bluuebunny/crossref_metadata_2025 huggingface dataset:
novoStoic2.0: An integrated framework for pathway synthesis, thermodynamic evaluation, and enzyme selection
Authors: Upadhyay Vikas , Anand Mohit , Maranas Costas D.
Published: September 2024

Computational pathway design and retro-biosynthetic approaches can facilitate the development of innovative biochemical production routes, biodegradation strategies, and the funneling of multiple precursors into a single bioproduct. However, effective pathway design necessitates a comprehensive understanding of biochemistries, enzyme activities, and thermodynamic feasibility. Herein, we introduce novoStoic2.0, an integrated platform that combines tools for estimating overall stoichiometry, designing de novo synthesis pathways, assessing thermodynamic feasibility, and selecting enzymes. novoStoic2.0 offers a unified web-based interface as a part of the AlphaSynthesis platform (http://novostoic.platform.moleculemaker.org/) tailored for the synthesis of thermodynamically viable pathways as well as the selection of enzymes for re-engineering required for novel reaction steps. We exemplify the utility of the platform to identify novel pathways for hydroxytyrosol synthesis, which are shorter than the known pathways and require reduced cofactor usage. In summary, novoStoic2.0 aims to streamline the process of pathway design contributing to the development of sustainable biotechnological solutions.
On the diversity of nematode antagonists in an agricultural soil, and their steerability by root-knot nematode density and cover crops
Authors: Cazzaniga Sara G. , Belliard Philippe , Van Steenbrugge Joris J,M. , Van den Elsen Sven , Lombaers Carin , Visser Johnny , Macia Vicente Jose , Postma Joeke , Mommer Liesje , Helder Johannes
Published: October 2024

Plant-parasitic nematodes are harmful pathogens for many agricultural crops. Within this category, root-knot nematodes (RKN, Meloidogyne spp.) are worldwide regarded as the most impactful because of their wide geographical distribution and their polyphagous nature. Host plant resistances against RKN have been successfully introduced in a few crops only. As the use of nematicides is becoming increasingly restricted because of environmental and human health concerns, there is a need for alternative strategies to control RKN. One such approach is the stimulation of local nematode antagonists. We investigated this in an experimental field setting with two main variables: density of the Columbia root-knot nematode Meloidogyne chitwoodi, and the type of cover of crop. For each of the three M. chitwoodi densities, the effects of ten cover crop treatments were tested on both the resident (DNA) and the active (RNA) fractions of the bacterial and fungal communities. In our analyses, we focussed on changes in the abundance of plant-parasitic nematode antagonists. From the eight bacterial and 26 fungal genera known from global literature to harbour potential antagonists of plant-parasitic nematodes, we detected respectively five and 14 genera in our agricultural field. Among the bacterial genera, four included bacterial species for which nematode antagonism has been documented. The fungal genera included facultative nematode parasites (e.g., Arthrobotrys spp.), endophytes strengthening host defences (e.g., Acremonium spp.), as well as multiple obligatory nematophagous species. This study revealed that conventionally managed arable fields may harbour an unexpectedly high diversity of nematode antagonists. Multiple antagonists were stimulated by cover crops in a cover crop-specific manner, and, to a lesser extent, by increased RKN densities. The richness in putative nematode antagonists did not translate into M. chitwoodi suppression probably because most antagonists have a facultative nematophagous lifestyle and will only predate nematodes under poorer nutritional conditions.
Outer membrane changes enable evolutionary escape from bacterial predation
Authors: Mridha Subham , Loose Sarah-Luise , Mattmann Ove Ch. , Isele Saskia , Kümmerli Rolf , Huwiler Simona G.
Published: September 2024

To combat antimicrobial resistant pathogens, natural predatory bacteria, likeBdellovibrio bacteriovorus, represent potential alternatives.B. bacteriovoruscould be particularly potent as it kills a broad range of human bacterial pathogens, however, it remains unclear whether prey can evolve genetically-determined resistance against predation. Here, we show that the model bacteriumEscherichia coliK-12 consistently evolves resistance againstB. bacteriovorusduring experimental evolution. Selection for resistance scaled positively with predation pressure and was widespread after two cycles of predator exposure. Like antibiotics, predation resistance was costly, manifesting in a trade-off between predation resistance and fitness in the absence of predators. Genetic analysis identified changes in outer membrane porin OmpF as common resistance mechanism, while a mutation in cell envelope lipopolysaccharide-modifying enzyme WaaF was rarer but also conferred predation resistance. Our study uncovers evolutionary and mechanistic aspects of prey escape from predation, generating important knowledge on predator-prey interactions and to advance sustainable treatments.
Lymphatic dynamics visualized by native transient hypoxia imaging in areas of tissue damage, edema and in sentinel lymph nodes
Authors: Ochoa Marien I. , Reed Matthew S. , Zeng Weifeng , Poore Samuel O. , Hasan Tayyaba , Pogue Brian W.
Published: October 2024

Imaging lymphatic compartments and their function has always been challenging, yet this capability is key to understanding the dynamics of immune response and lymph dysfunction in disease states. This study reports the first ever visualization of murine lymphatic pumping and function imaged from the inherent transient hypoxia that occurs within the lymph ducts and nodes. The lymphatic system appears as one of the few naturally hypoxic areas in vivo. Hypoxia in lymphatics is detected via delayed fluorescence (DF) of endogenous protoporphyrin IX (PpIX), enabling real-time imaging. Lymph nodes and their function were localized by hypoxia transcutaneous imaging and in surgically exposed nodes, followed by correlation of localization to indocyanine green (ICG) local injection. The lymphatic pumping frequency was altered through progressive damage from mild, moderate, and severe wound injuries, and hypoxia appeared readily in the sentinel lymph nodes near tumor regions. Cyclical pumping was observed at sites of edema and in nodes near tumors. Control data from uninjured anesthetized mice showed little lymphatic contrast, whereas awake mice exhibited hypoxia localized to lymph nodes. Unlike contrast injection-based regional lymph node imaging by ICG or MRI, DF hypoxia imaging appears to provide a natural whole-body contrast mechanism, highlighting its potential for visualizing lymphatic function and associated hypoxia dynamics. The value for localization of sentinel lymph nodes and for allowing for visualization of damaged lymph has very practical potential applications.
Improved robustness to gene tree incompleteness, estimation errors, and systematic homology errors with weighted TREE-QMC
Authors: Han Yunheng , Molloy Erin K.
Published: October 2024

Summary methods are widely used to reconstruct species trees from gene trees while accounting for incomplete lineage sorting; however, it is increasingly recognized that their accuracy can be negatively impacted by incomplete and/or error-ridden gene trees. To address the latter, Zhang and Mirarab (2022) leverage gene tree branch lengths and support values to weight quartets within the popular summary method ASTRAL. Although these quartet weighting schemes improved the robustness of ASTRAL to gene tree estimation error, implementing the weighting schemes presented computational challenges, resulting in the authors abandoning ASTRAL’s original search algorithm (i.e., computing an exact solution within a constrained search space) in favor of search heuristics (i.e., hill climbing with nearest neighbor interchange moves from a starting tree constructed via randomized taxon addition). Here, we show that these quartet weighting schemes can be leveraged within the Quartet Max Cut framework of Snir and Rao (2010), with only a small increase in time complexity compared to the unweighted algorithm, which behaves more like a constant factor in our simulation study. Moreover, our new algorithm, implemented within the TREE-QMC software, was highly competitive with weighted ASTRAL, even outperforming it in terms of species tree accuracy on some challenging model conditions, such as large numbers of taxa. In comparing unweighted and weighted summary methods on two avian data sets, we found that weighting quartets by gene tree branch lengths improves their robustness tosystematichomology errors and is as effective as removing the impacted taxa from individual gene trees or removing the impacted gene trees entirely. Lastly, our study revealed that TREE-QMC is highly robust to high rates of missing data and is promising as a supertree method. TREE-QMC is written in C++ and is publicly available on Github:https://github.com/molloy-lab/TREE-QMC
Hastened fusion-dependent endosomal escape improves activity of delivered enzyme cargo
Authors: Vázquez Maldonado Angel Luis , Chen Teresia , Rodriguez Diego , Zoltek Madeline , Schepartz Alanna
Published: September 2024

There is enormous interest in strategies to efficiently traffic biologics–proteins, nucleic acids, and complexes thereof–into the mammalian cell cytosol and internal organelles. Not only must these materials reach the appropriate cellular locale fully intact and in therapeutically relevant concentrations, they must also retain activity upon arrival. The question of residual activity is especially critical when delivery involves exposure to the late endocytic pathway, whose acidic lumenal environment can denature and/or degrade internalized material. ZF5.3 is a compact, stable, rationally designed mini-protein that efficiently escapes intact from late endocytic vesicles, with or without covalently linked protein cargo. Here, using insights from mechanistic studies on the pathway of endosomal escape and classic principles of zinc(II) bioinorganic chemistry, we re-designed the sequence of ZF5.3 to successfully alter the timing (but not the efficiency) of endosomal escape. The new mini-protein we describe, AV5.3, escapes earlier than ZF5.3 along the endocytic pathway with no loss in efficiency, with or without enzyme cargo. More importantly, earlier endosomal escape translates into higher enzymatic activity of a pH-sensitive enzyme upon arrival in the cytosol. Delivery of the pH-sensitive enzyme dihydrofolate reductase (DHFR) with AV5.3 results in substantial catalytic activity in the cytosol, whereas delivery with ZF5.3 does not. The activity of AV5.3-DHFR upon delivery is sufficient to rescue a genetic DHFR deletion in CHO cells. This work provides evidence that programmed trafficking through the endosomal pathway is a viable strategy for the efficient cytosolic delivery of therapeutic proteins.

Abstract Figure
Lysophospholipid headgroup size, and acyl chain length and saturation differentially affect vacuole acidification, Ca2+transport, and fusion
Authors: Zhang Chi , Feng Yilin , Calderin Jorge D. , Balutowski Adam , Ahmed Razeen , Knapp Charlie , Fratti Rutilio A.
Published: September 2024

SNARE-mediated membrane fusion is regulated by the lipid composition of the engaged bilayers. Lipid composition impacts fusion through direct protein lipid interactions or through modulating the physical properties of membranes at the site of contact, including the induction of positive curvature by lysophospholipids (LPLs). The degree of positive curvature induced is due to the length and saturation of the single acyl chain in addition to the size of the head group. Here we examined how yeast vacuole fusion and ion transport were differentially affected by changes in lysolipid properties. We found that lysophosphatidylcholine (LPC) with acyl chains containing 14-18 carbons all inhibited fusion with IC50values ranging from ∼40-120 µM. The monounsaturation of LPC-18:1 had no effect when compared to its saturated counterpart LPC-18:0. On the other hand, head group size played a more significant role in blocking fusion as lysophosphatidic acid (LPA)-18:1 failed to fully inhibit fusion. We also show that both Ca2+uptake and SNARE-dependent Ca2+efflux was sensitive to changes in the acyl chain length and saturation of LPCs, while LPA only affected Ca2+efflux. Finally, we tested these LPLs on vacuole acidification by the V-ATPase. This showed that LPC-18:0 could fully inhibit acidification whereas other LPCs had moderate effects. Again, LPA had no effect. Together these data suggest that the effects of LPLs were due to a combination of head group size and acyl chain length leading to a range in degree of positive curvature.
irCLIP-RNP and Re-CLIP reveal patterns of dynamic protein associations on RNA
Authors: Ducoli Luca , Zarnegar Brian J. , Porter Douglas F. , Meyers Robin M. , Miao Weili , Riley Nicholas M. , Srinivasan Suhas , Jackrazi Leandra V. , Yang Yen-Yu , Li Zhouxian , Wang Yinsheng , Bertozzi Carolyn R. , Flynn Ryan A. , Khavari Paul A.
Published: September 2024

RNA binding proteins (RBPs) control varied processes, including RNA splicing, stability, transport, and translation1-3. Dysfunctional RNA-RBP interactions contribute to the pathogenesis of human disease1,4,5, however, characterizing the nature and dynamics of multiprotein assemblies on RNA has been challenging. To address this, non-isotopic ligation-based ultraviolet crosslinking immunoprecipitation6was combined with mass spectrometry (irCLIP-RNP) to identify RNA-dependent associated proteins (RDAPs) co-bound to RNA with any RBP of interest. irCLIP-RNP defined landscapes of multimeric protein assemblies on RNA, uncovering previously unknown patterns of RBP-RNA associations, including cell-type-selective combinatorial relationships between RDAPs and primary RBPs. irCLIP-RNP also defined dynamic RDAP remodeling in response to epidermal growth factor (EGF), uncovering EGF-induced recruitment of UPF1 adjacent to HNRNPC to effect splicing surveillance of cell proliferation mRNAs. To identify the RNAs simultaneously co-bound by multiple studied RBPs, a sequential immunoprecipitation irCLIP (Re-CLIP) method was also developed. Re-CLIP confirmed binding relationships seen in irCLIP-RNP and detected simultaneous HNRNPC and UPF1 co-binding on RND3 and DDX3X mRNAs. irCLIP-RNP and Re-CLIP provide a framework to identify and characterize dynamic RNA-protein assemblies in living cells.
Loss of Ing4 enhances hematopoietic regeneration in multipotent progenitor cells
Authors: Anderson Georgina A. , Hernandez Marco , Quinde Carlos Alfaro , Thompson Zanshé , Rodriguez Melanie , Binder-Blaser Vera , Taylor Alison M. , Kathrein Katie L.
Published: September 2024

Despite its critical role in survival, many aspects of hematopoiesis remain unresolved. In the classical model of the hematopoietic program, quiescent hematopoietic stem cells (HSCs) sit at the top of the hematopoietic hierarchy, with the ability to self-renew and differentiate as needed. HSCs give rise to more proliferative progenitor cells, which possess multipotent potential, but have largely or completely lost self-renewal capabilities. Here, we have identified the tumor suppressor, Inhibitor of Growth 4 (Ing4), as a critical regulator of multipotent progenitor (MPP) homeostasis. In the absence of Ing4, we show that MPPs express a transcriptional program of hematopoietic activation, yet they remain quiescent with low levels of reactive oxygen species. Functionally, Ing4-deficient MPPs are capable of robust regeneration following competitive bone marrow transplantation, resulting in substantially higher blood chimerism compared to wild-type MPPs. These data suggest Ing4 deficiency promotes a poised state in MPPs, quiescent, but transcriptionally primed for activation, and capable of converting the poised state into robust repopulation upon stress. Our model provides key tools for further identification and characterization of pathways that control quiescence and regeneration in MPPs.
Photosymbiosis Shaped Animal Genome Architecture and Gene Evolution as Revealed in Giant Clams
Authors: Li Ruiqi , Leiva Carlos , Lemer Sarah , Kirkendale Lisa , Li Jingchun
Published: October 2024

Symbioses are major drivers of organismal diversification and phenotypic innovation. However, how long-term symbioses shape whole genome evolution in metazoans is still underexplored. Here, we used a giant clam (Tridacna maxima) genome to demonstrate how symbiosis has left complex signatures in an animal’s genome. Giant clams thrive in oligotrophic waters by forming a remarkable association with photosymbiotic dinoflagellate algae. Genome-based demographic inferences uncovered a tight correlation betweenT. maximaglobal population change and major paleoclimate and habitat shifts, highlighting how abiotic and biotic factors dictateT. maximamicroevolution. Comparative analyses revealed unique symbiosis-driven genomic features, including expansion and contraction of immunity-related gene families and a large proportion of lineage-specific genes. Strikingly, about 70% of the genome is composed of repetitive elements, especially transposable elements, most likely resulting from a symbiosis-adapted immune system. This work greatly enhances our understanding of genomic drivers of symbiosis that underlie metazoan evolution and diversification.
Investigating the integration of sustainable food initiatives in healthcare institutions in Ontario, Canada: A grey literature scoping review protocol
Authors: Blank Lisa L. , Milano Alyssa D. , Andrade Lesley , Kirkpatrick Sharon I.
Published: September 2024

Introduction

The climate emergency and other sustainability challenges interact to threaten human and planetary health. Efforts to improve the sustainability of food initiatives within healthcare institutions could mitigate these threats by addressing the four pillars of sustainability: health, social, economic, and environmental. Understanding current initiatives to incorporate sustainability into food programs and the sustainability pillars that guide those initiatives is important to inform priorities for action.

This scoping review was undertaken to investigate the extent to which major healthcare institutions in Ontario, Canada have publicly committed to, discussed, planned, and/or implemented sustainable food initiatives.

Methods and Analysis

Steps are based on guidance from the Joanna Briggs Institute and Arksey & O’Malley. First, the current strategic plans of 57 healthcare institutions in Ontario, Canada, will be retrieved from their websites and used to examine whether they include any commitments to or discussion, planning, and/or implementation of relevant initiatives. The healthcare institution websites, along with those of selected sustainability organizations, will be searched for grey literature from 2015 to 2024 describing sustainable food initiatives within these institutions. Documents will be screened for eligibility by two researchers. Data related to the incorporation of sustainable food into institutional food programs, and the sustainability pillars addressed, will be extracted by one researcher, with 10% of entries verified by a second researcher. The data will be synthesized to summarize publicly reported progress toward integrating sustainable food into healthcare institutions.

Ethics and Dissemination

This review will use publicly available grey literature with no expectation of privacy and no research participants; therefore, no ethics clearance is required.

Results will be shared with stakeholders in sustainability organizations, as well as at relevant conferences and in peer-reviewed journals, such as the Healthy Cities Conference and the Journal of Canadian Food Studies.

This protocol is registered on the Open Science Framework and can be accessed at the following URL:https://doi.org/10.17605/OSF.IO/CU9P6
Mapping the Mind in the Virtual Metaverse: An Initial In-Depth Thematic Exploration of Youth Mental Health within VRChat
Authors: Um Tevin S , Ownby Raymond L , Chou Shinnyi
Published: October 2024

Background

VRChat is a virtual reality (VR) social video game where users play a personalized avatar and socialize with others in community-built worlds. Its immersive nature and diverse community may impact youth users’ mental health, yet no research has explored this. This study is a thematic exploration of VRChat’s effect on youth mental health.

Methods

Semi-structured interviews were conducted with 20 active VRChat users aged 18-24 years. Interviews were transcribed and analyzed using grounded theory principles to identify themes related to VRChat use and mental health.

Results

Nine major themes emerged: Frequency of use; purpose of use; positive effects on mental health; negative effects on mental health; social connection and COVID-19; anonymity; cyberbullying and toxic behaviors; addiction; and recommendations for a safer environment. Positive effects identified included enhanced social connection, creative expression, and self-confidence. Negative effects included addiction, cyberbullying, predatory behaviors, and worsening of existing mental illnesses. The anonymity of VRChat created a paradoxical environment that can foster both genuine self-expression and toxic behaviors. During the COVID-19 pandemic, VRChat served as an important substitute for real-life social interactions. Participants also provided recommendations for developers, parents, and clinicians to promote a safer VRChat experience.

Conclusion

The immersive and interactive social experience offered by VRChat is conducive for positive effects on youth users’ mental health, such as social connectivity and self-expression. However, it also presents risks such as addiction and toxic behaviors. Given the interactive nature of VRChat, further research may elucidate its potential as a psychotherapeutic intervention for youth mental health.
A repository of Ogden syndrome patient derived iPSC lines and isogenic pairs by X-chromosome screening and genome-editing
Authors: Wesely Josephine , Rusielewicz Tom , Chen Yu-Ren , Hartley Brigham , McKenzie Dayna , Yim Matthew K. , Maguire Colin , Bia Ryan , Franklin Sarah , Makwana Rikhil , Marchi Elaine , Nikte Manali , Patil Soha , Sapar Maria , Moroziewicz Dorota ,   NYSCF Global Stem Cell Array® Team, Bauer Lauren , Lee Jeannie T. , Monsma Frederick J. , Paull Daniel , Lyon Gholson J.
Published: September 2024

Amino-terminal (Nt-) acetylation (NTA) is a common protein modification, affecting 80% of cytosolic proteins in humans. The human essential gene,NAA10,encodes the enzyme NAA10, as the catalytic subunit for the N-terminal acetyltransferase A (NatA) complex, including the accessory protein, NAA15. The first human disease directly involvingNAA10was discovered in 2011, and it was named Ogden syndrome (OS), after the location of the first affected family residing in Ogden, Utah, USA. Since that time, other variants have been found inNAA10andNAA15.Here we describe the generation of 31 iPSC lines, with 16 from females and 15 from males. This cohort includes CRISPR-mediated correction to the wild-type genotype in 4 male lines, along with editing one female line to generate homozygous wild-type or mutant clones. Following the monoclonalizaiton and screening for X-chromosome activation status in female lines, 3 additional pairs of female lines, in which either the wild type allele is on the active X chromosome (Xa) or the pathogenic variant allele is on Xa, have been generated. Subsets of this cohort have been successfully used to make cardiomyocytes and neural progenitor cells (NPCs). These cell lines are made available to the community via the NYSCF Repository.
Optimal Frequency for Seizure Induction with Electroconvulsive Therapy and Magnetic Seizure Therapy
Authors: Peterchev Angel V. , Deng Zhi-De , Sikes-Keilp Christopher , Feuer Elyssa C. , Rosa Moacyr A. , Lisanby Sarah H.
Published: October 2024

Electroconvulsive therapy (ECT) and magnetic seizure therapy (MST) are effective in the treatment of medication-resistant depression. Determining the stimulus frequency resulting in the lowest seizure threshold could produce fewer adverse effects by reducing the overall stimulus intensity. To determine the optimal frequency for seizure induction, four male rhesus macaques were titrated with an increasing number of pulses at fixed frequencies ranging from 5 to 240 pulses per second (pps) using ultrabrief-pulse right-unilateral ECT and circular-coil-on-vertex MST. The seizure threshold dependence on stimulus frequency was similar for ECT and MST. While higher frequencies required progressively shorter trains to induce a seizure, the middle frequency range was associated with the fewest pulses (and hence the least charge and energy), with a minimum at 16 pps and similarly low thresholds for 10 and 25 pps. The number of pulses at seizure threshold increased markedly at lower and higher frequencies. The lowest stimulus frequencies, 5 and 10 pps, were associated with the greatest ictal power measured by electroencephalography. While this study did not assess efficacy or side effects, the results highlight the significance of stimulus frequency for seizure induction, suggest efficient titration schedules that minimize exposure to the electrical stimulus, and can inform studies to assess the impact on clinical outcomes.
Reversible tuning of membrane sterol levels by cyclodextrin in a dialysis setting
Authors: Alsayyah Cynthia , Rodrigues Emmanuel , Hach Julia , Renne Mike F. , Ernst Robert
Published: October 2024

Large unilamellar vesicles (LUVs) are popular membrane models for studying the impact of lipids and bilayer properties on the structure and function of individual membrane proteins. The functional reconstitution of transmembrane in liposomes can be challenging, especially, if the hydrophobic thickness of the protein does not match the thickness of the surrounding lipid bilayer. During the reconstitution procedure Such hydrophobic mismatch causes low yields and protein aggregation, which are exacerbated in sterol-rich membranes featuring low membrane compressibility. Here, we explore new approaches to reversibly tune membrane sterol contents proteoliposomes after their formation. Both cholesterol delivery and extraction are mediated by methyl-β-cyclodextrin in a dialysis setting, which maintains (proteo)liposomes in a confined compartment. This makes it possible to reversibly tune the cholesterol level without losing membrane material simply by placing the dialysis cassette in a new bath containing either empty or cholesterol-loaded methyl-β-cyclodextrin. Cholesterol delivery and removal is monitored with the solvatochromic probe C-Laurdan, which reports on lipid packing. Using Förster-resonance energy transfer, we show that cholesterol delivery to proteoliposomes induces the oligomerization of a membrane property sensor, while the subsequent removal of cholesterol demonstrates the full reversibility. We propose that tuning membrane compressibility by methyl-β-cyclodextrin-meditated cholesterol delivery and removal in a dialysis setup provides a new handle to study its impact on membrane protein structure, function, and dynamics.

Statement of significance

Generating complex, sterol-rich, biomimetic membranes for studying the structure and function of reconstituted membrane proteins is challenging. As an important step towards asymmetric, sterol-rich, complex model membrane systems, we have established a procedure to control the membrane sterol level of liposomes and proteoliposomes using methyl-β-cyclodextrin in a dialysis setup. We demonstrate the feasibility of this approach by C-Laurdan and dehydroergosterol fluorescence spectroscopy and gain control over the membrane sterol content. We explore several parameters that affect the rate of cholesterol delivery and show that the oligomerization of a membrane property sensor, which is on the unfolded protein response sensor protein Ire1, is controlled by the sterol content of the surrounding lipid bilayer.
Exploring the Correlation Between UVB Sensitivity and SLE Activity: Insights into UVB-Driven Pathogenesis in Lupus Erythematosus
Authors: He Jiayu , Guo Yuanning , Chen Jiamin , Xu Jinhua , Zhu Xiaohua
Published: October 2024

Lupus erythematosus (LE) comprises various autoimmune inflammatory diseases, with significant overlap between cutaneous LE (CLE) and systemic LE (SLE). A key feature of both CLE and SLE is UV photosensitivity, particularly in UV-exposure-related skin inflammation. Despite this, reliable and objective UVB photosensitivity indicators closely correlating with LE activity have yet to be identified, and the underlying cellular and molecular mechanisms linking UVB sensitivity with LE onset and progression remain unclear. We discovered that ultraviolet B minimal erythema dose (UVB-MED), a quantitative photosensitivity measure, is a significant and independent risk factor for SLE activity, demonstrating a negative correlation with SLEDAI (r = -0.58, P < 0.0001). Comprehensive transcriptomic analyses of large-scale CLE and SLE samples revealed more pronounced and extensive UVB-response gene dysregulation in skin tissues compared to blood. Additionally, 14 lupus activity-correlated, UVB-response genes (UVBACGs) were identified, including eight interferon-stimulated genes (IRF7, ISG20, ISG15, IFI44, IFITM1, MX1, LY6E, OASL) and others (JUN, PTTG1, HLA-F, CAV1, HOPX, RPL3), with dysregulation evident in both skin and blood cells, particularly immunocytes. Conventional LE therapies were found to be associated with these genes and may potentially regulate them, thereby contributing to therapeutic effects. These findings highlight the role of UVB in triggering autoimmune inflammation in the skin, which may subsequently spread to systemic inflammation via immune cells and factors. UVBACGs play a critical role in this process and may serve as targets for precise LE therapies, providing insight into the link between UVB photosensitivity and LE progression.
Real-time affinity measurements of proteins synthesized in cell-free lysate using fluorescence correlation spectroscopy
Authors: Liu Chao , Hoang-Phou Steven A. , Ye Congwang , Laurence Emma J. , Laurence Matthew J. , Fong Erika J. , Hammond Nikki M. , Dillon Vannest B. , Watkins Nicholas N. , Laurence Ted A. , Coleman Matthew A.
Published: October 2024

Rapid, high throughput measurements of biomolecular interactions are essential across medicine and bioscience. Traditional methods for affinity-screening proteins require a long and costly process involving cell-based expression, purification, and titration of multiple concentrations to arrive at a binding curve. In contrast, we have developed a fast and simple approach that yields a wealth of information about the expression of the protein and its binding characteristics, all in a “one-pot reaction” and done in under several hours without the need for protein purification. The method uses cell-free protein synthesis to produce the protein of interest in the presence of its binding partner, while simultaneously using fluorescence correlation spectroscopy (FCS) to measure the increasing concentration of the protein and its binding to the binding partner. We characterize the sensitivity limits of this method by measuring the binding between the green fluorescent protein (GFP) and a low picomolar-affinity anti-GFP antibody and found that we can quantifyKDs down to the high picomolar to low-nanomolar range. We further demonstrate the method in a potentially ultrahigh-throughput sample format, in which FCS measurements are collected inside microcapsules. This work lays the foundation for a platform aimed at production and in situ affinity screening of thousands of different proteins.
Transcriptomic signatures of host immune responses in aphthous ulcers, the earliest lesions of Crohn’s disease, suggest that a bacterial invasive challenge, rather than global dysbiosis, is the initiating factor
Authors: Whiley Phillip J , Dixit Ojas V A , Gupta Mukta Das , Patel Hardip , Zhao Guoyan , Connor Susan J , Summers Kim M , Hume David A , Pavli Paul , O’Brien Claire L
Published: September 2024

Crohn’s disease is a chronic, transmural inflammatory disease of the human gut. Changes in the faecal microbial composition and a reduction in species diversity (dysbiosis) are consistent features in studies of Crohn’s disease patients, but whether dysbiosis is a cause or consequence of inflammation remains unresolved. Genetic susceptibility also plays a role in the development of Crohn’s disease and has been linked to genes involved in recognition of intestinal bacteria by cells of the mononuclear phagocyte system. The earliest visible lesions in Crohn’s disease are aphthous ulcers, overlying Peyer’s patches and lymphoid follicles. To identify mechanisms underlying the earliest stages of disease initiation we compared gene expression in aphthous ulcers, Peyer’s patches, inflamed and endoscopically normal mucosa from patients and controls using total RNA-seq.

The resulting data was subjected to network analysis to identify co-regulated gene expression signatures of cell types and processes. These results were compared to recent single cell RNA-seq analysis of intestinal macrophages in normal and diseased mucosa. The analysis of aphthous ulcers revealed signatures of epithelial stress and antimicrobial defence, plasma cell activation and immunoglobulin production, monocyte recruitment, inflammatory gene expression and induction of interferon-γ and downstream target genes. These signatures were not present in the normal appearing mucosa adjacent to aphthous ulcers which were similar to healthy control mucosa.

We conclude that the initial lesion in Crohn’s disease arises from an invasive bacterial challenge leading to intense activation of multiple host defence pathways rather than the breakdown of epithelial barrier integrity and widespread bacterial translocation.
Cryo-EM visualizes multiple steps of dynein activation pathway
Authors: Kendrick Agnieszka , Nguyen Kendrick , MA Wen , Karasmanis Eva , Amaro Rommie E , Reck-Peterson Samara , Leschziner Andres
Published: October 2024

Cytoplasmic dynein-1 (dynein) is an essential molecular motor controlled in part by autoinhibition. We recently identified a structure of partially autoinhibited dynein bound to Lis1, a key dynein regulator mutated in the neurodevelopmental disease lissencephaly. This structure provides an intermediate state in dynein activation pathway; however, other structural information is needed to fully explain Lis1 function in dynein activation. Here, we used cryo-EM and samples incubated with ATP for different times to reveal novel conformations that we propose represent intermediate states in the dynein activation pathway. We solved sixteen high-resolution structures, including seven distinct dynein and dynein-Lis1 structures from the same sample. Our data also support a model in which Lis1 relieves dynein autoinhibition by increasing its basal ATP hydrolysis rate and promoting conformations compatible with complex assembly and motility. Together, this analysis advances our understanding of dynein activation and the contribution of Lis1 to this process.
Natural variations in maternal behaviour shape trait anxiety and hippocampal neurogenesis in offspring
Authors: Grieco Fabio , van Gelderen Pauline , Larrieu Thomas , Toni Nicolas
Published: September 2024

Understanding the early origins of emotional traits and baseline anxiety is crucial for the development of personalized medicine in mood disorders. While previous research primarily focused on extreme conditions like chronic maternal deprivation, in the present study, we investigated how natural variations in maternal care influence anxiety-related behaviour and neurogenesis in C57BL/6J offspring in a longitudinal manner. We observed that mothers engaging in low maternal care (LMC) displayed lower adult neurogenesis in both the olfactory bulb and the dentate gyrus of the hippocampus compared to high maternal care (HMC) mothers. We then observed that LMC-reared pups exhibited increased anxiety-related behaviour at postnatal day (PND) 5, 9, and 22. Furthermore, maternal behaviour induced the development of emotional individuality at early stage. This mood-related phenotype in LMC-reared offspring was associated with decreased neurogenesis after weaning at PND24. In another group of litters, we further examined neurogenesis at an earlier age (PND9) and already found a reduction in the population of adult neural progenitor cells and cell proliferation in the subgranular zone of the dentate gyrus of LMC-reared pups. These results highlight that natural variations in early life experiences such as maternal care, shape long-term brain plasticity and behaviour in offspring. This underscores the relevance of maternal care and adult neurogenesis in shaping personality-like traits related to mood disorders.
Molecular and Neural Circuit Mechanisms Underlying Sexual Experience-dependent Long-Term Memory in Drosophila.
Authors: Sun Dongyu , Zhang Xiaoli , Miao Hongyu , Zhang Tianmu , Kim Woo Jae
Published: September 2024

The neural and molecular underpinnings of sexual experience-dependent long-term memory (SELTM) in male Drosophila melanogaster remain poorly understood, despite its significance for reproductive success. Here, we dissect the role of a specific class of neurons, termed 'Yuelao' (YL) neurons, in the formation of SELTM and the associated molecular pathways. We employed a combination of genetic manipulations, immunohistochemistry, and behavioral assays in Drosophila to investigate the function of YL neurons and their molecular interactors in SELTM. Utilizing the RNA sequencing data and transgenic tools, we delineated the neural circuits involved in taste and pheromone processing relevant to SELTM. Our findings reveal that the YL neurons, expressing the Orb2 scaffolding protein, are indispensable for the formation of SELTM following sexual experience. These neurons are regulated by the neuromodulator short neuropeptide F (sNPF) and its receptor (sNPF-R), which modulate glutamate release via NMDAR2. We demonstrate that sexual experience triggers synaptic plasticity in YL neurons, characterized by an increase in dendritic and presynaptic terminal areas, and a decrease in intracellular calcium levels. Furthermore, we show that YL neurons are specialized for the generation of appetitive sexual experience-dependent memory and project to brain regions implicated in memory formation. Our study uncovers the YL neurons as a key neural substrate for SELTM in Drosophila, shedding light on the molecular and circuit mechanisms that mediate the formation of long-term memories following sexual experience. These findings provide novel insights into the neural basis of taste-related memory and have broader implications for understanding the interplay between experience, memory formation, and behavior.
Stiffening Symphony of Aging: How Senescent Osteocytes Lose Their Elastic Rhythm
Authors: Tilton Maryam , Weivoda Megan , Potes Maria Astudillo , Gingery Anne , Liu Alan Y. , Tchkonia Tamara , Lu Lichun , Kirkland James L.
Published: October 2024

Senescent osteocytes are key contributors to age-related bone loss and fragility; however, the impact of mechanobiological changes in these cells remains poorly understood. This study provides a novel analysis of these changes in primary osteocytes following irradiation-induced senescence. By integrating sub-cellular mechanical measurements with gene expression analyses, we identified significant, time-dependent alterations in the mechanical properties of these cells. Increases in SA-β-Gal activity and p16Ink4a expression levels confirmed the senescence status post-irradiation. Key findings include a time-dependent increase in cytoskeletal Young’s modulus and altered viscoelastic properties of the plasma membrane, affecting the contractility of primary osteocytes. Additionally, we observed a significant increase in Sclerostin (Sost) expression 21 days post-irradiation. These mechanobiological changes may impair osteocyte mechanosensation and mechanotransduction, contributing to bone fragility. This is the first study to time-map senescence-associated mechanical changes in the osteocyte cytoskeleton. Our findings highlight the potential of biophysical markers as indicators of cellular senescence, providing more specificity than traditional, variable biomolecular markers. We believe these results support biomechanical stimulation as a potential therapeutic strategy to rejuvenate aging osteocytes and enhance bone health.
Levodopa impairs lysosomal function in sensory neurons in vitro
Authors: Olaoye Oyedele J. , Aksoy Asya Esin , Hyytiäinen Santeri V. , Narits Aia A. , Hickey Miriam A.
Published: October 2024

Parkinson’s disease (PD) is the second-most common neurodegenerative disease world-wide. Patients are diagnosed based upon movement disorders, including bradykinesia, tremor and stiffness of movement. However, non-motor signs, including constipation, rapid eye movement sleep behavior disorder, smell deficits and pain are well recognized. Peripheral neuropathy is also increasingly recognized, as the vast majority of patients show reduced intraepidermal nerve fibers, and sensory nerve conduction and sensory function is also impaired. Many case studies in the literature show that high-dose levodopa, the primary drug used in the treatment of PD, may exacerbate neuropathy, thought to involve levodopa’s metabolism to homocysteine. Here, we treated primary cultures of dorsal root ganglia and a sensory neuronal cell line with levodopa to examine effects on cell morphology, mitochondrial content and physiology, and lysosomal function. High-dose levo-dopa reduced mitochondrial membrane potential. At concentrations observed in the patient, levo-dopa enhanced immunoreactivity to beta III tubulin. Critically, levodopa reduced lysosomal content and also reduced the proportion of lysosomes that were acidic while homocysteine tended to have the opposite effect. Levodopa is a critically important drug for the treatment of PD. However, our data suggests that at concentrations observed in the patient, it has deleterious effects on sensory neurons that are not related to homocysteine.

Simple Summary

Parkinson’s disease (PD) is one of the most common chronic, degenerative brain diseases worldwide. Patients are diagnosed on the basis of slowness of movement and/or tremor and/or stiffness. However, many symptoms that are not movement related are now well recognized. Patients show changes in skin sensation, and the vast majority of patients show loss of sensory neurites, which enable sensation in skin. These changes in skin sensation occur prior to diagnosis; however, sensory issues may also be exacerbated by levodopa, an important drug used in the treatment of PD. Undoubtedly, levodopa is critical for the treatment of PD, but at high doses, it has repeatedly been shown to impair sensation in PD patients. Here, we show for the first time that high-dose levodopa impairs function of sensory neurons. Importantly, we also show for the first time that lysosomes, a critical organelle involved in recycling, are impaired by levodopa concentrations observed in patients. These data are important given the well-known lysosomal dysfunction observed in PD. Our data sheds light on how levodopa, the most important drug in the treatment of PD, may contribute to sensory deficits in PD.
Basic interactions responsible for thymus function explain convoluted medulla shape
Authors: Muramatsu David , Weyer Henrik , Gartner Florian , Frey Erwin
Published: October 2024

The thymus is one of the most important organs of the immune system. It is responsible for both the production of T cells and the prevention of their autoimmunity. It comprises two types of tissue: the cortex, where nascent T cells (thymocytes) are generated; and the medulla, embedded within the cortex, where autoreactive thymocytes are eliminated through negative selection. In mice, the medulla exhibits a complex, convoluted morphology, which has raised the question of whether its form impacts its function. Intriguingly, experiments also reveal a reverse dependency: the interactions between medullary stroma and thymocytes shape the medullary structure. However, understanding the underlying mechanisms of medulla morphogenesis emerging from these interactions remains elusive. Here, we present a conceptual theoretical model which shows that central, experimentally verified signaling pathways suffice to shape the convoluted medullary structure. The mathematical analysis of the model explains the observed effects of chemotaxis on thymocyte localization, as well as the reported morphological changes resulting from the modulation of thymocyte production. Our findings reveal that the established cross-talk between medulla growth and negative selection of thymocytes not only regulates medullary volume but also orchestrates the morphology of the thymus medulla. This mechanism of structure formation robustly organizes the medulla in a way that accelerates thymocyte negative selection by improving their chemotactic migration into the medulla. Thereby, we identify a feedback between the function of the thymus medulla and its form. Our theoretical study motivates further experimental analysis of the spatial distribution of thymic cell populations and predicts morphological changes under genetic perturbations.
Efficacy and safety of an mRNA-based RSV preF vaccine in preclinical studies
Authors: Bai Huarong , Li Qin , Gao Yue , Zhao Yubin , Yu Xueliang , Hu Rongkuan
Published: October 2024

The transmembrane fusion (F) protein of RSV plays important roles in RSV pathogenesis as it mediates the fusion between virus and the target cell membrane. During the fusion process, F protein transits from a metastable state (prefusion, preF) to a stable state after merging of virus and cell membranes (postfusion, postF). The majority of highly neutralizing antibodies induced by natural infection or immunization targets the preF form, making it the preferred antigen for vaccine development. Here, we evaluate the mRNA vaccine candidate, STR-V003, which encapsulates the modified mRNA encoding the preF protein in lipid nanoparticles (LNPs). This vaccine demonstrated robust immunogenic in both mice and cotton rats. STR-V003 induced high levels of neutralizing antibodies and RSV preF-specific IgG antibodies, and significantly reduced the RSV viral loads in the lung and nose tissue of challenged animals. In addition, STR-V003 did not have obvious enhancement of lung pathology without causing vaccine enhanced disease (VED). The repeated dose general toxicology studies and local tolerance studies of STR-V003 were evaluated in rats. Therefore, STR-V003 has an acceptable safety profile and robust protective immunity against RSV, and has been approved by the FDA to enter phase I clinical study (NCT06344975).