bluuebunny/crossref_metadata_2025 huggingface dataset:
Genome-wide identification of silencers in human cells
Authors: Zhu Xiusheng , Wang Chao , Kong Dashuai , Luo Jing , Deng Biao , Gu Yi , Kong Siyuan , Huang Lei , Liu Yuwen , Zhang Yubo
Published: June 2023

Transcriptional regulation is a complex process that is controlled by a variety of factors, including enhancers and silencers. Silencers, also known as repressor elements, play a crucial role in the fine-tuning of gene expression by inhibiting or suppressing transcription in the human genome. Although significant progresses have been made, genome-wide silencer research is still in its early stages. Here, we used a genome-wide method called massively parallel reporter assays (MPRAs) to identify silencers in three human cell lines: K562, LNCap, and HEK293T. We identified 739,434, 643,484, and 491,952 silencer regions in these cell lines, respectively. We found that most of the silencers we identified had inhibitory activity and significantly enriched inhibitory motifs. These results confirm that silencers are ubiquitous in the human genome and play an important role in regulating gene expression. Therefore, our study provides a general strategy for genome-wide functional identification of silencer elements. This information could be used to better understand the mechanisms of gene regulation and to develop new therapeutic strategies for diseases that are caused by dysregulation of gene expression.
Reversal of carbapenem resistance inPseudomonas aeruginosaby camelid single domain antibody fragment (VHH) against the C4 dicarboxylate transporter
Authors: Nagraj Anil Kumar , Shukla Manjiri , Kulkarni Mansi , Patil Pratik , Borgave Mrunal , Banerjee Sanjiban K
Published: June 2023

The COVID-19 pandemic has hastened the problem of nosocomial drug-resistant pathogens and is exerting a huge toll on hospitalized patients in critical care settings. Most small molecule antibiotics are susceptible to bacterial resistance mechanisms and constantly becoming ineffective leading to rapid shrinkage of the antibiotic armamentarium.Pseudomonas aeruginosa, one of the most common pathogens in hospital-born infection and considered critically dangerous by WHO and CDC, is extremely difficult to treat with frontline drugs, the carbapenems. In this study, we developed a camelid antibody fragment (VHH) library against wholeP. aeruginosaand isolated a highly potent neutralizing hit (PsC23) that selectively targetsP. aeruginosa .At 25 µg/mL, PsC23 inhibited growth of the ATCC 27853 and a locally isolated carbapenem resistant MCC 50428P. aeruginosa. The target of PsC23 is the C4 dicarboxylate transporter that transports C4 metabolites to the glyoxylate shunt during oxidative stress that is present in pathogens but not the human host. This ultimately results in the blockade of the shunt affecting bacterial energy transduction that leads to disruption of drug efflux. Interestingly, in a neutropenic mouse with MCC 50428 systemic infection, PsC23 in combination with meropenem completely reversed the drug resistance and eliminated the pathogens from the blood. PsC23 was stable in human serum and had no hemolytic or cytotoxic effect on human cells. Taken together, this VHH if co administered with the currently available carbapenems would reverse carbapenem resistance and could be used to effectively controlP. aeruginosain critical care settings.
Mapping MAVE data for use in human genomics applications
Authors: Arbesfeld Jeremy A. , Da Estelle Y. , Stevenson James S. , Kuzma Kori , Paul Anika , Farris Tierra , Capodanno Benjamin J. , Grindstaff Sally B. , Riehle Kevin , Saraiva-Agostinho Nuno , Safer Jordan F. , Milosavljevic Aleksandar , Foreman Julia , Firth Helen V. , Hunt Sarah E. , Iqbal Sumaiya , Cline Melissa S. , Rubin Alan F. , Wagner Alex H.
Published: June 2023

The large-scale experimental measures of variant functional assays submitted to MaveDB have the potential to provide key information for resolving variants of uncertain significance, but the reporting of results relative to assayed sequence hinders their downstream utility. The Atlas of Variant Effects Alliance mapped multiplexed assays of variant effect data to human reference sequences, creating a robust set of machine-readable homology mappings. This method processed approximately 2.5 million protein and genomic variants in MaveDB, successfully mapping 98.61% of examined variants and disseminating data to resources such as the UCSC Genome Browser and Ensembl Variant Effect Predictor.
Differential roles of diet on development and spinal cord regeneration in larval zebrafish
Authors: Purifoy Emily J. , Mruk Karen
Published: June 2023

The zebrafish is a powerful model organism for studying development and regeneration. However, there is a lack of a standardized reference diet. Most studies evaluate the rate of growth, survival and fecundity. In this study, we compare three diets and their effects on growth and regeneration after a spinal cord injury (SCI). Fish were fed daily for one week with daily measurements of overall length and width of spinal injury. Significant different in length after the trial were observed between live feed and commercial feeds. Similarly, differences in rate of regeneration were observed. Our data highlights the need for establishing a standardized diet for regeneration studies to improve research reproducibility.
RIBO-former: leveraging ribosome profiling information to improve the detection of translated open reading frames
Authors: Clauwaert Jim , McVey Zahra , Gupta Ramneek , Menschaert Gerben
Published: June 2023

Abstract

Ribosome profiling is a deep sequencing technique used to chart translation by means of mRNA ribosome occupancy. It has been instrumental in the detection of non-canonical coding sequences. Because of the complex nature of next-generation sequencing data, existing solutions that seek to identify translated open reading frames from the data are still not perfect. We propose RIBO-former, a new approach featuring several innovations for thede novoannotation of translated coding sequences. RIBO-former is built using recent transformer models that have achieved considerable advancements in the field of natural language processing. The presented deep learning approach allows to omit several pre-processing steps as features are automatically extracted from the data. We discuss various steps that improve the detection of coding sequences and show that read length information of all mapped reads can be leveraged to improve the predictive performance of the tool. Our results show RIBO-former to outperform previous methodologies. Additionally, through our study we find support for the existence of translated non-canonical ORFs, present along existing coding sequences or on long non-coding RNAs. Furthermore, several polycistronic mRNAs with multiple translated coding regions were detected.
1LocusSim a mobile-friendly simulator for teaching population genetics
Authors: Carvajal-Rodríguez Antonio
Published: June 2023

Biology students often struggle with the fundamental concepts of evolutionary genetics, including genetic drift, mutation, and selection. To address this problem, 1LocusSim was developed to simulate the interaction of different factors, such as population size, mutation, selection, and dominance, to study their effect on allelic frequency during evolution. With 1LocusSim, students can compare theoretical results with simulation outputs and solve and analyze different problems of population genetics. The 1LocusSim web has a responsive design which means that it has been specifically designed to be used on smartphones. To demonstrate its use, I review the classical overdominance model of population genetics and highlight a characteristic that is often not explicitly stated. Specifically, it is emphasized that the equilibrium of the model does not depend on the homozygous selection coefficients but rather on the ratio of the selection coefficients. This is already clear from the classical formula but maybe not so much for students. Also it implies that the equilibrium can be expressed solely in terms of the dominance coefficienth. To verify these theoretical prediction, I utilize the simulator and calculate the equilibrium for the well-known case of sickle cell anaemia.

Simulating basic population genetic models on smartphones can be a powerful learning aid that fosters critical skills and opens up new opportunities for Biology students. By utilizing this tool, students can learn at their own pace and convenience, anywhere and anytime.

Software and data availability

1LocusSim if freely available athttps://1LocusSim-biosdev.pythonanywhere.com/. Website implemented under the Bottle micro web-framework for Python, with all major browsers supported.

Contact

acraaj@uvigo.es

Supplementary information

The manual and examples are available from the help button of the program or directly athttps://acraaj.webs.uvigo.es/1LocusSim/1LocusSim_EN.html. Both the program and the manual pages are in English and Spanish.
The Role of Sympathetic Innervation in Neonatal Muscle Growth and Neuromuscular Contractures
Authors: Runkel Mason T. , Tarabishi Albaraa , Shay-Winkler Kritton , Emmert Marianne E. , Goh Qingnian , Cornwall Roger
Published: June 2023

Neonatal brachial plexus injury (NBPI), a leading cause of pediatric upper limb paralysis, results in disabling and incurable muscle contractures that are driven by impaired longitudinal growth of denervated muscles. A rare form of NBPI, which maintains both afferent and sympathetic muscle innervation despite motor denervation, protects against contractures. We have previously ruled out a role for NRG/ErbB signaling, the predominant pathway governing antegrade afferent neuromuscular transmission, in modulating the formation of contractures. Our current study therefore investigated the contributions of sympathetic innervation of skeletal muscle in modulating NBPI-induced contractures. Through chemical sympathectomy and pharmacologic modification with a β2-adrenergic agonist, we discovered that sympathetic innervation alone is neither required nor sufficient to modulate contracture formation in neonatal mice. Despite this, sympathetic innervation plays an intriguing sex-specific role in mediating neonatal muscle growth, as the cross-sectional area (CSA) and volume of normally innervated male muscles were diminished by ablation of sympathetic neurons and increased by β-adrenergic stimulation. Intriguingly, the robust alterations in CSA occurred with minimal changes to normal longitudinal muscle growth as determined by sarcomere length. Instead, β-adrenergic stimulation exacerbated sarcomere overstretch in denervated male muscles, indicating potentially discrete regulation of muscle width and length. Future investigations into the mechanistic underpinnings of these distinct aspects of muscle growth are thus essential for improving clinical outcomes in patients affected by muscle disorders in which both length and width are affected.
Body mass and growth rates predict protein intake across animals
Authors: Talal Stav , Harrison Jon F. , Farington Ruth , Youngblood Jacob P. , Medina Hector E. , Overson Rick , Cease Arianne J.
Published: June 2023

Organisms require dietary macronutrients in specific ratios to maximize performance, and variation in macronutrient requirements plays a central role in niche determination. Although it is well-recognized that development and body size can have strong and predictable effects on many aspects of organismal function, we lack a predictive understanding of ontogenetic or scaling effects on macronutrient intake. We determined protein and carbohydrate intake throughout development on lab populations of locusts and tested whether lab responses can predict results for field populations. Self-selected protein:carbohydrate targets declined dramatically through ontogeny, due primarily to declines in mass-specific protein consumption rates which were highly correlated with declines in specific growth rates. Importantly, lab results for protein consumption rates predicted results for field-collected locusts. However, field locusts consumed nearly double the carbohydrate, likely due to higher activity and metabolic rates. Combining our results with the available data for animals, both across species and during ontogeny, protein consumption scaled predictably and hypometrically, demonstrating a new scaling rule key for understanding nutritional ecology.
A Machine Learning Approach for Real-time Cortical State Estimation
Authors: Weiss David A , Borsa Adriano MF , Pala Aurélie , Sederberg Audrey J , Stanley Garrett B
Published: June 2023

Objective

Cortical function is under constant modulation by internally-driven, latent variables that regulate excitability, collectively known as “cortical state”. Despite a vast literature in this area, the estimation of cortical state remains relatively ad hoc, and not amenable to real-time implementation. Here, we implement robust, data-driven, and fast algorithms that address several technical challenges for online cortical state estimation.

Approach

We use unsupervised Gaussian Mixture Models (GMMs) to identify discrete, emergent clusters in spontaneous local field potential (LFP) signals in cortex. We then extend our approach to a temporally-informed Hidden semi-Markov Model (HSMM) with Gaussian observations to better model and infer cortical state transitions. Finally, we implement our HSMM cortical state inference algorithms in a real-time system, evaluating their performance in emulation experiments.

Main results

Unsupervised clustering approaches reveal emergent state-like structure in spontaneous electrophysiological data that recapitulate arousal-related cortical states as indexed by behavioral indicators. HSMMs enable cortical state inferences in a real-time context by modeling the temporal dynamics of cortical state switching. Using HSMMs provides robustness to state estimates arising from noisy, sequential electrophysiological data.

Significance

To our knowledge, this work represents the first implementation of a real-time software tool for continuously decoding cortical states with high temporal resolution (40 ms). The software tools that we provide can facilitate our understanding of how cortical states dynamically modulate cortical function on a moment-by-moment basis and provide a basis for state-aware brain machine interfaces across health and disease.
IFT cargo and motors associate sequentially with IFT trains to enter cilia
Authors: Mitra Aniruddha , Loseva Elizaveta , Peterman Erwin J.G.
Published: June 2023

Intraflagellar transport (IFT) orchestrates entry of proteins into primary cilia. At the ciliary base, assembled IFT trains, driven by kinesin-2 motors, can transport cargo proteins into the cilium, across the crowded transition zone (TZ). How trains assemble at the base and how proteins associate with them is far from understood. Here, we use single-molecule imaging in the cilia ofC. eleganschemosensory neurons to directly visualize the entry of kinesin-2 motors kinesin-II and OSM-3, as well as anterograde cargo proteins IFT dynein and tubulin. Single-particle tracking shows that IFT components associate with trains sequentially, both in time and space. Super-resolution maps of IFT components in wild-type and mutant worms reveal ciliary ultrastructure and show that kinesin-II is essential for axonemal organization. Finally, imaging cilia lacking kinesin-II and/or TZ function uncovers the interplay of kinesin-II and OSM-3 in driving efficient transport of IFT trains across the TZ.
Multi_CycGT: a DL-based multimodal model for membrane permeability prediction of cyclic peptides
Authors: Cao Lujing , Xu Zhenyu , Shang Tianfeng , Zhang Chengyun , Wu Xinyi , Wu Yejian , Zhai Silong , Ma Liefeng , Duan Hongliang
Published: June 2023

As a highly versatile therapeutic modality, cyclic peptides have gained significant attention due to their exceptional binding affinity, minimal toxicity and capacity to target the surface of conventionally “undruggable” proteins. However, the development of cyclic peptides with therapeutic effects by targeting intracellular biological targets has been hindered by the issue of limited membrane permeability. In this paper, we have conducted an extensive benchmarking analysis of a proprietary dataset consisting of 6941 cyclic peptides, employing machine learning and deep learning models. In addition, we propose an innovative multimodal model called Multi_CycGT which combines a Graph Convolutional Network (GCN) and a Transformer to extract 1D and 2D features. These encoded features are then fused for the prediction of cyclic peptide permeability. The cross-validation experiments demonstrate that the proposed Multi_CycGT model achieved the highest level of accuracy on the test set, with an accuracy value of 0.8206 and an AUC value of 0.8650. This paper introduces a pioneering deep learning-based approach that demonstrates enhanced effectiveness in predicting the membrane permeability of cyclic peptides. It also represents the first attempt in this field. We hope that this work will help to accelerate the design of cyclic peptide active drugs in medicinal chemistry and chemical biology applications.
Spatiotemporal Control of Genomics and Epigenomics by Ultrasound
Authors: Wu Yiqian , Huang Ziliang , Liu Yahan , Yoon Chi Woo , Sun Kun , Situ Yinglin , Ho Phuong , Yuan Zhou , Zhu Linshan , Eyquem Justin , Zhao Yunde , Liu Thomas , Kwong Gabriel A , Chien Shu , Wang Yingxiao
Published: June 2023

CRISPR (clustered regularly interspaced short palindromic repeats) is a revolutionary technology for genome editing. Its derived technologies such as CRISPR activation (CRISPRa) and CRISPR interference (CRISPRi) further allow transcriptional and epigenetic modulations. Focused ultrasound (FUS) can penetrate deep in biological tissues and induce mild hyperthermia in a confined region to activate heat-sensitive genes. Here we engineer a set of CRISPR(a/i) tools containing heat-sensitive genetic modules controllable by FUS for the regulation of genome and epigenome in live cells and animals. We demonstrated the capabilities of FUS-inducible CRISPRa, CRISPRi, and CRISPR (FUS-CRISPR(a/i)) to upregulate, repress, and knockout exogenous and/or endogenous genes, respectively, in different cell types. We further targeted FUS-CRISPR to telomeres in tumor cells to induce telomere disruption, inhibiting tumor growth and enhancing tumor susceptibility to killing by chimeric antigen receptor (CAR)-T cells. FUS-CRISPR-mediated telomere disruption for tumor priming combined with CAR-T therapy demonstrated synergistic therapeutic effects in xenograft mouse models. The FUS-CRISPR(a/i) toolbox allows the remote, noninvasive, and spatiotemporal control of genomic and epigenomic reprogramming in vivo, with extended applications in cancer treatment.
The ALOG domain defines a new family of plant-specific Transcription Factors acting during Arabidopsis flower development
Authors: Rieu Philippe , Beretta Veronica , Caselli Francesca , Thévénon Emmanuel , Lucas Jérémy , Rizk Mahmoud , Franchini Emanuela , Caporali Elisabetta , Nanao Max , Kater Martin , Dumas Renaud , Zubieta Chloe , Parcy François , Gregis Veronica
Published: June 2023

The ALOGs (ArabidopsisLIGHT-DEPENDENT SHORT HYPOCOTYLS 1and OryzaG1) are Transcription Factors (TFs) from an evolutionarily conserved plant-specific family shown to play critical roles in meristem identity, inflorescence architecture and organ boundaries in diverse species from mosses to higher flowering plants. However, the DNA binding-specificity and molecular determinants of protein-DNA interactions of this family were uncharacterized. Usingin vitrogenome-wide studies, we identified the conserved DNA motif bound by ALOG proteins from the liverwortMarchantia polymorphaand the flowering plants Arabidopsis, tomato and rice. In order to determine the amino acids responsible for DNA-binding specificity, we solved the 2.1Å structure of the ALOG DNA binding domain in complex with its cognate DNA. The ALOG DBD is an all-alpha helical domain with a structural zinc ribbon insertion and an N-terminal disordered NLS. The NLS sequence forms an integral part of the DNA binding domain and contributes to direct base read-out. To define the function of a group of redundant ALOG proteins in the model plant Arabidopsis thaliana, we generated a series ofalogmutants and uncovered their participation in a gene regulatory network involving the other floral regulators LEAFY, BLADE-ON-PETIOLE and PUCHI, all active in defining boundary regions between reproductive meristems and repressing bracts development. Taken together, this work provides the biochemical and structural basis for DNA-binding specificity of an evolutionarily conserved TF family and reveals its role as a key player in defining organ boundaries in Arabidopsis.

Significance Statement

Transcription Factors (TFs) are key proteins that bind specific regions in the genome and regulate the expression of associated genes. Not all organisms possess the same set of TFs and some, like the ALOGs, are specific to the plant kingdom. These TFs have been shown to play important roles from mosses to flowering plants. However, it was not known what DNA motif they recognize and how they bind DNA. Here we identify this motif, we show it is widely conserved in evolution and we understand how this new type of DNA binding domain works at the structural level. In addition, we also show that severalALOGgenes from Arabidopsis share a redundant function within the genetic network underlying correct floral meristem development.
Overcoming migratory exhaustion: Expression of Matrix metalloproteinase-8 (MMP-8) promotes tumor infiltration and anti-tumor activity of CAR-T cells
Authors: Bandey Irfan N. , Montalvo Melisa J. , Singh Harjeet , Varadarajan Navin
Published: June 2023

Despite the encouraging success of chimeric antigen receptor (CAR) T-cell therapy in treating hematological malignancies, the translation of adoptive cell therapies to solid tumors remains a challenge. Several studies have attributed the inability of tumor-infiltrating T cells to traffic to solid tumors, primarily to the presence of the extracellular matrix (ECM) and immunosuppressive environment of solid tumors. The ability of the transferred T cells to infiltrate the tumor is an essential prerequisite for anti-tumor activity. We show here that upon activation and expansion, T cells quickly lose their migratory capacity, leading to migratory exhaustion. At the molecular level, migratory exhaustion could be attributed to the downregulation of matrix metalloproteinase 8 (MMP8). To overcome this, we hypothesized that T cells genetically modified to secrete the mature form of matrix metalloproteinase 8 (mMMP8) would facilitate migration across matrix barriersin vitroandin vivo. We demonstrated that CAR T cells that co-express mMMP8 demonstrate robust migration across Matrigel and can kill tumor cells embedded in Matrigelin vitro. We tested the efficacy of these mMMP8 engineered cells in both leukemic and ovarian cancer cell models embedded in Matrigel in xenograft mouse models. Our results illustrate that unlike parental CAR T cells that have minimal anti-tumor efficacy in these models, CAR T cells that secrete mMMP8 promote T-cell infiltration, leading to the eradication of the tumors and survival. We anticipate that the co-expression of mMMP8 can be broadly utilized to improve the infiltration and efficacy of CAR T cells targeting many different antigens.
Macromolecular Crowding Promotes Reentrant Liquid-Liquid Phase Separation of Human Serum Transferrin and Prevents Surface-Induced Fibrillation
Authors: Patel Chinmaya Kumar , Rani Chanchal , Kumar Rajesh , Mukherjee Tushar Kanti
Published: June 2023

Protein aggregation and inactivation upon surface immobilization are major limiting factors for analytical applications in biotechnology related fields. Protein immobilization on solid surfaces often requires multi-step surface passivation which is time consuming and inefficient. Herein, we have discovered that biomolecular condensates of biologically active human serum transferrin (Tf) can effectively prevent surface-induced fibrillation and preserve the native-like conformation of phase separated Tf over a period of 30-days. It has been observed that macromolecular crowding promotes homotypic liquid-liquid phase separation (LLPS) of Tf through enthalpically driven multivalent hydrophobic interactions possibly via the involvement of its low complexity domain (residue 3–20) containing hydrophobic amino acids. The present LLPS of Tf is a rare example of salt-mediated reentrant phase separation in a broad range of salt concentrations (0–3 M) solely via the involvement of hydrophobic interactions. Notably, no liquid-to-solid-like phase transition has been observed over a period of 30-days, suggesting the intact conformational integrity of phase separated Tf as revealed from single droplet Raman, circular dichroism, and Fourier transform infrared spectroscopy measurements. More importantly, we discovered that the phase separated condensates of Tf completely inhibit the surface-induced fibrillation of Tf, illustrating the protective role of these liquid-like condensates against denaturation and aggregation of biomolecules. The cell mimicking aqueous compartments of biomolecular condensates with a substantial amount of interfacial water preserve the structure and functionality of biomolecules. Our present study highlights an important functional aspect of biologically active protein condensates and may have wide-ranging implications in cell physiology and biotechnological applications.
Antioxidant Polysulfide Nanoparticles Ameliorate Ischemia-Reperfusion Injury and Improve Porcine Kidney Function Post-Transplantation
Authors: Stone John Philip , D'Arcy Richard , Geraghty Abbey , Amin Kavit , Montero-Fernandez Angeles , Dabare Dilan , Ghibaudi Matilde , Francini Nora , Coradello Giulia , Bramhall Jo , Galwey Nicholas William , Ruseva Marieta , Tirelli Nicola , Fildes James
Published: June 2023

Ischemia-reperfusion injury (IRI) is a significant complication in kidney transplantation, often affecting the viability and function of organs. Normothermic machine perfusion (NMP) is a technique used to improve the condition of organs prior to transplantation. In this study, we show that incorporating antioxidant poly(propylene sulfide) nanoparticles (PPS-NPs) during cold-storage and NMP significantly enhances its efficacy in reducing IRI upon porcine kidney transplantation. We found that by scavenging reactive oxygen species, PPS-NPs reduced oxidative stress and inflammation that occurs during ischemia-reperfusion with oxidized DNA reduced 5.3x and both TNF-α and complement activation approximately halved. Our studies show that this approach led to significantly improved hemodynamics, better renal function, and tissue health compared to NMP alone. The results suggest that incorporating PPS-NPs into transplantation protocols may expand the pool of kidneys suitable for transplantation and enhance overall transplantation success rates. The broader impact of this work could extend to other organ transplants, suggesting a wider application for nanoantioxidant technologies in organ preservation. Methods: A randomised, two-stage, preclinical trial was used to assess the impact of poly(propylene sulfide) (polysulfide) nanoparticles (PPS-NPs) on parameters associated with IRI in a renal NMP system (experiment 1, n=6 vs 6). Paired porcine kidneys were randomised to receive either an NP-preservation flush followed by 6 hours of NMP with NP-perfusate, or control preservation flush and standard NMP. Following this, an allogeneic transplant- reperfusion model was used to evaluate if treatment with PPS-NPs improved renal haemodynamics post-transplantation (experiment 2, n=6 vs 6). Kidneys were perfused for 3 hours with or without NP, before being reperfused on a circuit primed with matched blood from genetically different donor pigs for 6 hours, without immunosuppression. Results: In experiment 1, all kidneys perfused well for 6 hours with physiological renal haemodynamics and biochemistry. Kidneys perfused with PPS-NPs had improved regional tissue perfusion on infra-red imaging. In experiment 2, renal haemodynamics were significantly improved during allogeneic reperfusion (post-transplant) after treatment with NP. Complement activation remained significantly lower in treated kidneys with a diminished TNF-a response. This translated into an improvement in tissue integrity. Conclusion: IRI was ameliorated following treatment with NPs during preservation and NMP. This was evidenced by an improvement in renal haemodynamics and diminished inflammatory markers upon reperfusion with allogeneic blood.
Enrichment of hard sweeps on the X chromosome compared to autosomes in sixDrosophilaspecies
Authors: Harris Mariana , Kim Bernard , Garud Nandita
Published: June 2023

The X chromosome, being hemizygous in males, is exposed one third of the time increasing the visibility of new mutations to natural selection, potentially leading to different evolutionary dynamics than autosomes. Recently, we found an enrichment of hard selective sweeps over soft selective sweeps on the X chromosome relative to the autosomes in a North American population ofDrosophila melanogaster. To understand whether this enrichment is a universal feature of evolution on the X chromosome, we analyze diversity patterns across six commonly studiedDrosophilaspecies. We find an increased proportion of regions with steep reductions in diversity and elevated homozygosity on the X chromosome compared to autosomes. To assess if these signatures are consistent with positive selection, we simulate a wide variety of evolutionary scenarios spanning variations in demography, mutation rate, recombination rate, background selection, hard sweeps, and soft sweeps, and find that the diversity patterns observed on the X are most consistent with hard sweeps. Our findings highlight the importance of sex chromosomes in driving evolutionary processes and suggest that hard sweeps have played a significant role in shaping diversity patterns on the X chromosome across multipleDrosophilaspecies.
Positive selection and relaxed purifying selection contribute to rapid evolution of male-biased genes in a dioecious flowering plant
Authors: Zhao Lei , Zhou Wei , He Jun , Li De-Zhu , Li Hong-Tao
Published: June 2023

Sex-biased genes offer insights into the evolution of sexual dimorphism. Sex-biased genes, especially those with male bias, show elevated evolutionary rates of protein sequences driven by positive selection and relaxed purifying selection in animals. Although rapid sequence evolution of sex-biased genes and evolutionary forces have been investigated in animals and brown algae, less is known about evolutionary forces in dioecious angiosperms. In this study, we separately compared the expression of sex-biased genes between female and male floral buds and between female and male flowers at anthesis in dioeciousTrichosanthes pilosa(Cucurbitaceae). In floral buds, sex-biased gene expression was pervasive, and had significantly different roles in sexual dimorphism such as physiology. We observed higher rates of sequence evolution for male-biased genes in floral buds compared to female-biased and unbiased genes. Male-biased genes under positive selection were mainly associated with functions to abiotic stress and immune responses, suggesting that high evolutionary rates are driven by adaptive evolution. Additionally, relaxed purifying selection may contribute to accelerated evolution in male-biased genes generated by gene duplication. Our findings, for the first time in angiosperms, suggest evident rapid evolution of male-biased genes, advance our understanding of the patterns and forces driving the evolution of sexual dimorphism in dioecious plants.
Molecular physiological characterization of the dynamics of persister formation inStaphylococcus aureus
Authors: Liu Shiqi , Huang Yixuan , Jensen Sean , Laman Paul , Kramer Gertjan , Zaat Sebastian A. J. , Brul Stanley
Published: June 2023

Bacteria possess the ability to enter a growth arrested state known as persistence in order to survive antibiotic exposure. Clinically, persisters are regarded as the main causative agents for chronic and recurrent infectious diseases. To combat this antibiotic-tolerant population, a better understanding of the molecular physiology of persisters is required. In this study, we collected samples at different stages of the biphasic kill curve to reveal the dynamics of the cellular molecular changes that occur in the process of persister formation. After exposure to antibiotics with different modes of action, namely vancomycin and enrofloxacin, similar persister levels were obtained. Both shared and distinct stress responses were enriched for the respective persister populations. However, the dynamics of the presence of proteins linked to the persister phenotype throughout the biphasic kill curve and the molecular profiles in a stable persistent population did show large differences depending on the antibiotic used. This suggests that persisters at the molecular level are highly stress specific, emphasizing the importance of characterizing persisters generated under different stress conditions. Additionally, although generated persisters exhibited cross-tolerance toward tested antibiotics, combined therapies were demonstrated to be a promising approach to reduce persister levels. In conclusion, this investigation sheds light on the stress-specific nature of persisters, highlighting the necessity of tailored treatment approaches and the potential of combined therapy.

Importance

By monitoring proteome and metabolites duringStaphylococcus aureuspersister formation under vancomycin and enrofloxacin exposure, we revealed the dynamic information of the molecular physiology of persister formation upon exposure to two different antibiotics with different modes of action. The data shows that cells that phenotypically are similarly classified as persisters, do have several molecular characteristics in common but, remarkably so, differ substantially in a significant number of other aspects of their molecular makeup. These contrasts provided valuable insights into persister eradication, which holds considerable clinical relevance.
Discovery, Characterization, and Bioactivity of the Achromonodins: Lasso Peptides Encoded byAchromobacter
Authors: Carson Drew V. , Zhang Yi , So Larry , Cheung-Lee Wai Ling , Cartagena Alexis Jaramillo , Darst Seth A. , Link A. James
Published: June 2023

Through genome mining efforts, we discovered two lasso peptide biosynthetic gene clusters (BGCs) within two different species ofAchromobacter, a genus that contains pathogenic organisms that can infect patients with cystic fibrosis. Using gene-refactored BGCs inE. coli, we heterologously expressed two lasso peptides, which we named achromonodin-1 and achromonodin-2. Achromonodin-1 is naturally encoded by certain isolates from the sputum of patients with cystic fibrosis. We solve the NMR structure of achromonodin-1, demonstrating that it is a threaded lasso peptide with a large loop and short tail structure, reminiscent of previously characterized lasso peptides that inhibit RNA polymerase (RNAP). We then show that achromonodin-1 inhibits RNAPin vitroand has potent but narrow-spectrum activity towardsAchromobacter pulmonis, another isolate from the sputum of a cystic fibrosis patient. Our efforts expand the repertoire of antimicrobial lasso peptides and provide insights into howAchromobacterisolates from certain ecological niches may interact with each other.
ALS-related p97 R155H mutation disrupts lysophagy in iPSC-derived motor neurons
Authors: Klickstein Jacob A. , Johnson Michelle A. , Antonoudiou Pantelis , Maguire Jamie , Paulo Joao A. , Gygi Steve P. , Weihl Chris , Raman Malavika
Published: June 2023

Mutations in the AAA+ ATPase p97 (also known as valosin containing protein, VCP) cause multisystem proteinopathy 1 (MSP-1) which includes amyotrophic-lateral sclerosis (ALS); however, the pathogenic mechanisms that contribute to motor neuron loss in familial ALS caused by p97 mutations remain obscure. Here, we use two distinct induced pluripotent stem cell models differentiated into spinal motor neurons to investigate how p97 mutations perturb the motor neuron proteome. Using multiplexed quantitative proteomics in these cells, we find that motor neurons harboring the p97 R155H mutation have deficits in lysosome quality control and the selective autophagy of lysosomes (lysophagy). p97 R155H motor neurons are unable to efficiently clear damaged lysosomes and have reduced viability. Additionally, lysosomes in mutant motor neurons have increased pH compared to their wildtype counterparts. The endo-lysosomal damage repair (ELDR) complex is required for clearance of damaged lysosomes and involves UBXD1-p97 interaction which is disrupted in mutant motor neurons. Finally, we report that inhibition of the ATPase activity of p97 R155H using the ATP competitive inhibitor CB-5083 rescues lysophagy defects in mutant motor neurons. These results add to the increasing evidence that endo-lysosomal dysfunction is a key aspect of disease pathogenesis in p97-related disorders.
Functional interaction of torsinA and its activators in liver lipid metabolism
Authors: Hernandez-Ono Antonio , Zhao Yi Peng , Murray John W. , Östlund Cecilia , Lee Michael J. , Shi Angsi , Dauer William T. , Worman Howard J. , Ginsberg Henry N. , Shin Ji-Yeon
Published: June 2023

TorsinA is an atypical ATPase that lacks intrinsic activity unless it is bound to its activators lamina-associated polypeptide 1 (LAP1) in the perinuclear space or luminal domain-like LAP1 (LULL1) throughout the endoplasmic reticulum. However, the interaction of torsinA with LAP1 and LULL1 has not yet been shown to modulate a defined physiological process in mammalsin vivo. We previously demonstrated that depletion of torsinA from mouse hepatocytes leads to reduced liver triglyceride secretion and marked steatosis, whereas depletion of LAP1 had more modest similar effects. We now show that depletion of LULL1 alone does not significantly decrease liver triglyceride secretion or cause steatosis. However, simultaneous depletion of both LAP1 and LULL1 from hepatocytes leads to defective triglyceride secretion and marked steatosis similar to that observed with depletion of torsinA. Our results demonstrate that torsinA and its activators dynamically regulate a physiological process in mammalsin vivo.
Network States in the Basolateral Amygdala Predicts Voluntary Alcohol Consumption
Authors: DiLeo Alyssa , Antonodiou Pantelis , Blandino Katrina , Conlin Eli , Melón Laverne , Maguire Jamie L.
Published: June 2023

Although most adults in the United States will drink alcohol in their life, only about 6% will go on to develop an alcohol use disorder (AUD). While a great deal of work has furthered our understanding of the cycle of addiction, it remains unclear why certain people transition to disordered drinking. Altered activity in regions implicated in AUDs, like the basolateral amygdala (BLA), has been suggested to play a role in the pathophysiology of AUDs, but how these networks contribute to alcohol misuse remains unclear. Our recent work demonstrated that alcohol can modulate BLA network states and that GABAergic parvalbumin (PV) interneurons are crucial modulators of network activity in the BLA. Further, our lab has demonstrated that δ subunit-containing GABAAreceptors, which are modulated by alcohol, are highly expressed on PV interneurons in the BLA. These receptors on PV interneurons have also been shown to influence alcohol intake in a voluntary binge drinking paradigm and anxiety-like behavior in withdrawal. Therefore, we hypothesized that alcohol may impact BLA network states via δ subunit-containing GABAAreceptors on PV interneurons to impact the extent of alcohol use. To test this hypothesis, we measured the impact of acute alcohol exposure on oscillatory states in the basolateral amygdala and then assessed the relationship to the extent of voluntary ethanol consumption in the Intermittent Access, Drinking-in-the-Dark-Multiple Scheduled Access, and Chronic Intermittent Ethanol exposure paradigms. Remarkably, we demonstrate that the average alcohol intake negatively correlates with δ subunit-containing GABAAreceptor expression on PV interneurons and gamma power in the BLA after the first exposure to alcohol. These data implicate δ subunit-containing GABAAreceptor expression on PV interneurons in the BLA in voluntary alcohol intake and suggest that BLA network states may serve as a useful biomarker for those at risk for alcohol misuse.

Significance Statement

Oscillatory states in the BLA have been demonstrated to drive behavioral states involved in emotional processing, including negative valence processing. Given that negative emotional states/hyperkatifeia contribute to the cycle of AUDs, our previous work demonstrating the ability of alcohol to modulate BLA network states and thereby behavioral states suggests that this mechanism may influence alcohol intake. Here we demonstrate a relationship between the ability of alcohol to modulate oscillations in the BLA and future alcohol intake such that the extent to which alcohol influences BLA network states predict the extent of future voluntary alcohol intake. These findings suggest that individual variability in the sensitivity of the BLA network to alcohol influences voluntary alcohol consumption.
Developmental exposure to the Parkinson’s disease-associated organochlorine pesticide dieldrin alters dopamine neurotransmission in α- synuclein pre-formed fibril (PFF)-injected mice
Authors: Boyd Sierra L. , Kuhn Nathan C. , Patterson Joseph R. , Stoll Anna C. , Zimmerman Sydney A. , Kolanowski Mason R. , Neubecker Joseph J. , Luk Kelvin C. , Ramsson Eric S. , Sortwell Caryl E. , Bernstein Alison I.
Published: June 2023

Parkinson’s disease (PD) is the most common movement disorder and one of the fastest-growing neurological diseases worldwide. This increase outpaces the rate of aging and is most rapid in recently industrialized areas, suggesting the role of environmental factors. Consistent with this, epidemiological studies show an association between exposure to persistent organic pollutants and an increased risk of PD. When combined with post-mortem analysis and mechanistic studies, a role for specific compounds, including the organochlorine pesticide dieldrin, emerges. In mouse models, developmental dieldrin exposure causes male-specific exacerbation of neuronal susceptibility to MPTP and synucleinopathy. Specifically, our novel two-hit model combining developmental dieldrin exposure with the α-synuclein (α-syn) pre-formed fibril (PFF) model showed a male-specific exacerbation of PFF-induced increases in striatal dopamine (DA) turnover and motor deficits on the challenging beam 6 months post-PFF injection in male offspring developmentally exposed to dieldrin. Here, we hypothesized that alterations in DA handling contribute to the observed changes and assessed vesicular monoamine transporter 2 (VMAT2) function and DA release in this dieldrin/PFF two-hit model. Female C57BL/6 mice were exposed to 0.3 mg/kg dieldrin or vehicle every 3 days, starting at 8 weeks of age by feeding and continuing throughout breeding, gestation, and lactation. Male offspring from independent litters underwent unilateral, intrastriatal injections of α-syn PFFs via stereotaxic surgery at 12 weeks of age and DA handling was assessed 4 months post-PFF injection via vesicular3H-DA uptake assay and fast-scan cyclic voltammetry (FSCV). We observed no dieldrin-associated change in VMAT2 activity, but a dieldrin-induced increase in DA release in striatal slices in PFF-injected animals. These results suggest that developmental dieldrin exposure alters the dopaminergic response to synucleinopathy-triggered toxicity and supports our hypothesis that alterations in DA handling may underly the observed exacerbation of PFF-induced deficits in motor behavior and DA turnover.

Graphical Abstract
Soil carbon mineralization and microbial community dynamics in response to PyOM addition
Authors: Zeba Nayela , Berry Timothy D. , Fischer Monika S. , Traxler Matthew F. , Whitman Thea
Published: June 2023

Wildfires can either negatively impact soil carbon (C) stocks through combustion or increase soil carbon stocks through the production of pyrogenic organic matter (PyOM), which is highly persistent and can affect non-pyrogenic soil organic carbon (SOC) mineralization rates. In this study, we used fine-resolution13CO2flux tracing to investigate PyOM-C mineralization, soil priming effects, and their impacts on soil microbial communities in a Californian mixed conifer forest Xerumbrept soil burned in the 2014 King Fire. We added PyOM produced from pine biomass at 350 °C and 550 °C to the soil and separately traced the mineralization of13C-labeled water-extractable and non-water-extractable PyOM-C fractions in a short-term incubation.

Our results indicate that the water-extractable fraction is 10-50x more mineralizable in both 350 °C and 550 °C PyOM treatments than the SOC or non-water-extractable PyOM fraction. 350 °C PyOM addition led to a short-term positive priming effect, likely due to co-metabolism of easily mineralizable PyOM-C and the SOC, whereas 550 °C PyOM addition induced negative priming, potentially due to physical protection of SOC. We observed significant shifts in bacterial community composition in response to both 350 °C and 550 °C PyOM, with positive PyOM responders belonging to the generaNoviherbaspirillum,Pseudonocardia, andGemmatimonas. In contrast, fungal communities were less responsive to PyOM additions. Our findings expand our understanding of the post-fire cycling of PyOM and SOC, providing insights into the microbial mineralization of different PyOM-C fractions and their influence on soil C dynamics in fire-affected ecosystems.
Identification of ERAD-dependent degrons for the endoplasmic reticulum lumen
Authors: Sharninghausen Rachel , Hwang Jiwon , Dennison Devon D. , Baldridge Ryan D.
Published: June 2023

Degrons are minimal protein features that are sufficient to target proteins for degradation. In most cases, degrons allow recognition by components of the cytosolic ubiquitin proteasome system. Currently, all of the identified degrons only function within the cytosol. UsingSaccharomyces cerevisiae, we identified the first short linear sequences that function as degrons from the endoplasmic reticulum (ER) lumen. We show that when these degrons are transferred to proteins, they facilitate proteasomal degradation through the ERAD system. These degrons enable degradation of both luminal and integral membrane ER proteins, expanding the types of proteins that can be targeted for degradation in budding yeast and mammalian tissue culture. This discovery provides a framework to target proteins for degradation from the previously unreachable ER lumen and builds toward therapeutic approaches that exploit the highly-conserved ERAD system.