bluuebunny/crossref_metadata_2025 huggingface dataset:
Individual-specific memory reinstatement patterns within human face-selective cortex
Authors: Chen Yvonne Y. , Areti Aruni , Yoshor Daniel , Foster Brett L.
Published: August 2023

Humans have the remarkable ability to vividly retrieve sensory details of past events. According to the theory of sensory reinstatement, during remembering, brain regions involved in the sensory processing of prior events are reactivated to support this perception of the past. Recently, several studies have emphasized potential transformations in the spatial organization of reinstated activity patterns. In particular, studies of scene stimuli suggest a clear anterior shift in the location of retrieval activations compared with those during perception. However, it is not clear that such transformations occur universally, with evidence lacking for other important stimulus categories, particularly faces. Critical to addressing these questions, and to studies of reinstatement more broadly, is the growing importance of considering meaningful variations in the organization of sensory systems across individuals. Therefore, we conducted a multi-session neuroimaging study to first carefully map individual participants face-selective regions within ventral temporal cortex (VTC), followed by a second session to examine the correspondence of activity patterns during face memory encoding and retrieval. Our results showed distinct configurations of face-selective regions within the VTC across individuals. While a significant degree of overlap was observed between face perception and memory encoding, memory retrieval engagement exhibited a more selective and constricted reinstatement pattern within these regions. Importantly, these activity patterns were consistently tied to individual-specific neural substrates, but did not show any consistent direction of spatial transformation (e.g., anteriorization). To provide further insight to these findings, we also report on unique human intracranial recordings from VTC under the same experimental conditions. Our findings highlight the importance of considering individual variations in functional neuroanatomy in the context of assessing the nature of cortical reinstatement. Consideration of such factors will be important for establishing general principles shaping the neural transformations that occur from perception to memory.
Development of Cell-Derived Plasma Membrane Vesicles as a Nanoparticle Encapsulation and Delivery System
Authors: Kheradmandi Mahsa , Farnoud Amir M. , Burdick Monica M.
Published: August 2023

Background

Developing non-invasive delivery platforms with a high level of structural and/or functional similarity to biological membranes is highly desirable to reduce toxicity and improve targeting capacity of nanoparticles. Numerous studies have investigated the impacts of physicochemical properties of engineered biomimetic nanoparticles on their interaction with cells, yet technical difficulties have led to the search for better biomimetics, including vesicles isolated directly from live cells. Cell-derived giant plasma membrane vesicles (GPMVs), in particular, offer a close approximation of the intact cell plasma membrane by maintaining the latter’s compositional complexity, protein positioning in a fluid-mosaic pattern, and physical and mechanical properties. Thus, to overcome technical barriers of prior nanoparticle delivery approaches, we aimed to develop a novel method using GPMVs to encapsulate a variety of engineered nanoparticles, then use these core-shell, nanoparticle-GPMV vesicle structures to deliver cargo to other cells.

Results

The GPMV system in this study was generated by chemically inducing vesiculation in A549 cells, a model human alveolar epithelial line. These cell-derived GPMVs retained encapsulated silica nanoparticles (50 nm diameter) for at least 48 hours at 37 °C. GPMVs showed nearly identical lipid and protein membrane profiles as the parental cell plasma membrane, with or without encapsulation of nanoparticles. Notably, GPMVs were readily endocytosed in the parental A549 cell line as well as the human monocytic THP-1 cell line. Higher cellular uptake levels were observed for GPMV-encapsulated nanoparticles compared to control groups, including free nanoparticles. Further, GPMVs delivered a variety of nanoparticles to parental cells with reduced cytotoxicity compared to free nanoparticles at concentrations that were otherwise significantly toxic.

Conclusions

We have introduced a novel technique to load nanoparticles within the cell plasma membrane during the GPMV vesiculation process. These GPMVs are capable of (a) encapsulating different types of nanoparticles (including larger and not highly-positively charged bodies that have been technically challenging cargoes) using a parental cell uptake technique, and (b) improving delivery of nanoparticles to cells without significant cytotoxicity. Ultimately, endogenous surface membrane proteins and lipids can optimize the physicochemical properties of cell membrane-derived vesicles, which could lead to highly effective cell membrane-based nanoparticle/drug delivery systems.
The roles of RNA editing in cancer immunity through interacting with interferon
Authors: Wu Sijia , Qin Xinyu , Lu Zhennan , Wen Jianguo , Yang Mengyuan , Kim Pora , Zhou Xiaobo , Huang Liyu
Published: August 2023

The interferon-activated tumor innate immunity can be primed by specific double-stranded RNA (dsRNA) sensors upon stimulation. A-to-I RNA editing in the dsRNA regions can have a potential function to regulate interferon-related cancer immunity. A systematical analysis of both the editing enzyme and specific enriched editing region in patients, tissues, and cell lines is performed to reveal the underlying mechanisms. We then validate the preferred editing of dsRNA regions, identify the hyper-editing in severe tumors, and discover the negative effect of editing on cancer immunity. Specifically, RNA editing acts as an inhibitor ofPKR- andMDA5-related interferon pathways through the regulations of miRNAs and RNA-binding proteins and the deactivation of dsRNA sensors. With the alteration of interferons, subsequently, RNA editing represses the infiltration of CD8 and CD4 T cells and reduces the sensitivities of cancer drugs, such as cisplatin. These analyses on A-to-I RNA editing can improve the knowledge of tumorigenesis, immunology, and cancer-targeted immunotherapy.

Highlights

The preferred dsRNA region for RNA editing is validated.

Upregulation of RNA editing in severe tumors is discovered.

RNA editing inhibits PKR- and MDA5-related cancer immunity.

RNA editing represses the infiltration of CD8 and CD4 T cells.

RNA editing reduces the sensitivities of cancer drugs.
RFC1AAGGG pentanucleotide repeats form parallel G-quadruplex: structural implications for aberrant molecular cascades in CANVAS
Authors: Wang Yang , Wang Junyan , Yan Zhenzhen , Hou Jianing , Wan Liqi , Yi Jie , Guo Pei , Han Da
Published: August 2023

A pentanucleotide repeat expansion (PRE) of (AAGGG)nin the replication factor C subunit 1 (RFC1) gene is recently identified as the genetic cause of cerebellar ataxia, neuropathy, and vestibular areflexia syndrome (CANVAS), and also linked to several other neurodegenerative disorders including the Parkinson’s disease. However, the molecular mechanism by whichRFC1PRE drives pathology remains poorly understood. Here, for the first time we discovered and determined the high-resolution structures of parallel G-quadruplex formed by AAGGG repeats within the pathogenicRFC1PRE, revealing an intriguing conformational plasticity at the 3’-termi that allows stacking of multiple G4s. We further identify a molecular mechanism by which the DNA G4 inRFC1PRE impedes polymerase processivity leading to replication stalling and transcription inhibitionin vitroin a repeat-length-dependent manner, and the transcription inhibition could partially contribute to a reduced gene expression in cells. Our results demonstrate that the DNA G-quadruplex ofRFC1PRE participate in aberrant molecular cascades, and provide an unprecedented high-resolution structural target to discover helicases and ligands that resolve the pathogenic G4 for therapeutic intervention.
A competitive disinhibitory network for robust optic flow processing inDrosophila
Authors: Erginkaya Mert , Cruz Tomás , Brotas Margarida , Steck Kathrin , Nern Aljoscha , Torrão Filipa , Varela Nélia , Bock Davi , Reiser Michael , Chiappe M Eugenia
Published: August 2023

Many animals rely on optic flow for navigation, using differences in eye image velocity to detect deviations from their intended direction of travel. However, asymmetries in image velocity between the eyes are often overshadowed by strong, symmetric translational optic flow during navigation. Yet, the brain efficiently extracts these asymmetries for course control. While optic flow sensitive-neurons have been found in many animal species, far less is known about the postsynaptic circuits that support such robust optic flow processing. In the flyDrosophila melanogaster, a group of neurons called the horizontal system (HS) are involved in course control during high-speed translation. To understand how HS cells facilitate robust optic flow processing, we identified central networks that connect to HS cells using full brain electron microscopy datasets. These networks comprise three layers: convergent inputs from different, optic flow-sensitive cells, a middle layer with reciprocal, and lateral inhibitory interactions among different interneuron classes, and divergent output projecting to both the ventral nerve cord (equivalent to the vertebrate spinal cord), and to deeper regions of the fly brain. By combining two-photon optical imaging to monitor free calcium dynamics, manipulating GABA receptors and modeling, we found that lateral disinhibition between brain hemispheres enhance the selectivity to rotational visual flow at the output layer of the network. Moreover, asymmetric manipulations of interneurons and their descending outputs induce drifts during high-speed walking, confirming their contribution to steering control. Together, these findings highlight the importance of competitive disinhibition as a critical circuit mechanism for robust processing of optic flow, which likely influences course control and heading perception, both critical functions supporting navigation.
OSGN-1 is a conserved flavin-containing monooxygenase required to stabilize the intercellular bridge in late cytokinesis
Authors: Goupil Eugénie , Lacroix Léa , Brière Jonathan , Guga Sandra , Saba-El-Leil Marc K. , Meloche Sylvain , Labbé Jean-Claude
Published: August 2023

Cytokinesis is the last step of cell division and is regulated by the small GTPase RhoA. RhoA activity is required for all steps of cytokinesis, including prior to abscission when daughter cells are ultimately physically separated. Like germ cells in all animals, theC. elegansembryonic germline founder cell initiates cytokinesis but does not complete abscission, leaving a stable intercellular bridge between the two daughter cells. Here we identify and characterizeC. elegansOSGN-1 as a novel cytokinetic regulator that promotes RhoA activity during late cytokinesis. Sequence analyses and biochemical reconstitutions reveal that OSGN-1 is a flavin-containing monooxygenase. Genetic analyses indicate that the monooxygenase activity of OSGN-1 is required to maintain active RhoA at the end of cytokinesis in the germline founder cell and to stabilize the intercellular bridge. Deletion ofOSGIN1in human cells results in an increase in binucleation as a result of cytokinetic furrow regression, and this phenotype can be rescued by expressing a catalytically-active form ofC. elegansOSGN-1, indicating that OSGN-1 and OSGIN1 are functional orthologs. We propose that OSGN-1 and OSGIN1 are novel, conserved monooxygenase enzymes required to maintain RhoA activity at the intercellular bridge during late cytokinesis and thus promote its stability, enabling proper abscission in human cells and bridge stabilization inC. elegansgerm cells.
Using deep long-read RNAseq in Alzheimer’s disease brain to assess medical relevance of RNA isoform diversity
Authors: Heberle Bernardo Aguzzoli , Brandon J. Anthony , Page Madeline L. , Nations Kayla A. , Dikobe Ketsile I. , White Brendan J. , Gordon Lacey A. , Fox Grant A. , Wadsworth Mark E. , Doyle Patricia H. , Williams Brittney A. , Fox Edward J. , Shantaraman Anantharaman , Ryten Mina , Goodwin Sara , Ghiban Elena , Wappel Robert , Mavruk-Eskipehlivan Senem , Miller Justin B. , Seyfried Nicholas T. , Nelson Peter T. , Fryer John D. , Ebbert Mark T. W.
Published: August 2023

Due to alternative splicing, human protein-coding genes average over eight RNA isoforms, resulting in nearly four distinct protein coding sequences per gene. Long-read RNAseq (IsoSeq) enables more accurate quantification of isoforms, shedding light on their specific roles. To assess the medical relevance of measuring RNA isoform expression, we sequenced 12 aged human frontal cortices (6 Alzheimer’s disease cases and 6 controls; 50% female) using one Oxford Nanopore PromethION flow cell per sample. Our study uncovered 53 new high-confidence RNA isoforms in medically relevant genes, including several where the new isoform was one of the most highly expressed for that gene. Specific examples includeWDR4(61%; microcephaly),MYL3(44%; hypertrophic cardiomyopathy), andMTHFS(25%; major depression, schizophrenia, bipolar disorder). Other notable genes with new high-confidence isoforms includeCPLX2(10%; schizophrenia, epilepsy) andMAOB(9%; targeted for Parkinson’s disease treatment). We identified 1,917 medically relevant genes expressing multiple isoforms in human frontal cortex, where 1,018 had multiple isoforms with different protein coding sequences, demonstrating the need to better understand how individual isoforms from a single gene body are involved in human health and disease, if at all. Exactly 98 of the 1,917 genes are implicated in brain-related diseases, including Alzheimer’s disease genes such asAPP(Aβ precursor protein; five),MAPT(tau protein; four), andBIN1(eight). As proof of concept, we also found 99 differentially expressed RNA isoforms between Alzheimer’s cases and controls, despite the genes themselves not exhibiting differential expression. Our findings highlight the significant knowledge gaps in RNA isoform diversity and their medical relevance. Deep long-read RNA sequencing will be necessary going forward to fully comprehend the medical relevance of individual isoforms for a “single” gene.
Differential conformational dynamics in two type-A RNA-binding domains drive the double-stranded RNA recognition and binding
Authors: Parvez Firdousi , Sangpal Devika , Paithankar Harshad , Amin Zainab , Chugh Jeetender
Published: August 2023

TAR RNA binding protein (TRBP) has emerged as a key player in the RNA interference (RNAi) pathway, wherein it binds to different pre-miRNAs and siRNAs, each varying in sequence and/or structure. We hypothesize that TRBP displays dynamic adaptability to accommodate heterogeneity in target RNA structures. Thus, it is crucial to ascertain the role of intrinsic and RNA-induced protein dynamics in RNA recognition and binding. We have previously elucidated the role of intrinsic and RNA-induced conformational exchange in the double-stranded RNA-binding domain 1 (dsRBD1) of TRBP in shape-dependent RNA recognition. The current study delves into the intrinsic and RNA-induced conformational dynamics of the TRBP-dsRBD2 and then compares it with the dsRBD1 study carried out previously. Remarkably, the two domains exhibit differential binding affinity to a 12 bp dsRNA owing to the presence of critical residues and structural plasticity. Further, we report that dsRBD2 depicts constrained conformational plasticity when compared to dsRBD1. Although, in the presence of RNA, dsRBD2 undergoes induced conformational exchange within the designated RNA-binding regions and other residues, the amplitude of the motions remains modest when compared to those observed in dsRBD1. We propose a dynamics-driven model of the two tandem domains of TRBP, substantiating their contributions to the versatility of dsRNA recognition and binding.

Significance Statement

Exploring the intricacies of RNA-protein interactions by delving into dynamics-based measurements not only adds valuable insights into the mechanics of RNA-protein interactions but also underscores the significance of conformational dynamics in dictating the functional outcome in such tightly regulated biological processes. In this study, we measure intrinsic and RNA-induced conformational dynamics in the second dsRBD, i.e., TRBP-dsRBD2, and compare the same with that carried out in the first dsRBD (TRBP-dsRBD1) of TRBP protein, a key player of the RNAi pathway. The study unveils the differential conformational space accessible to the two domains of TRBP, even though they both adopt a canonical dsRBD fold, thereby affecting how they interact with target RNAs.
A novel conceptual model of heart rate autonomic modulation based on a small-world modular structure and heterogeneous innervation of the sinoatrial node
Authors: Maltsev Alexander V , Stern Michael D , Lakatta Edward G , Maltsev Victor A
Published: August 2023

The current theory of cardiac pacemaker rate modulation by the autonomic nervous system is based on the concept that a primary pacemaker cell or a group of cells in the center of the sinoatrial node (SAN) can change its AP firing rate within a broad range, driving respective myocardium contractions to commensurate body demands. Experimental data show, however, that pacemaker cells are extremely heterogeneous, with different areas of the SAN or cell clusters specializing to drive APs at specific rates. Thus, higher heart rates under stress are mainly driven by cell clusters in superior SAN, whereas low rates by cell clusters in inferior SAN, with basal state rates generated somewhere in the middle of the node. Cells within different clusters feature different intrinsic electrophysiological and Ca cycling properties, sympathetic and parasympathetic innervation, and vasculature, thereby supporting effective shift of the system to an optimal rate (under given conditions) accompanied by respective shifts in the leading pacemaker site. Thus, the popular single-cell-based pacemaker theory does not capture this complex emerging paradigm of pacemaker function of SAN tissue revealed by recent experimental studies. Here we propose a more realistic, conceptual model of heart rate autonomic modulation based on these studies. Our new model (the ‘gear model’) simulates the SAN as a brain-like structure featuring a small world of loosely connected clusters (functional modules) of tightly coupled cells, modeled as Maltsev-Lakatta coupled-clock system. One module (the higher chronotropic gear) generates higher AP rates in basal state and under β-adrenergic stimulation, but its activity is strongly suppressed by parasympathetic stimulation. The other module (the lower gear) generates lower rates and has low sensitivity to parasympathetic stimulation. Such modular, gear-like system reproduces the respective shifts of the leading pacemaker site observed experimentally and features a wide range of rate modulation and robust function whilst conserving energy. In perspective, future refinement and application of this new pacemaker tissue mechanism will provide better understanding of cardiac pacemaker function, its deterioration in aging and disease, and ultimately the creation of new therapies to treat sick sinus syndrome and other SAN function-related cardiac arrythmias.
Assessment of emissions and potential occupational exposure to carbon monoxide during biowaste composting
Authors: Sobieraj Karolina , Giez Karolina , Koziel Jacek A. , Białowiec Andrzej
Published: August 2023

To date, only a few studies focused on the carbon monoxide (CO) production during waste composting; all targeted on CO inside piles. Here, the CO net emissions from compost piles and the assessment of worker’s occupational risk of exposure to CO at large-scale composting plants are shown for the first time. CO net emissions were measured at two plants processing green waste, sewage sludge, or undersize fraction of municipal solid waste. Effects of the location of piles (hermetised hall vs. open yard) and turning (before vs. after) were studied. Higher CO net emission rates were observed from piles located in a closed hall. The average CO flux before turning was 23.25 and 0.60 mg‧m−2‧h−1for hermetised and open piles, respectively, while after – 69.38 and 5.11 mg‧m−2‧h−1. The maximum CO net emissions occurred after the compost was turned (1.7x to 13.7x higher than before turning). The top sections of hermetised piles had greater CO emissions compared to sides. Additionally, 5% of measurement points of hermetised piles switched to ‘CO sinks’. The 1-h concentration in hermetised composting hall can reach max. ∼50 mg CO·m−3before turning, and >115 mg CO·m−3after, exceeding the WHO thresholds for a 1-h and 15-min exposures, respectively.
CURCUMIN HAS LOWER IC50 VALUES AGAINST A549 LUNG CANCER CELLS
Authors: Kaçar Sedat , Tomsuk Özlem , Şahintürk Varol
Published: August 2023

In this study, we aimed to determine the toxic effect of curcumin on A549 lung cancer cells and to show how this effect is reflected in morphology. Firstly, the toxic doses of curcumin against A549 lung cancer cells were determined via MTT and neutral red cytotoxicity tests by using the doses of 6.25, 12.5, 25, 50, 100, 200 and 400 µM. The morphological examination was performed using inverted microscope and light microscope with hematoxylin-eosin eosin.As a result, the IC50 dose of curcumin against A549 cells was 33 µM according to MTT cytotoxicity test and 52 µM according to the neutral red cytotoxicity test. When these doses were administered, the rounded and shrunken cells were visible in the inverted microscope, while apoptotic hallmarks such as nucleus condensation, renal nucleus structure and cellular shrinkage were detected in hematoxylin-eosin staining. In conclusion, the low IC50 values of curcumin for 24 hours indicate that curcumin is effective against A549 cells. It has the potential to be used either alone or in combination with other agents in cancer studies in case its low toxic effect on normal was assured.
Environmental effects on reproductive performance in preindustrial women
Authors: Colejo-Durán Lidia , Pelletier Fanie , Dillon Lisa , Gagnon Alain , Bergeron Patrick
Published: August 2023

Early life environments can have long-lasting effects on adult reproductive performance, but disentangling the influence of early and adult life environments on fitness is challenging, especially for long-lived species. Using a detailed dataset spanning over two centuries, we studied how early and adult life environments impact reproductive performance in preindustrial women. We assessed the effect of environmental conditions associated with the parish of birth and the parish where the women had their offspring. We considered resource availability differences between rural, urban, northern, and southern parishes by comparing women who switched environments during their lifetime with those who did not. We found that urban-born women had an earlier age at first reproduction and lower offspring survival to adulthood than rural-born women, while South Québec women had more offspring born than North Québec women. Moreover, switching from urban to rural led to higher offspring survival, while the reverse had the opposite effect. In addition, moving from South to North resulted in fewer offspring born and surviving, whereas moving from North to South had the inverse effect. Finally, women who changed from rural to urban and from South to North had their first child at an older age compared to those who stayed in the same urbanity type. Our study underscores the complex and interactive effects of early and adult life environments on reproductive traits, highlighting the need to consider both when studying environmental effects on reproductive outcomes.
RACER-m Leverages Structural Features for Sparse T Cell Specificity Prediction
Authors: Wang Ailun , Lin Xingcheng , Chau Kevin Ng , Onuchic José N. , Levine Herbert , George Jason T.
Published: August 2023

Reliable prediction of T cell specificity against antigenic signatures is a formidable task, complicated primarily by the immense diversity of T cell receptor and antigen sequence space and the resulting limited availability of training sets for inferential models. Recent modeling efforts have demonstrated the advantage of incorporating structural information to overcome the need for extensive training sequence data, yet disentangling the heterogeneous TCR-antigen structural interface to accurately predict the MHC-allele-restricted TCR-peptide binding interactions remained challenging. Here, we present RACER-m, a coarse-grained structural template model leveraging key biophysical information from the diversity of publicly available TCR-antigen crystal structures. We find explicit inclusion of structural content substantially reduces the required number of training examples for reliable prediction of TCR-recognition specificity and sensitivity across diverse biological contexts. We demonstrate that our structural model capably identifies biophysically meaningful point-mutants that affect overall binding affinity, distinguishing its ability in predicting TCR specificity of point mutants peptides from alternative sequence-based methods. Collectively, our approach combines biophysical and inferential learning-based methods to predict TCR-peptide binding events using sparse training data. Its application is broadly applicable to studies involving both closely-related and structurally diverse TCR-peptide pairs.
A parametrized two-domain thermodynamic model explains diverse mutational effects on protein allostery
Authors: Liu Zhuang , Gillis Thomas , Raman Srivatsan , Cui Qiang
Published: August 2023

New experimental findings continue to challenge our understanding of protein allostery. Recent deep mutational scanning study showed that allosteric hotspots in the tetracycline repressor (TetR) and its homologous transcriptional factors are broadly distributed rather than spanning well-defined structural pathways as often assumed. Moreover, hotspot mutation-induced allostery loss was rescued by distributed additional mutations in a degenerate fashion. Here, we develop a two-domain thermodynamic model for TetR, which readily rationalizes these intriguing observations. The model accurately captures the in vivo activities of various mutants with changes in physically transparent parameters, allowing the data-based quantification of mutational effects using statistical inference. Our analysis reveals the intrinsic connection of intra- and inter-domain properties for allosteric regulation and illustrate epistatic interactions that are consistent with structural features of the protein. The insights gained from this study into the nature of two-domain allostery are expected to have broader implications for other multidomain allosteric proteins.
Analyses of cell wall synthesis inClostridioides difficilereveal a diversification in cell division mechanisms in endospore-forming bacteria
Authors: Shrestha Shailab , Taib Najwa , Gribaldo Simonetta , Shen Aimee
Published: August 2023

Current models of bacterial cell division assume that the core synthases of the multiprotein divisome complex, FtsW-FtsI, are the primary drivers of septal peptidoglycan (PG) synthesis. These enzymes are typically encoded in the highly conserved division and cell wall (dcw) cluster and are considered to be universally essential for cell division. Here, we combine bioinformatics analyses with functional characterization in the pathogenClostridioides difficileto show thatdcw-encoded PG synthases have undergone a surprising specialization in the sole endospore-forming phylum, Firmicutes, to fulfill sporulation-specific roles. We describe a novel role for these enzymes in synthesizing septal PG during the sporulation-specific mode of cell division inC. difficile. Although these enzymes are directly regulated by canonical divisome components during this process,dcw-encoded PG synthases and their divisome regulators are unexpectedly dispensable for cell division during normal growth. Instead,C. difficileuses its sole bifunctional class A penicillin-binding protein (aPBP) to drive cell division, revealing a previously unreported role for this class of PG synthases as the core divisome enzyme. Collectively, our findings reveal how the emergence of endosporulation in the Firmicutes phylum was a key driver for the functional repurposing of an otherwise universally conserved cellular process such as cell division. Moreover, they indicate thatC. difficile,and likely other clostridia, assemble a divisome that differs markedly from previously studied bacteria, thus representing an attractive, unique target for therapeutic purposes.
WITHDRAWN: A Survey of CRISPR Across Pectobacterium atrosepticum
Authors: Alahmadi Ramzi Atiah
Published: August 2023

Withdrawal Statement

The authors have withdrawn their manuscript because permission to publish the genome data has not yet been obtained from the owners. Therefore, the authors do not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author.
Distinct modulation of calcium-activated chloride channel TMEM16A by a novel drug-binding site
Authors: Roh Jae Won , Gee Heon Yung , Wainger Brian , Kim Woo Kyung , Lee Wook , Nam Joo Hyun
Published: August 2023

TMEM16A is a calcium-activated chloride channel with significant role in multiple cellular processes. Several TMEM16A inhibitors have been identified; however, their binding sites and inhibitory mechanisms remain unclear. Using magnolol and honokiol, the two regioisomeric inhibitors, as chemical probes, we have identified a novel drug-binding site distinct from the pore region, in TMEM16A, which is described here. With electrophysiology, unbiased molecular docking and clustering, molecular dynamics simulations, and experimental validation with mutant cycle analysis, we show that magnolol and honokiol utilize different drug-binding sites, pore and non-pore pockets. The pore blocker utilizes amino acids crucial for chloride passage, whereas the non-pore blocker allosterically modulates the pore residues to hinder ion permeation. Among 17 inhibitors tested, 11 were pore blockers and six were non-pore blockers, indicating the importance of this newly identified non-pore pocket. Our study provides insights into drug-binding mechanism in TMEM16A together with a rationale for future drug development.
Evaluation of information flows in the RAS-MAPK system using transfer entropy measurements
Authors: Umeki Nobuhisa , Kabashima Yoshiyuki , Sako Yasushi
Published: August 2023

The RAS-MAPK system plays an important role in regulating various cellular processes, including growth, differentiation, apoptosis, and transformation. Dysregulation of this system has been implicated in genetic diseases and cancers affecting diverse tissues. To better understand the regulation of this system, we employed information flow analysis based on transfer entropy (TE) between the activation dynamics of two key elements in cells stimulated with EGF: SOS, a guanine nucleotide exchanger for the small GTPase RAS, and RAF, a RAS effector serine/threonine kinase. TE analysis allows for model-free assessment of the timing, direction, and strength of the information flow regulating the system response. We detected significant amounts of TE in both directions between SOS and RAF, indicating feedback regulation. Importantly, the amount of TE did not simply follow the input dose or the intensity of the causal reaction, demonstrating the uniqueness of TE. TE analysis proposed regulatory networks containing multiple tracks and feedback loops and revealed temporal switching in the reaction pathway primarily responsible for reaction control. This proposal was confirmed by the effects of a MEK inhibitor on TE. Furthermore, TE analysis identified the functional disorder of a SOS mutation associated with Noonan syndrome, a human genetic disease, of which the pathogenic mechanism has not been precisely known yet. TE assessment holds significant promise as a model-free analysis method of reaction networks in molecular pharmacology and pathology.

Significance

Intracellular signal processing relies on intricate regulatory networks composed of various molecular reactions. Ordinary differential equation (ODE) modeling, which proposes the reaction formula and parameter values, is a conventional approach to analyzing the mechanism of reaction networks. In contrast, statistical analyses focus on the correlations among the reactions and complement ODE modeling. Among them, TE analysis is advantageous in dealing with the regulation of the reaction by capturing the dynamics of causality between the reactions. In this study, we analyzed the TE in the cell signaling pathway. The results demonstrate the usefulness of TE assessment as a model-free network analysis method.
Baseline and treatment-emergent bedaquiline resistance in drug-resistant tuberculosis: A systematic review and meta-analysis
Authors: Perumal Rubeshan , Bionghi Neda , Nimmo Camus , Letsoalo Marothi , Cummings Matthew J. , Hopson Madeleine , Wolf Allison , Jubaer Shamim Al , Padayatchi Nesri , Naidoo Kogieleum , Larsen Michelle H. , O’Donnell Max
Published: August 2023

Rationale

Bedaquiline is a novel antimycobacterial agent for drug-resistant tuberculosis (DR-TB) and is classified as a World Health Organization (WHO) Group A drug due to its excellent clinical efficacy, high bactericidal activity, and potent sterilizing effect. Baseline and treatment-emergent bedaquiline resistance have been described but prevalence and incidence have not been reported, leading to gaps in the knowledge required to design strategies to optimize MDR-TB clinical outcomes and prevent the amplification of bedaquiline resistance.

Methods

We performed a systematic review and meta-analysis to estimate the frequency of, and mutations associated with, baseline and acquired (treatment-emergent) bedaquiline resistance in clinicalMtbisolates. Pooled estimates of bedaquiline resistance were generated by proportional meta-analysis in R version 4.2.2 using dmetar, metafor and meta packages. Resistance associated variants associated with prevalent and incident bedaquiline resistance were identified.

Results

Data from 14 studies were included; 14 and 9 studies reported on pre-treatment and acquired bedaquiline resistance, respectively. The pooled prevalence of pre-treatment bedaquiline resistance was 2.4% (95% CI 1.7 – 3.5), with significant heterogeneity across all studies (I266%, p<0.01). The pooled prevalence of treatment-emergent bedaquiline resistance was 2.1% (95% CI 1.4 - 3.0), with no significant heterogeneity across the included studies (I20%, p=0.97).

Discussion

We found a concerning frequency of bedaquiline resistance present at baseline and acquired during treatment. Urgent strategies are required to mitigate further resistance to this crucial drug.
Loss of HSC stemness identity is associated with exhaustion and hyporesponsiveness in GATA2 deficiency syndrome
Authors: Largeaud Laetitia , Fregona Vincent , Jamrog Laura , Hamelle Camille , Dufrechou Stéphanie , Prade Naïs , Sellam Esmaa , Enfedaque Pauline , Bayet Manon , Hébrard Sylvie , Bouttier Mathieu , Didier Christine , Delabesse Eric , Gerby Bastien , Pasquet Marlène , Broccardo Cyril
Published: August 2023

GermlineGATA2mutations lead to a syndrome involving both immunodeficiency and myeloid malignancies. Since GATA2 is a key player in hematopoietic initiation and development, we specify the impact of these germline mutations on hematopoietic homeostasis by generated a knock-in mouse model expressing the recurrentGata2R396Q missense mutation. These mice exhibit a hematopoietic stem and progenitor cell (HSPC) compartment profoundly impacted with increased HSC number, decreased self-renewal potential and inability to respond to acute inflammatory stimuli. Moreover, mutated HSPCs are predisposed to be hyporesponsive, as evidenced by lower interferon signaling and enrichment of inflammatory stress signatures. Furthermore, a Gata2 allelic specific expression results in a molecular and functional heterogeneity of the mutated Long Term-HSC population. Altogether, we highlight that Gata2 plays a crucial role in the ability of HSCs to perceive and respond to their environment, and that germline mutation contributes to the decline in HSC functionality.
The shape of Cannabis sativa L. seeds for the differentiation of commercial medicinal cultivars from Argentina
Authors: Torne Francisco Fernandez , Bosco Tomas , Idaszkin Yanina L. , Bigatti Gregorio , Garcés Natahiel , Lozada Mariana , González-José Rolando , Márquez Federico
Published: August 2023

The Cannabis sativa L. plant has been used since ancient times as food, a source of fiber, and medicine, resulting in crosses that led to hybridization that currently does not allow for morphological differentiation among the three varieties of the genus (sativa, indica, and ruderalis). Currently, three chemotypes are differentiated based on their cannabinoid content (THC/CBD). Starting in the year 2023, seeds of two contrasting chemotype medicinal cultivars developed by CONICET (National Council for Scientific and Technical Research) and registered with INASE (National Institute of Seeds) can be commercialized in Argentina. In a previous study, we reported a relationship between the shape of Cannabis seeds and the chemical fingerprint associated with the chemotype. The objective of this study is to morphometrically characterize the seeds of two cultivars of Cannabis sativa L. with contrasting chemotypes: type I (high THC) and type III (high CBD). For this purpose, 2D geometric morphometrics based on landmarks and semilandmarks were used, allowing for the independent study of shape and size variation. Seed size between cultivars was compared using univariate statistics of an unbiased size estimator. To understand the magnitude and direction of shape change and determine shape characters that maximize separation between cultivars, a multivariate statistical approach was employed. Seeds belonging to the Malvina cultivar (type I cultivar, THC only) had, on average, smaller size and a rounded shape, whereas seeds from the Pachamama cultivar (type III, high CBD content) had larger size and a tendency towards an elongated oval shape. The use of a discriminant function based on seed shape allowed for over 97% correct assignments between cultivars. Our results could be used for implementing seed shape as a quality and authenticity seal for registered cultivars in the Argentine Cannabis market.
A kinome-wide synthetic lethal CRISPR/Cas9 screen reveals that mTOR inhibition prevents adaptive resistance to CDK4/CDK6 blockade in HNSCC
Authors: Goto Yusuke , Koshizuka Keiichi , Ando Toshinori , Izumi Hiroki , Wu Xingyu , Ford Kyle , Feng Xiaodong , Wang Zhiyong , Arang Nadia , Allevato Michael M. , Kishore Ayush , Mali Prashant , Gutkind J. Silvio
Published: August 2023

The comprehensive genomic analysis of the head and neck cancer (HNSCC) oncogenome revealed frequent loss of p16INK4A(CDKN2A) in most HPV negative HNSCC lesions, often concomitant with amplification of the cyclin D1 (CCND1) gene locus. However, cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors as single agents have shown modest effect in the clinic, even when combined with cetuximab. The aberrant activation of PI3K/mTOR pathway is highly prevalent in HNSCC, and recent clinical trials targeting mTOR showed promising results in terms of objective responses and progression free survival. However, the clinical efficacy of mTOR inhibitors (mTORi) for advanced HNSCC patients may be limited due to intrinsic or acquired resistance. By a kinome-wide CRISPR/Cas9 screen, we identified cell cycle inhibition as a synthetic lethal target for mTORi. Combination of mTORi and palbociclib, a CDK4/6 specific inhibitor, showed strong synergism in HNSCC-derived cells in vitro and in vivo. Remarkably, we found that adaptive increase in cyclin E1 (CCNE1) expression upon palbociclib treatment underlies the rapid acquired resistance to this CDK4/6 inhibitor in HNSCC. Mechanistically, mTORi inhibits the formation of eIF4G-CCNE1mRNA complexes, with the consequent reduction in mRNA translation and CCNE1 protein expression. Our findings suggest that concomitant mTOR blockade reverts the adaptive resistance to palbociclib, thereby providing a novel multimodal therapeutic option for HNSCC patients by co-targeting mTOR and CDK4/6. Our findings may have broad implications to halt the emergence of palbociclib resistance.
Modelling Cell Shape in 3D Structured Environments: A Quantitative Comparison with Experiments
Authors: Link Rabea , Jaggy Mona , Bastmeyer Martin , Schwarz Ulrich S.
Published: August 2023

Cell shape plays a fundamental role in many biological processes, including migration, division and tissue morphogenesis, but it is not clear which shape model best predicts three-dimensional cell shape in structured environments. Here, we compare different modelling approaches with experimental data. 3D scaffolds for cell adhesion were manufactured using 3D laser nanoprinting and the shapes of single mesenchymal NIH/3T3 cells were compared by a Fourier method with surfaces that minimize area under the given adhesion and volume constraints. For the minimized surface model, we found marked differences to the experimentally observed cell shapes, which necessitated the use of more advanced shape models. We used different variants of the cellular Potts model, which effectively includes both surface and bulk contributions. The simulations revealed that the Hamiltonian with linear area energy outperformed the elastic area constraint in accurately modeling the 3D shapes of cells in structured environments. Explicit modeling the nucleus did not improve the accuracy of the simulated cell shapes. Overall, our work identifies effective methods for accurately modeling cellular shapes in complex environments, which in the future will enable the rational design of scaffolds for desired cell shapes.

Author summary

Cell shape and forces have emerged as important determinants of cell function and thus their prediction is essential to describe and control the behaviour of cells in complex environments. While there exist well-established models for the two-dimensional shape of cells on flat substrates, it is less clear how cell shape should be modeled in three dimensions. Different from the philosophy of the vertex model often used for epithelial sheets, we find that models based only on cortical tension as a constant geometrical surface tension are not sufficient to describe the shape of single cells in 3D. Therefore, we employ different variants of the cellular Potts model, where either a target area is prescribed by an elastic constraint or the area energy is described with a linear surface tension. By comparing the simulated shapes to experimental images of cells in 3D scaffolds manufactured with direct laser writing, we can identify parameters that accurately model the 3D cell shape.
Detection and characterization of a novel bat filovirus (Dehong virus, DEHV) in fruit bats
Authors: He Biao , Hu Tingsong , Yan Xiaomin , Zhang Fuqiang , Tu Changchun
Published: August 2023

Although bats are natural hosts of most filoviruses (FiVs), with the pathogenic ones frequently causing deadly haemorrhagic fevers in Africa, the viruses are difficult to trace. Recently, multiple divergent FiVs have been uncovered in China, raising concerns about their threat to public health. Here we report the detection of bat-borne FiVs at the interface of orchards between bats and humans, eventually resulting in the discovery of a novel FiV, Dehong virus (DEHV), fromRousettus leschenaultii(R. leschenaultii) bats. DEHV has the largest genome, and the phylogeny places it between the generaDianlovirusandMarburgvirus, suggesting its classification as a prototype of a new genus within the familyFiloviridae. Our study emphasizes the importance of further understanding the distribution and potential risk of FiVs in the region.
The circadian clock is a pacemaker of the axonal regenerative ability
Authors: Virgiliis Francesco De , Mueller Franziska , Palmisano Ilaria , Chadwick Jessica S , Gutierrez Lucía Luengo , Giarrizzo Angela , Yan Yuyang , Danzi Matt Christopher , Muñoz Carmen Picón , Zhou Luming , Kong Guiping , Serger Elisabeth , Hutson Thomas Haynes , Maldonado ines , Song Yayue , Scheiermann Christoph , Brancaccio Marco , Giovanni Simone Di
Published: August 2023

Peripheral nervous system injuries lead to long-term neurological disability due to limited axonal regenerative ability. Injury-dependent and -independent physiological mechanisms have provided important molecular insight into neuronal regeneration. However, whether common molecular denominators underpin both injury-dependent and independent biological processes remain unclear. Here, we performed a comparative analysis of recently generated transcriptomic datasets associated with the regenerative ability of sciatic dorsal root ganglia (DRG). Surprisingly, circadian rhythms were identified as a the most significantly enriched biological process associated with regenerative capability. We demonstrate that DRG neurons possess an endogenous circadian clock with a 24h oscillations of circadian genes and that their regenerative ability displays a diurnal oscillation in a mouse model of sciatic nerve injury. Consistently, transcriptomic analysis of DRG neurons showed a significant time-of-day dependent enrichment for processes associated with axonal regeneration, development and growth, as well as circadian associated genes, including the core clock genesBmal1andClock. Indeed, DRG-specific ablation of the non-redundant clock geneBmal1showed that it is required for regenerative gene expression, neuronal intrinsic circadian regeneration and target reinnervation. Lastly, lithium, a chrono-active compound, enhanced nerve regeneration, in wildtype but not inBmal1andCry1/2-deficient mice. Together, these data demonstrate that daily rhythms and the molecular clock fine-tune the regenerative response of DRG neurons, and that chrono-active drugs, such as lithium, are a novel potential approach to nerve repair.
Structure of the human outer kinetochore KMN network complex
Authors: Yatskevich Stanislau , Yang Jing , Bellini Dom , Zhang Ziguo , Barford David
Published: August 2023

Faithful chromosome segregation requires robust, load-bearing attachments of chromosomes to the mitotic spindle, a function accomplished by large macromolecular complexes termed kinetochores. In most eukaryotes, the constitutive centromere-associated network (CCAN) complex of the inner kinetochore recruits to centromeres the ten-subunit outer kinetochore KMN network, which comprises the KNL1C, MIS12C and NDC80C complexes. The KMN network directly attaches CCAN to microtubules through MIS12C and NDC80C. Here, we determined a high-resolution cryo-EM structure of the human KMN network. This showed an intricate and extensive assembly of KMN subunits, with the central MIS12C forming rigid interfaces with NDC80C and KNL1C. The redundancy and strength of inter-subunit connections explains how KMN withstands strong forces applied during chromosome segregation. We also observed that unphosphorylated MIS12C exists in an auto-inhibited state that suppresses its capacity to interact with CCAN. Ser100 and Ser109 of the N-terminal segment of the MIS12C subunit Dsn1, two key targets of Aurora B kinase, directly stabilize this auto-inhibition. Our work provides a molecular mechanism for how selectively relieving this auto-inhibition through Ser100 and Ser109 phosphorylation would restrict outer kinetochore assembly to functional centromeres during cell division.