters expressed in the same synapses, we determine the time course of changes in VGLUT1 recycling in relation to changes in presynaptic calcium concentration. Inhibition of P/Q- and N-type calcium channels reduces calcium levels, as well as the rate of synaptic vesicle exocytosis and the fraction of vesicles released

Characterization of reference genes for RT-qPCR in the desert moss Syntrichia caninervis in response to abiotic stress and desiccation/rehydration

Syntrichia caninervis is the dominant bryophyte of the biological soil crusts found in the Gurbantunggut desert. The extreme desert environment is characterized by prolonged drought, temperature extremes, high radiation and frequent cycles of hydration and dehydration. S. caninervis is an ideal organism for the identification and characterization of genes related to abiotic stress tolerance. RT-qPCR expression analysis is a powerful analytical technique that requires the use of stable reference genes. Using available S. caninervis transcriptome data, we selected 15 candidate reference genes and analyzed their relative expression stabilities in S. caninervis gametophores exposed to a range of abiotic stresses or a hydration-desiccation-rehydration cycle. The programs geNorm, NormFinder, and RefFinder were used to assess and rank the expression stability of the 15 candidate genes. The stability ranking results of reference genes under each specific experimental condition showed high consistency using different algorithms. For abiotic stress treatments, the combination of two genes (α-TUB2 and CDPK) were sufficient for accurate normalization. For the hydration-desiccation-rehydration process, the combination of two genes (α-TUB1 and CDPK) were sufficient for accurate normalization. 18S was among the least stable genes in all of the experimental sets and was unsuitable as reference gene in S. caninervis. This is the first systematic investigation and comparison of reference gene selection for RT-qPCR work in S. caninervis. This research will facilitate gene expression studies in S. caninervis, related moss species from the Syntrichia complex and other mosses

Frequency analysis of storm-scale soil erosion and characterization of extreme erosive events by linking the DWEPP model and a stochastic rainfall generator

Soil erosion affects agricultural landscapes worldwide, threatening food security and ecosystem viability. In arable environments, soil loss is primarily caused by short, intense rainstorms, typically characterized by high spatio-temporal variability. The complexity of erosive events challenges modeling efforts and explicit inclusion of extreme events in long-term risk assessment is missing. This study is intended to bridge this gap by quantifying the discrete and cumulative impacts of rainstorms on plot-scale soil erosion and providing storm-scale erosion risk analyses for a cropland region in northern Israel. Central to our analyses is the coupling of (1) a stochastic rainfall generator able to reproduce extremes down to 5-minute temporal resolutions; (2) a processes-based event-scale cropland erosion model (Dynamic WEPP, DWEPP); and, (3) a state-of-the-art frequency analysis method that explicitly accounts for rainstorms occurrence and properties. To our knowledge, this is the first study in which DWEPP runoff and soil loss are calibrated at the plot-scale on cropland (NSE is 0.82 and 0.79 for event runoff and sediment, respectively). We generated 300-year stochastic simulations of event runoff and sediment yield based on synthetic precipitation time series. Based on this data, themean annual soil erosion in the study site is 0.1 kgm(-2) [1.1 t ha(-1)]. Results of the risk analysis indicate that individual extreme rainstorms (>50 return period), characterized by high rainfall intensities (30-minute maximal intensity>similar to 60mmh(-1)) and high rainfall depth (>similar to 200 mm), can trigger soil losses even one order of magnitude higher than the annual mean. The erosion efficiency of these rainstorms is mainly controlled by the short-duration (<= 30 min) maximal intensities. The results demonstrate the importance of incorporating the impact of extreme events into soil conservation and management tools. We expect our methodology to be valuable for investigating future changes in soil erosion with changing climate. (C) 2021 Elsevier B.V. All rights reserved

High-density genetic mapping identifies new major loci for tolerance to low-phosphorus stress in soybean

Phosphorus (P) is essential for all living cells and organisms, and low-P stress represents a major constraint on plant growth and yield worldwide. Soybean is an important economical resource of protein and oil for human and animals, and soybean is also a high-P demand species that is sensitive to low-P stress, which is considered a major constraint on soybean production. However, P efficiency is an important complex quantitative trait involving multiple genes, and the mechanisms underlying soybean P efficiency are largely unknown. Here, we reported the construction of a high-density genetic map using a specific-locus amplified fragment sequencing (SLAF-seq) strategy in soybean. This map, spanning 3020.59 cM in length, contained 6,159 markers on 20 chromosomes, with an average distance of 0.49 cM between adjacent markers. Based on this map, 20 loci, including eight novel loci, associated with P efficiency-related traits were identified across multiple years and treatments. The confidence intervals of almost all QTLs were refined significantly, and the accuracy of this map was evidenced by coincident detections of the previously identified P efficiency-related genes GmACP1 and GmPT1. Notably, a highly significant novel QTL located on chromosome 4, q4-2, was identified across traits, years and treatments. Several candidate genes, such as a pectin methylesterase-encoding gene (Glyma.04G214000) and a protein kinase gene (Glyma.13G161900), with significantly differential expression upon low-P stress were considered as promising candidates involved in regulating soybean P efficiency. Markers that tightly associated with P efficiency could be used for marker-assisted selection in a soybean P efficient breeding program. Further dissection of these QTLs will facilitate gene cloning underlying P efficiency in soybean, and increase our understanding of efficient use of P in enhancing crop yield

Soil biodiversity supports the delivery of multiple ecosystem functions in urban greenspaces

22 páginas.-. 5 figuras.- 64 referencias.- Supplementary Information This pdf file includes: Supplementary Figs. 1–5, Tables 1–11 and references. https://static-content.springer.com/esm/art%3A10.1038%2Fs41559-022-01935-4/MediaObjects/41559_2022_1935_MOESM1_ESM.pdfWhile the contribution of biodiversity to supporting multiple ecosystem functions is well established in natural ecosystems, the relationship of the above- and below-ground diversity with ecosystem multifunctionality remains virtually unknown in urban greenspaces. Here we conducted a standardized survey of urban greenspaces from 56 municipalities across six continents, aiming to investigate the relationships of plant and soil biodiversity (diversity of bacteria, fungi, protists and invertebrates, and metagenomics-based functional diversity) with 18 surrogates of ecosystem functions from nine ecosystem services. We found that soil biodiversity across biomes was significantly and positively correlated with multiple dimensions of ecosystem functions, and contributed to key ecosystem services such as microbially driven carbon pools, organic matter decomposition, plant productivity, nutrient cycling, water regulation, plant–soil mutualism, plant pathogen control and antibiotic resistance regulation. Plant diversity only indirectly influenced multifunctionality in urban greenspaces via changes in soil conditions that were associated with soil biodiversity. These findings were maintained after controlling for climate, spatial context, soil properties, vegetation and management practices. This study provides solid evidence that conserving soil biodiversity in urban greenspaces is key to supporting multiple dimensions of ecosystem functioning, which is critical for the sustainability of urban ecosystems and human wellbeing.This study was supported by a 2019 Leonardo Grant for Researchers and Cultural Creators, BBVA Foundation (URBANFUN), and by the BES grant agreement number LRB17\1019 (MUSGONET). M.D.-B. acknowledges support from the Spanish Ministry of Science and Innovation for the I+D+i project PID2020-115813RA-I00 funded by MCIN/AEI/10.13039/501100011033. M.D.-B. is also supported by a project of the Fondo Europeo de Desarrollo Regional (FEDER) and the Consejería de Transformación Económica, Industria, Conocimiento y Universidades of the Junta de Andalucía (FEDER Andalucía 2014-2020 Objetivo temático ‘01-Refuerzo de la investigación, el desarrollo tecnológico y la innovación’) associated with the research project P20_00879 (ANDABIOMA). H.C. was supported by the Strategic Priority Research Program of Chinese Academy of Sciences (XDA28020202), National Key R&D Program of China (2022YFD1500202) and the National Natural Science Foundation of China (91951109, 42230511, 92251305). K.F. was supported by Young Elite Scientist Sponsorship Program by CAST (2021QNRC001) and China Postdoctoral Science Foundation (2021M703302). F.D.A. and S.A. were supported by ANID FONDECYT 11180538 and 1170995. J.P.V. was supported by SERB (SIR/2022/000626, EEQ/2021/001083), DST (DST/INT/SL/P-31/2021) and Banaras Hindu University, IoE (6031) incentives grant for plant-microbe interaction and soil microbiome research. T.G. and T.U.N were supported by the Slovenian Research Agency grants P4-0107, J4-3098 and J4-4547.Peer reviewe