bluuebunny/crossref_metadata_2025 huggingface dataset:
Photosynthetic light harvesting and thylakoid organization in a CRISPR/Cas9 Arabidopsis thaliana LHCB1 knockout mutant
Authors: Sattari Vayghan Hamed , Nawrocki Wojciech J. , Schiphorst Christo , Tolleter Dimitri , Hu Chen , Douet Veronique , Glauser Gaëtan , Giovanni Finazzi , Croce Roberta , Wientjes Emilie , Longoni Fiamma
Published: December 2021

Light absorbed by chlorophylls of photosystem II and I drives oxygenic photosynthesis. Light-harvesting complexes increase the absorption cross-section of these photosystems. Furthermore, these complexes play a central role in photoprotection by dissipating the excess of absorbed light energy in an inducible and regulated fashion. In higher plants, the main light-harvesting complex is the trimeric LHCII.

In this work, we used CRISPR/Cas9 to knockout the five genes encoding LHCB1, which is the major component of the trimeric LHCII. In absence of LHCB1 the accumulation of the other LHCII isoforms was only slightly increased, thereby resulting in chlorophyll loss leading to a pale green phenotype and growth delay. Photosystem II absorption cross-section was smaller while photosystem I absorption cross-section was unaffected. This altered the chlorophyll repartition between the two photosystems, favoring photosystem I excitation. The equilibrium of the photosynthetic electron transport was partially maintained by a lower photosystem I over photosystem II reaction center ratio and by the dephosphorylation of LHCII and photosystem II. Loss of LHCB1 altered the thylakoid structure, with less membrane layers per grana stack and reduced grana width. Stable LHCB1 knock out lines allow characterizing the role of this protein in light harvesting and acclimation and pave the way for future in vivo mutational analyses of LHCII.
Restricted proliferation during neurogenesis contributes to regionalization of the amphioxus nervous system
Authors: Gattoni Giacomo , Andrews Toby GR , Benito-Gutiérrez Elia
Published: December 2021

The central nervous system of the cephalochordate amphioxus consists of a dorsal neural tube with an anterior brain. Two decades of gene expression analyses in developing amphioxus embryos have shown that despite the lack of overt segmentation the amphioxus neural tube is highly regionalized at the molecular level. However, little is known about the mechanisms that generate such precise regionalization. Proliferation is a key driver of pattern formation and cell type diversification, but in amphioxus it has never been studied in detail nor in the specific context of neurogenesis. Here, we describe the dynamics of cell division during the formation of the central nervous system in amphioxus embryos and its contributions to the regionalization of the neural axis. We show that after gastrulation, proliferation pauses to become spatially restricted to the anterior and posterior ends of the neural tube at neurula stages. Only at the onset of larval life, proliferation resumes in the central part of the nervous system. By marking specific populations and inhibiting cell division during neurulation, we demonstrate that proliferation in the anterior cerebral vesicle is required to establish the full cell type repertoire of the frontal eye complex and the putative hypothalamic region of the amphioxus brain, while posterior proliferating progenitors, which were found here to derive from the dorsal lip of the blastopore, contribute to elongate the caudal floor plate. Between these proliferative domains, we find trunk nervous system differentiation is independent from cell division, which decreases during neurulation and resumes at the early larval stage. Taken together, our results highlight multiple roles for proliferation in shaping the amphioxus nervous system.
Distinct patterns of genetic variation at low-recombining genomic regions represent haplotype structure
Authors: Ishigohoka Jun , Bascón-Cardozo Karen , Bours Andrea , Fuß Janina , Rhie Arang , Mountcastle Jacquelyn , Haase Bettina , Chow William , Collins Joanna , Howe Kerstin , Uliano-Silva Marcela , Fedrigo Olivier , Jarvis Erich D. , Pérez-Tris Javier , Illera Juan Carlos , Liedvogel Miriam
Published: December 2021

Genetic variation of the entire genome represents population structure, yet individual loci can show distinct patterns. Such deviations identified through genome scans have often been attributed to effects of selection instead of randomness. This interpretation assumes that long enough genomic intervals average out randomness in underlying genealogies, which represent local genetic ancestries. However, an alternative explanation to distinct patterns has not been fully addressed: too few genealogies to average out the effect of randomness. Specifically, distinct patterns of genetic variation may be due to reduced local recombination rate, which reduces the number of genealogies in a genomic window. Here, we associate distinct patterns of local genetic variation with reduced recombination rates in a songbird, the Eurasian blackcap (Sylvia atricapilla), using genome sequences and recombination maps. We find that distinct patterns of local genetic variation reflect haplotype structure at low-recombining regions either shared in most populations or found only in a few populations. At the former species-wide low-recombining regions, genetic variation depicts conspicuous haplotypes segregating in multiple populations. At the latter population-specific low-recombining regions, genetic variation represents variance among cryptic haplotypes within the low-recombining populations. With simulations, we confirm that these distinct patterns of haplotype structure evolve due to reduced recombination rate, on which the effects of selection can be overlaid. Our results highlight that distinct patterns of genetic variation can emerge through evolution of reduced local recombination rate. Recombination landscape as an evolvable trait therefore plays an important role determining the heterogeneous distribution of genetic variation along the genome.
A Single-copy Knock In Translating Ribosome ImmunoPrecipitation (SKI TRIP) tool kit for tissue specific profiling of actively translated mRNAs in C. elegans
Authors: Wester Laura E. , Lanjuin Anne , Bruckisch Emanuel H. W. , Perez Matos Maria C. , Heintz Caroline , Denzel Martin S. , Mair William B.
Published: December 2021

Translating Ribosome Affinity Purification (TRAP) methods have emerged as a powerful approach to profile actively translated transcripts in specific cell and tissue types. Epitope tagged ribosomal subunits are expressed in defined cell populations and used to pull down ribosomes and their associated mRNAs, providing a snapshot of cell type-specific translation occurring in that space and time. Current TRAP toolkits available to the C. elegans community have been built using multi-copy arrays, randomly integrated in the genome. Here we introduce a Single-copy Knock In Translating Ribosome ImmunoPrecipitation (SKI TRIP) tool kit, a collection of C. elegans strains engineered by CRISPR in which tissue specific expression of FLAG tagged ribosomal subunit protein RPL-22 is driven by cassettes present in single copy from defined sites in the genome. In depth characterization of the SKI TRIP strains and methodology shows that 3xFLAG tagged RPL-22 expressed from its endogenous locus or within defined cell types incorporates into actively translating ribosomes and can be used to efficiently and cleanly pull-down cell type specific transcripts without impacting overall mRNA translation or fitness of the animal. We propose SKI TRIP use for the study of processes that are acutely sensitive to changes in translation, such as aging.
ERK2 MAP kinase regulates SUFU binding by multisite phosphorylation of GLI1
Authors: Bardwell A. Jane , Wu Beibei , Sarin Kavita Y. , Waterman Marian L. , Atwood Scott X. , Bardwell Lee
Published: December 2021

There is considerable evidence that crosstalk between the Hedgehog and MAPK signaling pathways occurs in several types of cancer, and contributes to the emergence of clinical resistance to Hedgehog pathway inhibitors. Here we show that MAP kinase-mediated phosphorylation weakens the binding of the GLI1 transcription factor to its negative regulator SUFU. ERK2 phosphorylates GLI1 on three evolutionarily-conserved target sites (S102, S116 and S130) located near the high-affinity binding site for SUFU; these phosphorylations cooperate to weaken the affinity of GLI1-SUFU binding by over 25 fold. Phosphorylation of any one, or even any two, of the three sites does not result in the level of SUFU release seen when all three sites are phosphorylated. Tumor-derived mutations in R100 and S105, residues bordering S102, also diminish SUFU binding, collectively defining a novel evolutionarily-conserved SUFU-affinity-modulating region. In cultured mammalian cells, GLI1 variants containing phosphomimetic substitutions of S102, S116 and S130 displayed an increased ability to drive transcription. We conclude that of multisite phosphorylation of GLI1 by ERK2 or other MAP kinases weakens GLI1-SUFU binding, thereby facilitating GLI1 activation and contributing to both physiological and pathological crosstalk.
Single-cell RNA sequencing reveals a role for reactive oxygen species and peroxiredoxins in fatty-acid-induced rat ß-cell proliferation
Authors: Vivoli Alexis , Ghislain Julien , Filali-Mouhim Ali , Angeles Zuraya Elisa , Castell Anne-Laure , Sladek Robert , Poitout Vincent
Published: December 2021

The functional mass of insulin-secreting pancreatic β cells expands to maintain glucose homeostasis in the face of nutrient excess, in part via replication of existing β cells. To decipher the underlying molecular mechanisms, we assessed β-cell proliferation in isolated rat islets exposed to glucose and oleate or palmitate for 48 h and analyzed the transcriptional response by single-cell RNA sequencing. Unsupervised clustering of pooled β cells identified subpopulations, including proliferating β cells. β-cell proliferation increased in response to oleate but not palmitate. Both fatty acids enhanced the expression of genes involved energy metabolism and mitochondrial activity. Comparison of proliferating vs. non-proliferating β cells and pseudotime ordering suggested the involvement of reactive oxygen species (ROS) and peroxiredoxin signaling. Accordingly, the antioxidant N-acetyl cysteine and the peroxiredoxin inhibitor Conoidin A both blocked oleate-induced β-cell proliferation. Our data reveal a key role for ROS signaling in β-cell proliferation in response to nutrients.
Structure of anchorless RML prion reveals motif variation between strains
Authors: Hoyt Forrest , Standke Heidi G. , Artikis Efrosini , Schwartz Cindi L. , Hansen Bryan , Li Kunpeng , Hughson Andrew G. , Manca Matteo , Thomas Olivia R. , Raymond Gregory J. , Baron Gerald S. , Caughey Byron , Kraus Allison
Published: December 2021

Little is known about the structural basis of prion strains. Here we provide a high (3.0 Å) resolution cryo-electron microscopy-based structure of brain-derived fibrils of the mouse anchorless RML scrapie strain which, like the recently determined hamster 263K strain, has a parallel in-register β-sheet-based core. However, detailed comparisons reveal that variations in shared structural motifs provide a basis for prion strain determination.

One-sentence summary

Cryo-electron microscopy reveals a near-atomic structure of an infectious, brain-derived murine prion fibril and strain differences.
Biophysical and pharmacokinetic characterization of a small-molecule inhibitor of RUNX1/ETO tetramerization with anti-leukemic effects
Authors: Gopalswamy Mohanraj , Kroeger Tobias , Bickel David , Frieg Benedikt , Akter Shahina , Schott-Verdugo Stephan , Viegas Aldino , Pauly Thomas , Mayer Manuela , Przibilla Julia , Reiners Jens , Nagel-Steger Luitgard , Smits Sander H.J. , Groth Georg , Etzkorn Manuel , Gohlke Holger
Published: December 2021

Acute myeloid leukemia (AML) is a malignant disease of immature myeloid cells and the most prevalent acute leukemia among adults. The oncogenic homo-tetrameric fusion protein RUNX1/ETO results from the chromosomal translocation t(8;21) and is found in AML patients. The nervy homology region 2 (NHR2) domain of ETO mediates tetramerization; this oligomerization is essential for oncogenic activity. Previously, we identified the first-in-class small-molecule inhibitor of NHR2 tetramer formation, 7.44, which was shown to specifically interfere with NHR2, restore gene expression down-regulated by RUNX1/ETO, inhibit the proliferation of RUNX1/ETO-depending SKNO-1 cells, and reduce the RUNX1/ETO-related tumor growth in a mouse model. However, no biophysical and structural characterization of 7.44 binding to the NHR2 domain has been reported. Likewise, the compound has not been characterized as to physicochemical, pharmacokinetic, and toxicological properties. Here, we characterize the interaction between the NHR2 domain of RUNX1/ETO and 7.44 by biophysical assays and show that 7.44 interferes with NHR2 tetramer stability and leads to an increase in the dimer population of NHR2. The affinity of 7.44 with respect to binding to NHR2 is Klig = 3.95 ± 1.28 μM. By NMR spectroscopy combined with molecular dynamics simulations, we show that 7.44 binds with both heteroaromatic moieties to NHR2 and interacts with or leads to conformational changes in the N-termini of the NHR2 tetramer. Finally, we demonstrate that 7.44 has favorable physicochemical, pharmacokinetic, and toxicological properties. Together with biochemical, cellular, and in vivo assessments, the results reveal 7.44 as a lead for further optimization towards targeted therapy of t(8;21) AML.
Effects of α-crystallin gene knockout on zebrafish lens development
Authors: Posner Mason , Murray Kelly L. , Andrew Brandon , Brdicka Stuart , Roberts Alexis , Franklin Kirstan , Hussen Adil , Kaye Taylor , Kepp Emmaline , McDonald Mathew S. , Snodgrass Tyler , Zientek Keith , David Larry L.
Published: December 2021

The α-crystallin small heat shock proteins contribute to the transparency and refractive properties of the vertebrate eye lens and prevent the protein aggregation that would otherwise produce lens cataracts, the leading cause of human blindness. There are conflicting data in the literature as to what role the α-crystallins may play in early lens development. In this study, we used CRISPR gene editing to produce zebrafish lines with null mutations for each of the three α-crystallin genes (cryaa, cryaba and cryabb). The absence of normal protein was confirmed by mass spectrometry, and lens phenotypes were assessed with differential interference contrast microscopy and histology. Loss of αA-crystallin produced a variety of lens defects with varying severity in larval lenses at 3 and 4 dpf but little substantial change in normal fiber cell denucleation. Loss of either αBa- or full-length αBb-crystallin produced no substantial lens defects. Mutation of each α-crystallin gene did not alter the mRNA levels of the remaining two, suggesting a lack of genetic compensation. These data confirm a developmental role for αA-crystallin in lens development, but the range of phenotype severity suggests that its loss simply increases the chance for defects and that the protein is not essential. Our finding that cryaba and cryabb mutants lack noticeable lens defects is congruent with insubstantial transcript levels in lens epithelial and fiber cells. Future experiments can explore the molecular consequences of cryaa mutation and causes of lens defects in this null mutant, as well as the roles of other genes in lens development and function.
A discovery-based proteomics approach identifies protein disulfide isomerase (PDIA1) as a biomarker of β cell stress in type 1 diabetes
Authors: Syed Farooq , Singhal Divya , Raedschelders Koen , Krishnan Preethi , Bone Robert N. , McLaughlin Madeline R. , Van Eyk Jennifer E. , Mirmira Raghavendra G. , Yang Mei-Ling , Mamula Mark J. , Wu Huanmei , Liu Xiaowen , Evans-Molina Carmella
Published: December 2021

Background

Activation of stress pathways intrinsic to the β cell are thought to both accelerate β cell death and increase β cell immunogenicity in type 1 diabetes (T1D). However, information on the timing and scope of these responses is lacking.

Methods

To identify temporal and disease-related changes in islet β cell protein expression, data independent acquisition-mass spectrometry was performed on islets collected longitudinally from NOD mice and NOD-SCID mice rendered diabetic through T cell adoptive transfer.

Findings

In islets collected from female NOD mice at 10, 12, and 14 weeks of age, we found a time-restricted upregulation of proteins involved in the maintenance of β cell function and stress mitigation, followed by loss of expression of protective proteins that heralded diabetes onset. Pathway analysis identified EIF2 signaling and the unfolded protein response, mTOR signaling, mitochondrial function, and oxidative phosphorylation as commonly modulated pathways in both diabetic NOD mice and NOD-SCID mice rendered acutely diabetic by adoptive transfer, highlighting this core set of pathways in T1D pathogenesis. In immunofluorescence validation studies, β cell expression of protein disulfide isomerase A1 (PDIA1) and 14-3-3b were found to be increased during disease progression in NOD islets, while PDIA1 plasma levels were increased in pre-diabetic NOD mice and in the serum of children with recent-onset T1D compared to age and sex-matched non-diabetic controls.

Interpretation

We identified a common and core set of modulated pathways across distinct mouse models of T1D and identified PDIA1 as a potential human biomarker of β cell stress in T1D.
Extracting dynamical understanding from neural-mass models of mouse cortex
Authors: Siu Pok Him , Müller Eli , Zerbi Valerio , Aquino Kevin , Fulcher Ben D.
Published: December 2021

New brain atlases with high spatial resolution and whole-brain coverage have rapidly advanced our knowledge of the brain’s neural architecture, including the systematic variation of excitatory and inhibitory cell densities across the mammalian cortex. But understanding how the brain’s microscale physiology shapes brain dynamics at the macroscale has remained a challenge. While physiologically based mathematical models of brain dynamics are well placed to bridge this explanatory gap, their complexity can form a barrier to providing clear mechanistic interpretation of the dynamics they generate. In this work we develop a neural-mass model of the mouse cortex and show how bifurcation diagrams, which capture local dynamical responses to inputs and their variation across brain regions, can be used to understand the resulting whole-brain dynamics. We show that strong fits to resting-state functional magnetic resonance imaging (fMRI) data can be found in surprisingly simple dynamical regimes—including where all brain regions are confined to a stable fixed point—in which regions are able to respond strongly to variations in their inputs, consistent with direct structural connections providing a strong constraint on functional connectivity in the anesthetized mouse. We also use bifurcation diagrams to show how perturbations to local excitatory and inhibitory coupling strengths across the cortex, constrained by cell-density data, provide spatially dependent constraints on resulting cortical activity, and support a greater diversity of coincident dynamical regimes. Our work illustrates methods for visualizing and interpreting model performance in terms of underlying dynamical mechanisms, an approach that is crucial for building explanatory and physiologically grounded models of the dynamical principles that underpin large-scale brain activity.
Applying a Modified Metabarcoding Approach for the Sequencing of Macrofungal Specimens from Fungarium Collections
Authors: Olds C. Gary , Berta-Thompson Jessie W. , Loucks Justin J. , Levy Richard A. , Wilson Andrew W.
Published: December 2021

Premise:Fungaria are a largely untapped source for understanding fungal biodiversity. The effort and cost in producing DNA barcode sequence data for large numbers of fungal specimens can be prohibitive. This study applies a modified metabarcoding approach that provides a labor and cost-effective solution for sequencing the fungal DNA barcode from hundreds of specimens at once.

Methods:A two-step PCR approach uses nested barcoded primers to nrITS2 sequence data. We applied this to 766 macrofungal specimens that represent a broad taxonomic sampling of the Dikarya, of which 382Lactariusspecimens are used to identify molecular operational taxonomic units (MOTUs) through a phylogenetic approach. Scripts in Python and R were used to organize sequence data and execute packages CutAdapt and DADA2 were used for primer removal and assessing sequence quality. Sequences were compared to NCBI and UNITE databases and Sanger-produced sequences.

Results:Specimen taxonomic identities from nrITS2 sequence data are >90% accurate across all specimens sampled. Phylogenetic analysis ofLactariussequences identified 20 MOTUs.

Discussion:The results demonstrate the capacity of these methods to produce nrITS2 sequences from large numbers of fungarium specimens. This provides an opportunity to more effectively use fungarium collections in advancing fungal diversity identification and documentation.
Transcriptome-wide analysis of microRNA-mRNA correlations in unperturbed tissue transcriptomes identifies microRNA targeting determinants
Authors: Trinidad Barnech Juan Manuel , Fort Rafael Sebastián , Barnech Guillermo Trinidad , Garat Beatriz , Duhagon María Ana
Published: December 2021

MicroRNAs are small RNAs that regulate gene expression through complementary base pairing with their target mRNAs. Given the small size of the pairing region and the large number of mRNAs that each microRNA can control, identifying biologically relevant targets is difficult. Since current knowledge of target recognition and repression has mainly relied on in vitro studies, we sought to determine if the interrogation of gene expression data of unperturbed tissues could yield new insight into these processes. The transcriptome-wide repression of all the microRNA-mRNA canonical interaction sites (seed and 3’-supplementary regions, identified by sole base complementarity) was calculated as a normalized Spearman correlation (Z-score) between the abundance of the transcripts in the PRAD-TCGA tissues (RNA-seq and small RNA-seq data of 546 samples). Using the repression values obtained, we confirmed established properties or microRNA targeting efficacy, such as the preference for gene regions (3’UTR > CDS > 5’UTR), the correspondence between repression and seed length (6mer < 7mer < 8mer), and the contribution to the repression exerted by the 3’-supplementary pairing at nucleotides 13-16 of the microRNA. Our results suggest that the 7mer-A1 seed could be more repressive than the 7mer-m8, while they have similar efficacy when they interact using the 3’-supplementary pairing. The 6mer+suppl sites yielded a normalized Z-score of repression similar to the sole 7mer-A1 seeds, alerting its potential biological relevance. We then used the approach to further characterize the 3’-supplementary pairing using 39 microRNAs that hold repressive 3’-supplementary interactions. The analysis of the bridge between seed and 3’-supplementary pairing sites confirmed the optimum +1 offset previously evidenced, but higher offsets appear to have similar repressive strength. The selected microRNAs show a low GC content at positions 13-16 and base preferences that allow a sequence motif identification. Our study demonstrates that transcriptome-wide analysis of microRNA-mRNA correlations in large, matched RNA-seq and small-RNA-seq data can uncover hints of microRNA targeting determinants operating in the in vivo unperturbed set. Finally, we provide a bioinformatic tool to identify microRNA-mRNA candidate interactions based on sequence complementarity of the seed and 3’-supplementary regions.
Estimating Microbial Population Data from Optical Density
Authors: Mira Portia , Yeh Pamela , Hall Barry G.
Published: December 2021

The spectrophotometer has been used for decades to measure the density of bacterial populations as the turbidity expressed as optical density – OD. However, the OD alone is an unreliable metric and is only proportionately accurate to cell titers to about an OD of 0.1. The relationship between OD and cell titer depends on the configuration of the spectrophotometer, the length of the light path through the culture, the size of the bacterial cells, and the cell culture density. We demonstrate the importance of plate reader calibration to identify the exact relationship between OD and cells/ml. We use four bacterial genera and two sizes of micro-titer plates (96-well and 384-well) and show that the cell/ml per unit OD depends heavily on the bacterial cell size and plate size. We applied our calibration curve to real growth curve data and conclude the cells/ml – rather than OD – is a metric that can be used to directly compare results across experiments, labs, instruments, and species.
sRNAs enriched in outer membrane vesicles of pathogenic Flavobacterium psychrophilum interact with immune genes of rainbow trout
Authors: Chapagain Pratima , Ali Ali , Kidane Destaaalem T. , Farone Mary , Salem Mohamed
Published: December 2021

Outer membrane vesicles (OMVs) released by gram-negative bacteria during host-pathogen interactions harbor cargos, such as DNA, RNA, toxins, and virulence factors. We hypothesized that sRNAs carried within OMVs of Flavobacterium psychrophilum interact with host immune genes and affect their expression. OMVs were isolated from F. psychrophilum and visualized using transmission electron microscopy (TEM). RNA-Seq datasets generated from whole-cell F. psychrophilum and their OMVs indicated enrichment of specific sRNAs in the OMVs compared to the parent cell. Fluorescent in situ hybridization (FISH) and confocal microscopy confirmed the expression of a randomly chosen sRNA.

Integrated RNA-Seq analyses of host transcriptome and bacterial sRNAs on day 5 post-infection of F. psychrophilum-resistant and -susceptible rainbow trout genetic lines revealed 516 protein-coding, 595 lncRNA, and 116 bacterial sRNA differentially expressed (DE) transcripts. Integrated and network analyses of these DE transcripts revealed immune genes targeted by bacterial sRNAs. On the top of these genes, an isoform encoding anaphase-promoting complex subunit 13 (ANAPC13_1) was highly upregulated and exhibited interaction and reciprocal expression with 21 DE sRNAs enriched in OMVs and/or located in pathogenicity islands (PAIs). In vitro treatment of the rainbow trout epithelial cell line RTgill-W1 with OMVs showed signs of cell autolysis accompanied by dynamic changes in expression of host genes when profiled 24h following treatment. The OMV-enriched sRNAs, soFE013584 and soFE002123, showed high interactions with the protection of telomeres 1 gene (POT1); essential for chromosome stability and cellular viability. Modulation of the host gene expression following OMV-treatment, which favors elements from the phagocytic, endocytic, and antigen presentation pathways in addition to HSP70, HSP90, and cochaperone proteins, provided evidence for a potential role of OMVs in boosting the host immune response. In conclusion, our work identified novel microbial targets and inherent characteristics of OMVs that could open up new avenues of treatment and prevention of fish infections.
The antique genetic plight of the Mediterranean monk seal (Monachus monachus)
Authors: Salmona Jordi , Dayon Julia , Lecompte Emilie , Karamanlidis Alexandros A. , Aguilar Alex , Fernandez De Larrinoa Pablo , Pires Rosa , Mo Giulia , Panou Aliki , Agnesi Sabrina , Borrell Asunción , Danyer Erdem , Öztürk Bayram , Tonay Arda M. , Anestis Anastasios K. , González Luis M. , Dendrinos Panagiotis , Gaubert Philippe
Published: December 2021

Disentangling the impact of Late Quaternary climate change from human activities can have crucial implications on the conservation of endangered species. We investigated the population genetics and demography of the Mediterranean monk seal (Monachus monachus), one of the world’s most endangered marine mammals, through an unprecedented dataset encompassing historical (extinct) and extant populations from the eastern North Atlantic to the entire Mediterranean Basin. We show that Cabo Blanco (Western Sahara/Mauritania), Madeira, Western Mediterranean (historical range), and Eastern Mediterranean regions segregate into four populations. This structure is likely the consequence of recent drift, combined with long-term isolation by distance (R2= 0.7), resulting from prevailing short-distance (< 500 km) and infrequent long-distance dispersal (< 1,500 km). All populations (Madeira especially), show high levels of inbreeding and low levels of genetic diversity, seemingly declining since historical time, but surprisingly not being impacted by the 1997 massive die-off in Cabo Blanco. Approximate Bayesian Computation analyses support scenarios combining local extinctions and a major effective population size decline in all populations during Antiquity. Our results suggest that the early densification of human populations around the Mediterranean Basin coupled with the development of seafaring techniques were the main drivers of the decline of Mediterranean monk seals.
ROOT PENETRATION INDEX 3, a major quantitative trait locus (QTL) associated with root system penetrability in Arabidopsis
Authors: Bello Elohim Bello , Cambron Thelma Y. Rico , Álvarez Rubén Rellán , Estrella Luis Herrera
Published: December 2021

Soil mechanical impedance precludes root penetration, confining root system development to shallow soil horizons where mobile nutrients are scarce. Using a two-phase-agar system, we characterized Arabidopsis thaliana responses to low and high mechanical impedance at three root penetration stages. We found that seedlings whose roots fail to penetrate agar barriers show drastic changes in shoot and root morphology, while those capable of penetrating have only minor morphological effects. The assessment of 21 Arabidopsis accessions revealed that primary root penetrability (PRP) varies widely among accessions. To search for quantitative trait loci (QTLs) associated to root system penetrability, we evaluated a recombinant inbred population (RIL) derived from Landsberg erecta (Ler-0, with a high PRP) and Shahdara (Sha, with a low PRP) accessions. QTL analysis revealed a major-effect QTL localized in chromosome 3 (q-RPI3), which accounted for 29.98% (LOD = 8.82) of the total phenotypic variation. Employing an introgression line (IL-321), with a homozygous q-RPI3 region from Sha in the Ler-0 genetic background, we demonstrated that q-RPI3 plays a crucial role in root penetrability. This multiscale study revels new insights into root plasticity during the penetration process in hard agar layers, natural variation and genetic architecture behind primary root penetrability in Arabidopsis.

Highlight

We found a wide natural variation in the capacity of Arabidopsis accessions to penetrate hard agar layers. Using a Ler-0 x Sha recombinant inbred population, a major-effect QTL (q-RPI3) strongly associated with root penetrability of compact agar layers was identified.
A single-cell atlas of human fetal lung development between 14 and 19 weeks of gestation
Authors: Renesme Laurent , Lesage Flore , Cook David , Zhong Shumei , Hänninen Satu , Carpén Olli , Mižíková Ivana , Thébaud Bernard
Published: December 2021

Rationale

Human lung development has been mainly described in morphologic studies and the potential underlying molecular mechanisms were extrapolated from animal models. Therefore, there is a need to gather knowledge from native human lung tissue. In this study we describe changes at a single-cell level in human fetal lungs during the pseudoglandular stage.

Methods

We report the cellular composition, cell trajectories and cell-to-cell communication in developing human lungs with single-nuclei RNA sequencing (snRNA-seq) on 23,251 nuclei isolated from nine human fetuses with gestational ages between 14 to 19 weeks of gestation.

Results

We identified nine different cell types, including a rare pulmonary neuroendocrine cells population. For each cell type, marker genes are reported, and selected marker genes are used for spatial validation with fluorescent RNA in situ hybridization. Enrichment and developmental trajectory analysis provide insight into molecular mechanisms and signaling pathways within individual cell clusters according to gestational age. Lastly, ligand-receptor analysis highlights determinants of cell-to-cell communication among the different cell types through the pseudoglandular stage, including general developmental pathways (NOTCH and TGFB), as well as more specific pathways involved in vasculogenesis, neurogenesis, and immune system regulation.

Conclusion

These findings provide a clinically relevant background for research hypotheses generation in projects studying normal or impaired lung development and help to develop and validate surrogate models to study human lung development, such as human lung organoids.

TAKE HOME MESSAGE

Using a single-cell transcriptomic approach (single-nuclei RNA sequencing), we describe here, for the first time, the cellular landscape, cell developmental trajectories, and cell-to-cell communication in the developing human lung during the pseudoglandular stage.
Toll-like receptor 5 as a novel receptor for fungal zymosan
Authors: Reuter Stefanie , Herold Kristina , Domrös Jana , Mrowka Ralf
Published: December 2021

Microbial pathogens carry specific structural patterns which were termed pathogen-associated molecular patterns (PAMPs). Toll-like receptors (TLRs) as key elements for the recognition of microbial pathogens are necessary for the activation of innate immune pathways. TLRs are activated by binding PAMPs of bacteria, viruses and fungi and initiate a signaling pathway resulting in the activation of transcription factors which modulate the production of various proinflammatory cytokines. It is not fully clear in detail which microbial pattern is recognized by which TLR. Here we show for the first time that TLR5 is a strong receptor for the yeast particle zymosan. We have generated stable human cell lines with combinations of TLR2 and TLR5 knock in/knock out together with stable nuclear factor kappaB (NF-κB) luciferase reporters. We found that both receptors TLR5 and TLR2 lead to an independent activation of the NF-κB pathway when simulated with zymosan. Our results demonstrate that TLR5 is a receptor for the fungal particle zymosan in addition to bacterial fragments like flagellin. Distinct cytokine patterns might suggest that TLR5 is potentially important for the differentiation in the recognition of the specific type of the foreign microorganisms and in the specific host defense response.
In situ distance measurements in a membrane transporter using maleimide functionalized orthogonal spin labels and 5-pulse electron double resonance spectroscopy
Authors: Ketter Sophie , Dajka Marina , Rogozhnikova Olga , Dobrynin Sergey A. , Tormyshev Victor M. , Bagryanskaya Elena G. , Joseph Benesh
Published: December 2021

Spectroscopic investigation of membrane proteins in their native environment is a challenging task. Earlier we demonstrated the feasibility to measure precise distances within outer membrane proteins in E. coli and native membranes using methanethiosulfonate (MTS) functionalized labels combined with pulsed electron double resonance spectroscopy. Here we show the application of maleimide functionalized Gd(III), nitroxide, and trityl labels for in situ distance measurement using the cobalamin transporter BtuB. These labels enabled distance measurements for BtuB in E. coli and native outer membranes and in the membranes maleimide-Gd-DOTA also is effective. Further, we show that the observable dipolar evolution time can be significantly prolonged in the native environments using the Carr-Purcell 5-pulse electron double resonance sequence. For a nitroxide-nitroxide pair, application of sech/tanh inversion pulses substantially suppressed the 4-pulse artifact at the Q-band frequency. In the case of a nitroxide-trityl pair, Gaussian pump pulses of varying amplitude are sufficient to suppress the artifact to the typical noise level. The feasibility of a range of bioresistant spin labels and the 5-pulse electron double resonance offers promising tools for investigating heterooligomeric membrane protein complexes in their native environment.
Crosstalk between biochemical signaling and trafficking determines AMPAR dynamics in synaptic plasticity
Authors: Bell M. K. , Rangamani P.
Published: December 2021

Synaptic plasticity involves the modification of both biochemical and structural components of neurons. Many studies have revealed that the change in the number density of the glutamatergic receptor AMPAR at the synapse is proportional to synaptic weight update; increase in AMPAR corresponds to strengthening of synapses while decrease in AMPAR density weakens synaptic connections. The dynamics of AMPAR are thought to be regulated by upstream signaling, primarily the calcium-CaMKII pathway, trafficking to and from the synapse, and influx from extrasynaptic sources. Here, we have developed a set of models using compartmental ordinary differential equations to systematically investigate contributions of signaling and trafficking variations on AMPAR dynamics at the synaptic site. We find that the model properties including network architecture and parameters significantly affect the integration of fast upstream species by slower downstream species. Furthermore, we predict that the model outcome, as determined by bound AMPAR at the synaptic site, depends on (a) the choice of signaling model (bistable CaMKII or monostable CaMKII dynamics), (b) trafficking versus influx contributions, and (c) frequency of stimulus. Therefore, AMPAR dynamics can have unexpected dependencies when upstream signaling dynamics (such as CaMKII and PP1) are coupled with trafficking modalities.
Multimodal lesion network mapping to predict sensorimotor behavior in stroke patients
Authors: Jimenez-Marin Antonio , Bruyn Nele De , Gooijers Jolien , Llera Alberto , Meyer Sarah , Alaerts Kaat , Verheyden Geert , Swinnen Stephan P. , Cortes Jesus M.
Published: December 2021

Lesion network mapping (LNM) has proved to be a successful technique to map symptoms to brain networks after acquired brain injury. Beyond the characteristics of a lesion, such as its etiology, size or location, LNM has shown that common symptoms in patients after injury may reflect the effects of their lesions on the same circuits, thereby linking symptoms to specific brain networks. Here, we extend LNM to its multimodal form, using a combination of functional and structural connectivity maps drawn from data from 1000 healthy participants in the Human Connectome Project. We applied the multimodal LNM to a cohort of 54 stroke patients with the aim of predicting sensorimotor behavior, as assessed through a combination of motor and sensory tests. Test scores were predicted using a Canonical Correlation Analysis with multimodal brain maps as independent variables, and cross-validation strategies were employed to overcome overfitting. The results obtained led us to draw three conclusions. First, the multimodal analysis reveals how functional connectivity maps contribute more than structural connectivity maps in the optimal prediction of sensorimotor behavior. Second, the maximal association solution between the behavioral outcome and multimodal lesion connectivity maps suggests an equal contribution of sensory and motor coefficients, in contrast to the unimodal analyses where the sensory contribution dominates in both structural and functional maps. Finally, when looking at each modality individually, the performance of the structural connectivity maps strongly depends on whether sensorimotor performance was corrected for lesion size, thereby eliminating the effect of larger lesions that produce more severe sensorimotor dysfunction. By contrast, the maps of functional connectivity performed similarly irrespective of any correction for lesion size. Overall, these results support the extension of LNM to its multimodal form, highlighting the synergistic and additive nature of different types of imaging modalities, and the influence of their corresponding brain networks on behavioral performance after acquired brain injury.
Putting perception into action: Inverse optimal control for continuous psychophysics
Authors: Straub Dominik , Rothkopf Constantin A.
Published: December 2021

Psychophysical methods are a cornerstone of psychology, cognitive science, and neuroscience where they have been used to quantify behavior and its neural correlates for a vast range of mental phenomena. Their power derives from the combination of controlled experiments and rigorous analysis through signal detection theory. Unfortunately, they require many tedious trials and preferably highly trained participants. A recently developed approach, continuous psychophysics, promises to transform the field by abandoning the rigid trial structure involving binary responses and replacing it with continuous behavioral adjustments to dynamic stimuli. However, what has precluded wide adoption of this approach is that current analysis methods recover perceptual thresholds, which are one order of magnitude larger compared to equivalent traditional psychophysical experiments. Here we introduce a computational analysis framework for continuous psychophysics based on Bayesian inverse optimal control. We show via simulations and on previously published data that this not only recovers the perceptual thresholds but additionally estimates subjects’ action variability, internal behavioral costs, and subjective beliefs about the experimental stimulus dynamics. Taken together, we provide further evidence for the importance of including acting uncertainties, subjective beliefs, and, crucially, the intrinsic costs of behavior, even in experiments seemingly only investigating perception.
Structure and mechanism of a cytosine transport protein
Authors: Hatton Caitlin E. , Brotherton Deborah H. , Cameron Alexander D.
Published: December 2021

CodB is a cytosine transporter from the Nucleobase-Cation-Symport-1 (NCS1) transporter family, a member of the widespread LeuT superfamily. Previous experiments with the nosocomial pathogenPseudomonas aeruginosahave shown CodB also to be important in the uptake of 5-fluorocytosine, which has been suggested as a novel drug to combat antimicrobial resistance by suppressing virulence in the organism. Here we solve the crystal structure of CodB fromProteus vulgaris, at 2.4Å resolution in complex with cytosine. We show that the protein carries out the sodium-dependent uptake of cytosine and can bind 5-fluorocytosine. Comparison of the substrate-bound structures of CodB and the hydantoin transporter Mhp1, the only other NCS1 family member for which the structure is known, highlight the importance of the hydrogen bonds that the substrates make with the main chain at the breakpoint in the discontinuous helix, TM6. In contrast to other LeuT superfamily members, neither CodB nor Mhp1 make specific interactions with residues on TM1. Comparison of the structures provides insight into the intricate mechanisms of how these proteins transport substrates across the plasma membrane.
Gene by environment interaction mouse model reveals a functional role for 5-hydroxymethylcytosine in neurodevelopmental disorders
Authors: Papale Ligia A. , Madrid Andy , Zhang Qi , Chen Kailei , Sak Lara , Keleş Sündüz , Alisch Reid S.
Published: December 2021

Mouse knockouts of Cntnap2 show altered neurodevelopmental behavior, deficits in striatal GABAergic signaling, and a genome-wide disruption of an environmentally sensitive DNA methylation modification (5-hydroxymethylcytosine [5hmC]) in the orthologs of a significant number of genes implicated in human neurodevelopmental disorders. We tested adult Cntnap2 heterozygous mice (Cntnap2+/−; lacking behavioral or neuropathological abnormalities) subjected to a prenatal stress and found that prenatally stressed Cntnap2+/− female mice show repetitive behaviors and altered sociability, similar to the homozygote phenotype. Genomic profiling revealed disruptions in hippocampal and striatal 5hmC levels that are correlated to altered transcript levels of genes linked to these phenotypes (e.g., Reln, Dst, Trio, and Epha5). Chromatin immunoprecipitation coupled with high-throughput sequencing and hippocampal nuclear lysate pull-down data indicated that 5hmC abundance alters the binding of the transcription factor CLOCK near the promoters of these genes (e.g., Palld, Gigyf1, and Fry), providing a mechanistic role for 5hmC in gene regulation. Together, these data support gene-by-environment hypotheses for the origins of mental illness and provide a means to identify the elusive factors contributing to complex human diseases.
Tangible symmetry elements and space-group models to guide from molecular to solid-state composition
Authors: Graw Nico , Stalke Dietmar
Published: December 2021

The ability to imagine symmetry and the spatial arrangement of atoms and molecules is crucial in chemistry in general. Teaching and understanding crystallography and the composition of the solid state therefore require understanding of symmetry elements and their relationships. To foster the student's spatial imagination, models representing a range of concepts from individual rotation axes to complete space groups have been designed and built. These models are robust and large enough to be presented and operated in a lecture hall, and they enable students to translate conventional 2D notations into 3D objects and vice versa. Tackling them hands-on means understanding them.
CRISTAL-ITE: a single-crystal X-ray diffractometer scale model for scientific dissemination
Authors: Giorgi Michel , Berchadsky Yves
Published: December 2021

This article presents the design and manufacture of an automated scale model of a four-circle single-crystal X-ray diffractometer that can be used for scientific dissemination. The purpose of this device is to reach out to the wider public and students to introduce them in an entertaining way to one of the laboratory apparatuses to which they do not usually have access, to talk to them about crystallography in the broadest sense, to develop concepts in various fields of science and technology, and to initiate interest and discussions. The main technical aspects of the project are described, with the expectation that such an approach could be useful to anyone involved in scientific dissemination and could be developed for other laboratory equipment and other disciplines. This kind of device can also be the subject of scientific and technological projects in close collaboration with educational institutions.
anti-1-(Benzylamino)glyoxime. Corrigendum
Authors: Köysal Yavuz , Işık Şamil , Sarıkavaklı Nursabah , Eyduran Fatih
Published: December 2021

In the paper by Köysal et al. [Acta Cryst. (2004), E60, o515–o516], there was an error in the name of the last author.