bluuebunny/crossref_metadata_2025 huggingface dataset:
Multifocal Ectopic Purkinje Premature Contractions due to neutralization of anSCN5Anegative charge: structural insights into the gating pore hypothesis
Authors: Glazer Andrew M. , Yang Tao , Li Bian , Page Dana , Fouda Mohamed , Wada Yuko , Lancaster Megan C. , O’Neill Matthew J. , Muhammad Ayesha , Gao Xiaozhi , Ackerman Michael J. , Sanatani Shubhayan , Ruben Peter C. , Roden Dan M.
Published: February 2024

Background

We identified a novelSCN5Avariant, E171Q, in a neonate with very frequent ectopy and reduced ejection fraction which normalized after arrhythmia suppression by flecainide. This clinical picture is consistent with multifocal ectopic Purkinje-related premature contractions (MEPPC). Most previous reports of MEPPC have implicatedSCN5Avariants such as R222Q that neutralize positive charges in the S4 voltage sensor helix of the channel protein NaV1.5 and generate a gating pore current.

Methods and Results

E171 is a highly conserved negatively-charged residue located in the S2 transmembrane helix of NaV1.5 domain I. E171 is a key component of the Gating Charge Transfer Center, a region thought to be critical for normal movement of the S4 voltage sensor helix. We used heterologous expression, CRISPR-edited induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), and molecular dynamics simulations to demonstrate that E171Q generates a gating pore current, which was suppressed by a low concentration of flecainide (IC50 = 0.71±0.07 µM). R222Q shifts voltage dependence of activation and inactivation in a negative direction but we observed positive shifts with E171Q. E171Q iPSC-CMs demonstrated abnormal spontaneous activity and prolonged action potentials. Molecular dynamics simulations revealed that both R222Q and E171Q proteins generate a water-filled permeation pathway that underlies generation of the gating pore current.

Conclusion

Previously identified MEPPC-associated variants that create gating pore currents are located in positively-charged residues in the S4 voltage sensor and generate negative shifts in the voltage dependence of activation and inactivation. We demonstrate that neutralizing a negatively charged S2 helix residue in the Gating Charge Transfer Center generates positive shifts but also create a gating pore pathway. These findings implicate the gating pore pathway as the primary functional and structural determinant of MEPPC and widen the spectrum of variants that are associated with gating pore-related disease in voltage-gated ion channels.
A SENSORIZED 3D-PRINTED KNEE TEST RIG FOR EXPERIMENTAL VALIDATION OF PATELLAR TRACKING AND CONTACT SIMULATION AFTER TOTAL KNEE REPLACEMENT
Authors: Michaud Florian , Mouzo Francisco , Dopico Daniel , Cuadrado Javier
Published: February 2024

Total knee arthroplasty aims to relieve pain and restore function in the affected joint through artificial implants. Despite advances in design and surgical techniques, there are complications associated with surgery, especially concerning the patella. The use of musculoskeletal models in orthopedic surgery allows for objective prediction of postoperative function and optimization of results for each patient. To ensure that simulations are trustworthy and can be used for predictive purposes, comparing simulation results with experimental data is crucial. Although progress has been made in obtaining 3D bone geometry and estimating contact forces, validation of these predictions has been limited due to the lack of direct in vivo measurements and the economic and ethical constraints associated with available alternatives. In this study, an existing commercial surgical training station was transformed into a sensorized test bench. The use of 3D-printed models and sensors allowed low-cost and reproducible experimental validation of computer simulation (patellar movement and forces) while avoiding ethical issues.
Vitamin D is Glucoprotective in Aging Males but Not Females
Authors: Ginnard Olivia Z.B. , Morales Maria , Youn Ji Youn , Xu Yong , Sisley Stephanie R.
Published: February 2024

Type 2 diabetes is strongly linked to vitamin D deficiency in older adults. However, there is a discrepancy between clinical trials in adults on the efficacy of vitamin D treatment in prediabetes and diabetes. In addition, human data indicates there may be sexual dimorphism in the effect of vitamin D deficiency on dysglycemia that is more pronounced in men. These incongruities may be due to our limited understanding of the underlying mechanisms of vitamin D in glucose homeostasis among its vast target tissues across the body. Furthermore, vitamin D deficiency is diagnosed by low levels of a storage form of vitamin D, which may not be an accurate indicator of vitamin D status in these individuals. Thus, measuring expression levels of vitamin D receptor (VDR)-target genes across tissues involved in glucose regulation and vitamin D pathways may be a more promising marker of vitamin D status. Here we describe the sex-specific physiological effects of vitamin D supplementation in an aged, non-obese mouse model on glucose homeostasis and tissue-specific gene regulation.
Analog epigenetic memory revealed by targeted chromatin editing
Authors: Palacios Sebastian , Bruno Simone , Weiss Ron , Salibi Elia , Kane Andrew , Ilia Katherine , Del Vecchio Domitilla
Published: February 2024

Chemical modifications to histones and DNA play a crucial role in the regulation of transcription and in the maintenance of chromatin states that are not permissive to gene expression [1–3]. However, the landscape of gene expression states that these modifications stably maintain remains uncharted. Here, we show that gene expression can be memorized at a wide range of levels thus implementing analog epigenetic memory. Mechanistically, we find that DNA methylation serves a primary role in maintaining memory across cell divisions while histone modifications only follow DNA methylation to regulate gene expression. Employing targeted epigenetic editing and time-course analysis, we analyzed the temporal stability of gene expression and DNA methylation post removal of epigenetic effectors. We found that the grade of DNA methylation in the gene’s promoter, defined as the mean fraction of methylated CpGs, remains stable over time and inversely correlates with gene expression level. By contrast, Histone 3 lysine 9 trimethylation (H3K9me3) could not persist after removal of its writer in the absence of DNA methylation. These experimental findings, combined with our chromatin modification model, indicate that the absence of positive feedback mechanisms around DNA methylation - unlike those found in histone modifications - enable the temporal stability of the DNA methylation grade, which leads to analog memory. These results expand current knowledge on how epigenetic memory is achieved in natural systems. Moreover, we anticipate that analog memory through graded DNA methylation will enable to program mammalian cells with fine-grained information storage. This capability will significantly enhance the sophistication of engineered cell functionality in applications including tissue engineering, organoids, and cell therapies.
Cancer-Specific Alterations in Nuclear Matrix Proteins Determined by Multi-omics Analyses of Ductal Carcinomain Situ
Authors: Almutairy Ali , Alhamed Abdullah , Grant Stephen G. , Sarachine Falso Miranda J. , Day Billy W. , Simmons Colton R. , Latimer Jean J.
Published: February 2024

Breast cancer (BC) is the most common cancer affecting women in the United States. Ductal carcinomain situ(DCIS) is the earliest identifiable pre-invasive BC lesion. Estimates show that 14 to 50% of DCIS cases progress to invasive BC. Our objective was to identify nuclear matrix proteins (NMP) with specifically altered expression in DCIS and later stages of BC compared to non-diseased breast reduction mammoplasty and a contralateral breast explant using mass spectrometry and RNA sequencing to accurately identify aggressive DCIS. Sixty NMPs were significantly differentially expressed between the DCIS and non-diseased breast epithelium in an isogenic contralateral pair of patient-derived extended explants. Ten of the sixty showed significant mRNA expression level differences that matched the protein expression. These 10 proteins were similarly expressed in non-diseased breast reduction cells. Three NMPs (RPL7A, RPL11, RPL31) were significantly upregulated in DCIS and all other BC stages compared to the matching contralateral breast culture and an unrelated non-diseased breast reduction culture. RNA sequencing analyses showed that these three genes were upregulated increasingly with BC progression. Finally, we identified three NMPs (AHNAK, CDC37 and DNAJB1) that were significantly downregulated in DCIS and all other BC stages compared to the isogenically matched contralateral culture and the non-diseased breast reduction culture using both proteomics and RNA sequencing techniques.
Effects of ketamine and propofol on muscarinic plateau potentials in rat neocortical pyramidal cells
Authors: Fleiner Anne S. , Kolnier Daniel , Hagger-Vaughan Nicholas , Raeder Johan , Storm Johan Frederik
Published: February 2024

Propofol and ketamine are widely used general anaesthetics, but have different effects on consciousness: propofol gives a deeply unconscious state, with little or no dream reports, whereas vivid dreams are often reported after ketamine anaesthesia. Ketamine is an N-methyl-D-aspartate (NMDA) receptor antagonist, while propofol is a gamma-aminobutyric-acid (GABAA) agonist, but these mechanisms do not fully explain how these drugs alter consciousness. Most previous in vitro studies of cellular mechanisms of anaesthetics have used brain slices or neurons in a nearly "comatose" state, because no "arousing" neuromodulators were added. Here we tested mechanisms of anaesthetics in slices after adding the cholinergic agonist muscarine to partly mimic an "awake-like" state. Using whole-cell patch-clamp recordings from layer 2/3 pyramidal cells (L2/3PCs) in rat medial prefrontal cortex (mPFC) slices, we saw that muscarine induced long-lasting depolarizing plateau potentials (PPs) and spiking following brief depolarizing current injections. According to leading theories of consciousness and working memory, L2/3PCs and PPs are particularly important for these cognitive functions. After 2 hours of pre-incubation with ketamine or propofol, the muscarine-induced PPs were altered in different ways: 3 uM propofol reduced the PPs and (significantly) spiking, whereas 20 μM ketamine seemed to enhance PPs and spiking (non-significantly). Brief wash-in of these drug concentrations failed to induce such effects, probably due to insufficient equilibration by diffusion in the slices. In contrast, pre-incubation with 100 uM ketamine suppressed the PPs and spiking. The different effects on PPs may be related to contrasting clinical effects: ketamine causing atypical anaesthesia with vivid, "psychedelic" dreaming while propofol causes less dreaming. However, high ketamine or propofol concentrations both suppressed PPs, suggesting possible connections between PPs, desynchronized activity, and consciousness. More experiments are needed to test these tentative conclusions.
Deciphering the function of intrinsic and genomics-driven epigenetic heterogeneity in head and neck cancer progression with single-nucleus CUT&RUN
Authors: Womersley Howard J. , Muliaditan Daniel , DasGupta Ramanuj , Cheow Lih Feng
Published: February 2024

Interrogating regulatory epigenetic alterations during tumor progression at the resolution of single cells has remained an understudied area of research. Here we developedahighly sensitive single-nucleus CUT&RUN (snCUT&RUN) assay to profile histone modifications in isogenic primary, metastatic, and cisplatin-resistant head and neck squamous cell carcinoma (HNSCC) patient-derived tumor cell lines. We find that the epigenome can be involved in diverse modes to contribute towards HNSCC progression. First, we demonstrate that gene expression changes during HNSCC progression can be co-modulated by alterations in both copy number and chromatin activity, driving epigenetic rewiring of cell-states. Furthermore, intratumour epigenetic heterogeneity (ITeH) may predispose sub-clonal populations within the primary tumour to adapt to selective pressures and foster the acquisition of malignant characteristics. In conclusion, snCUT&RUN serves as a valuable addition to the existing toolkit of single-cell epigenomic assays and can be used to dissect the functionality of the epigenome during cancer progression.
Myosin-19 and Miro regulate mitochondria-endoplasmatic reticulum contacts and mitochondria inner membrane architecture
Authors: Attia Aya , Majstrowicz Katarzyna , Shembekar Samruddhi , Honnert Ulrike , Nikolaus Petra , Lohman Birgit , Bähler Martin
Published: February 2024

Mitochondrial dynamics is important for cellular health and includes morphology, fusion, fission, vesicle formation, transport and contact formation with other organelles. Myosin XIX (Myo19) is an actin-based motor which competes with TRAK1/2 adaptors of microtubule-based motors for binding to the outer mitochondrial membrane receptors Mitochondrial Rho GTPases 1/2 (Miro). Currently, it is poorly understood how Myo19 contributes to mitochondrial dynamics. Here, we report on a Myo19-deficient mouse model and the ultrastructure of the mitochondria from cells of Myo19-deficient mice and HEK cells, Miro-deficient HEK cells and TRAK1-deficient HAP1 cells. Myo19-deficient mitochondria in kidney, skeletal and cardiac muscle cells, MEFs and HEK cells have morphological alterations in the inner mitochondrial membrane with reduced numbers of malformed cristae. In addition, mitochondria in Myo19-deficient cells showed fewer ER-mitochondria contact sites (ERMCS). In accordance with the ultrastructural observations, Myo19-deficient MEFs had lower oxygen consumption rates and a reduced abundance of OXPHOS supercomplexes. The simultaneous loss of Miro1 and Miro 2 led to a comparable mitochondria phenotype and reduced ERMCS as observed upon loss of Myo19. However, the loss of TRAK1 caused only a reduction in the number of cristae, but not ERMCS. These results demonstrate that both actin- and microtubule-based motors regulate cristae formation, but only Myo19 and its membrane receptor Miro regulate ERMCS.
E93 controls adult differentiation by repressingbroadinDrosophila
Authors: Cruz Josefa , Ureña Enric , Franch-Marro Xavier , Martín David
Published: February 2024

InDrosophila melanogaster, successful development relies on the precise coordination of both spatial and temporal regulatory axes. The temporal axis governs stage-specific identity and developmental transitions through a number of genes, collectively forming the Metamorphic Gene Network. Among these, Ecdysone inducible protein 93F (E93) serves as the critical determinant for adult specification, but its mechanism of action remains unclear. Here, we found that, rather than acting as an instructive signal, E93 promotes adult differentiation through the repression of the pupal specifierbroad (br). In the absence of E93, sustained high levels of Br during the pupal stage strongly represses pupal-specific enhancers that are essential for the terminal differentiation of the wing. We also show that Br represses the pupal-enhancers during the larval and prepupal stages preventing the premature implementation of the adult genetic program, and that it also dampens the activity of larval enhancers during the latter stages of larval development. This mechanism of action seems to be a derived feature acquired in Diptera, as in the coleopteranTribolium castaneum, repression ofbrby E93 is not sufficient to allow adult differentiation. In summary, our study elucidates the crucial role of the intricate interplay between E93 and Br as the governing mechanism in the process of terminal differentiation inDrosophila. This discovery holds significant implications for advancing our understanding of the evolution of insect metamorphosis.
In vitro activity of cefiderocol against carbapenemase-producing and meropenem-nonsusceptible gram-negative bacteria collected in the Japan Antimicrobial Resistant Bacterial Surveillance (JARBS-GNR)
Authors: Kayama Shizuo , Kawakami Sayoko , Kondo Kohei , Kitamura Norikazu , Yu Liansheng , Hayashi Wataru , Yahara Koji , Sugawara Yo , Sugai Motoyuki
Published: February 2024

The treatment options available for infections caused by multidrug-resistant gram-negative pathogens are often limited. Cefiderocol (CFDC) is a novel siderophore cephalosporin that exhibits activity against multidrug-resistant gram-negative pathogens. Several studies have reported the in vitro activity of CFDC using clinical isolates from Europe, the United States, and China. However, no large-scale studies on the in vitro activities of CFDC have been conducted using all isolates with available genomic backgrounds based on whole-genome sequencing (WGS). We evaluated the antimicrobial activities of CFDC, ceftolozane/tazobactam (CTLZ/TAZ), imipenem-relebactam (IPM/REL), and ceftazidime/avibactam (CAZ/AVI) against carbapenemase-producing Enterobacterales, carbapenemase-non-producing meropenem-nonsusceptible Enterobacterales, and carbapenemase-producing non-fermentative bacteria. We selected 603 isolates (528 Enterobacterales, 18Pseudomonas aeruginosa, and 57Acinetobacterspp.) from the recent surveillance of clinical isolates in Japan using WGS data. Among these, 97.7% (300/307 strains) of carbapenemase-producing Enterobacterales, 100% (18/18 strains) of carbapenemase-producingP. aeruginosa, and 91.2% (52/57 strains) of carbapenemase-producingAcinetobacterspp. were susceptible to CFDC, showing better antimicrobial activity than the other antimicrobial agents evaluated in this study. In addition, CFDC was highly effective against class A, B, and D β-lactamase harboring isolates when compared to the other antimicrobial agents in this study. While β-lactam antibiotics were essentially ineffective against CFDC-resistant Enterobacterales, minocycline was the most effective, and gentamicin and amikacin were also effective. This is the first large-scale study to systematically demonstrate the efficacy of CFDC using carbapenemase-producing strains with transparent genomic backgrounds.
The NAPstar family of NADP redox state sensors highlights glutathione as the primary mediator of anti-oxidative electron flux
Authors: Scherschel Marie , Niemeier Jan-Ole , Jacobs Lianne J.H.C. , Hoffmann Markus , Diederich Anika , Bell Christopher , Höhne Pascal , Raetz Sonja , Kroll Johanna B. , Steinbeck Janina , Lichtenauer Sophie , Multhoff Jan , Zimmermann Jannik , Sadhanasatish Tanmay , Rothemann R. Alexander , Grashoff Carsten , Messens Joris , Ampofo Emmanuel , Laschke Matthias , Riemer Jan , Roma Leticia Prates , Schwarzländer Markus , Morgan Bruce
Published: February 2024

The NADPH/NADP+redox couple is central to metabolism and redox signalling. NADP redox state is differentially regulated by distinct enzymatic machineries at the subcellular compartment level. Nonetheless, a detailed understanding of subcellular NADP redox dynamics is limited by the availability of appropriate tools. Here, we introduce NAPstars, a family of genetically encoded, fluorescent protein-based NADP redox state biosensors. NAPstars offer real-time, specific, pH-resistant measurements, across a broad-range of NADP redox states, with subcellular resolution. We establish NAPstar measurements in yeast, plants and mammalian cell models, revealing a conserved robustness of cytosolic NADP redox homeostasis. NAPstars uncovered NADP redox oscillations linked to the cell cycle in yeast and illumination- and hypoxia-dependent NADP redox changes in plant leaves. By selectively impairing the glutathione and thioredoxin anti-oxidative pathways under acute oxidative challenge, NAPstars demonstrated an unexpected role for the glutathione system as the primary mediator of anti-oxidative electron flux that is conserved across eukaryotic kingdoms.
R-Loop Functions inBrca1-Associated Mammary Tumorigenesis
Authors: Chiang Huai-Chin , Qi Leilei , Mitra Payal , Hu Yanfen , Li Rong
Published: February 2024

Excessive R-loops, a DNA-RNA hybrid structure, are associated with genome instability andBRCA1mutation-related breast cancer. Yet the causality of R-loops in tumorigenesis remains unclear. Here we show that R-loop removal byRnaseh1overexpression (Rh1-OE) inBrca1-knockout (BKO) mouse mammary epithelium exacerbates DNA replication stress without affecting homology-directed DNA repair. R-loop removal also diminishes luminal progenitors, the cell of origin for estrogen receptor α (ERα)-negative BKO tumors. However, R-loop reduction does not dampen spontaneous BKO tumor incidence. Rather, it gives rise to a significant percentage of ERα-expressing BKO tumors. Thus, R-loops reshape mammary tumor subtype rather than promoting tumorigenesis.
Expanding the molecular grammar of polar residues and arginine in FUS prion-like domain phase separation and aggregation
Authors: Wake Noah , Weng Shuo-Lin , Zheng Tongyin , Wang Szu-Huan , Kirilenko Valentin , Mittal Jeetain , Fawzi Nicolas L
Published: February 2024

A molecular grammar governing low-complexity prion-like domains phase separation (PS) has been proposed based on mutagenesis experiments that identified tyrosine and arginine as primary drivers of phase separation via aromatic-aromatic and aromatic-arginine interactions. Here we show that additional residues make direct favorable contacts that contribute to phase separation, highlighting the need to account for these contributions in PS theories and models. We find that tyrosine and arginine make important contacts beyond only tyrosine-tyrosine and tyrosine-arginine, including arginine-arginine contacts. Among polar residues, glutamine in particular contributes to phase separation with sequence/position-specificity, making contacts with both tyrosine and arginine as well as other residues, both before phase separation and in condensed phases. For glycine, its flexibility, not its small solvation volume, favors phase separation by allowing favorable contacts between other residues and inhibits the liquid-to-solid (LST) transition. Polar residue types also make sequence-specific contributions to aggregation that go beyond simple rules, which for serine positions is linked to formation of an amyloid-core structure by the FUS low-complexity domain. Hence, here we propose a revised molecular grammar expanding the role of arginine and polar residues in prion-like domain protein phase separation and aggregation.
Breaking Barriers: pCF10 Type 4 Secretion System relies on a self-regulating muramidase to modulate the cell wall
Authors: Sun Wei-Sheng , Torrens Gabriel , Beek Josy ter , Cava Felipe , Berntsson Ronnie P-A
Published: February 2024

Conjugative Type 4 Secretion Systems (T4SS) are a main driver for the spread of antibiotic resistance genes and virulence factors in bacteria. To deliver the DNA substrate to recipient cells, it must cross the cell envelopes of both donor and recipient mating bacteria. In the T4SS from the enterococcal conjugative plasmid pCF10, PrgK is known to be the active cell wall degrading enzyme. It has 3 predicted extracellular hydrolase domains, LytM, SLT and CHAP. Here, we report the structure of the LytM domain, and show that its active site is degenerate and lacks the active site metal. Further, we show that only the predicted SLT domain is functional, and that it unexpectedly has a muramidase instead of a lytic transglycosylase activity. While we did not observe any peptidoglycan hydrolytic activity for the LytM or CHAP domain, we found that these domains downregulated the SLT muramidase activity. The CHAP domain was also found to be involved in PrgK dimer formation. Further, we show that PrgK interacts with PrgL, which likely targets PrgK to the rest of the T4SS. The presented data provides important information towards understanding the function of Gram-positive T4SSs.
The co-chaperone DNAJA2 buffers proteasomal degradation of cytosolic proteins with missense mutations
Authors: Baker Heather A , Bernardini Jonathan P , Csizmók Veronika , Madero Angel , Kamat Shriya , Eng Hailey , Lacoste Jessica , Yeung Faith Au , Comyn Sophie , Calabrese Gaetano , Raught Brian , Taipale Mikko , Mayor Thibault
Published: February 2024

Mutations can result in the loss of a protein’s native function due to protein misfolding, which is generally handled by an intricate protein quality control network. To better understand the triaging mechanisms of misfolded cytosolic proteins, we screened a human mutation library to identify a panel of unstable mutations. The degradation of these mutated cytosolic proteins is largely dependent on the ubiquitin proteasome system. Using BioID proximity labelling, we found that the co-chaperones DNAJA1 and DNAJA2 are key interactors of one of the mutated proteins. Notably, the absence of DNAJA2 increases the turnover of the mutant protein but not of the wild-type protein. Our work indicates that missense mutations in cytosolic proteins can promote interactions with molecular chaperones that normally do not occur. Assessment of the broader panel of cytosolic mutant proteins shows that the co-chaperone DNAJA2 exhibits three distinct behaviours: acting to stabilize solely the mutant, both the wild-type and mutant proteins, or being dispensable. Our work illustrates how distinct elements of the protein homeostasis network are utilized in the presence of a cytosolic misfolded protein.

Summary Statement

We identified a panel of cytosolic mutant proteins degraded by the proteasome. DNAJA2 is often required to prevent mutant protein turnover, even if it is sometimes dispensable for the wild-type protein.
Arf GTPase-Activating proteins ADAP1 and ARAP1 regulate incorporation of CD63 in multivesicular bodies
Authors: Suzuki Kasumi , Okawa Yoshitaka , Akter Sharmin , Ito Haruki , Shiba Yoko
Published: February 2024

Arf GTPase-Activating proteins (ArfGAPs) mediate the hydrolysis of GTP bound to ADP-ribosylation factors. ArfGAPs are critical for cargo sorting in the Golgi-to-ER traffic. However, the role of ArfGAPs in sorting into intralumenal vesicles (ILVs) in multivesicular bodies (MVBs) inpost-Golgi traffic remains unclear. Exosomes are extracellular vesicles (EVs) of endosomal origin. EVs mediate cell-to-cell communication, and CD63 is an EV marker. CD63 is enriched in intralumenal vesicles (ILVs) in MVBs of cells. However, the secretion of CD63 positive EVs has not been consistent with the data on CD63 localization in MVBs, and how CD63-containing EVs are formed is yet to be understood. To elucidate the mechanism of CD63 transport to ILVs, we focused on CD63 localization in MVBs and searched for the ArfGAPs involved in CD63 localization. We observed that ADAP1 and ARAP1 depletion inhibited CD63 localization to enlarged endosomes after Rab5Q79L overexpression. We tested epidermal growth factor (EGF) and CD9 localization in MVBs. We observed that ADAP1 and ARAP1 depletion affected the localization of EGF and CD9 differently. Our results indicate that there may be different populations of MVBs and that ADAP1 and ARAP1 regulate CD63 incorporation into ILVs in different MVBs.

Summary Statement

ADAP1 and ARAP1 regulate CD63 localization in endosomes.
Unlocking prophage potential:In silicoand experimental analysis of a novelMycobacterium fortuitumLysinB containing a peptidoglycan-binding domain
Authors: Das Ritam , Nadar Kanika , Arora Ritu , Bajpai Urmi
Published: February 2024

Endolysins are highly evolved bacteriophage-encoded lytic enzymes produced to damage the bacterial cell wall for phage progeny release. They offer promising potential as highly specific lytic proteins with a low chance of bacterial resistance. The diversity in lysin sequences and domain organization can be staggering.In silicoanalysis of bacteriophage and prophage genomes can help identify endolysins exhibiting unique features and high antibacterial activity, hence feeding the pipeline of narrow-spectrum protein antibiotics. Mycobacteriophage lysis cassettes mostly have two lytic enzymes, LysinA and LysinB. The enzyme LysinA targets peptidoglycan in the cell wall and possesses a modular architecture. LysinB typically contains a single domain and acts upon the mycolyl ester linkages in mycolyl-arabinogalactan-peptidoglycan (Payneet al., 2010). This study aimed to find novel LysinBs againstMycobacterium fortuitum. After a detailedin silicocharacterization of lysis cassettes from threeM. fortuitumprophages, we chose to work on a LysinB (hereafter described as LysinB_MF) found in an incomplete prophage (phiE1336, 9.4 kb in strain E1336).

LysinB_MF showed low sequence similarity with any other endolysins in the database and formed a separate clade on phylogenetic analysis. LysinB_MF’s structure, extracted from the AlphaFold Protein Structure Database, demonstrated a modular architecture with two structurally distinct domains: a peptidoglycan-binding domain (PGBD) at the N-terminal and the characteristic alpha/beta hydrolase domain connected via a linker peptide. We found the alpha/beta hydrolase domain, which is the enzyme-active domain (EAD), contains the conserved Ser-Asp-His catalytic triad with a tunnel-like topology and forms intermolecular hydrogen bonds. The PGBD shows structural similarity to the cell-wall binding domain of an amidase fromClostridium acetobutylicum,hinting at its acquisition due to domain mobility. Ourin silicoelectrostatic potential analysis suggested that PGBD might be essential to the enzyme activity. This was experimentally validated by generating a truncated version of the enzyme, which demonstrated about six-fold decreased activity compared to its native form. The antimycobacterial activity of this enzyme was also compromised in its absence. Based on our analysis, PGBD emerged as an integral constituent of enzymes with diverse functional properties and is predicted to be a conserved cross-kingdom. Overall, this study highlights the importance of mining mycobacterial prophages as a novel endolysin source. It also provides unique insights into the diverse architecture of mycobacteriophage-encoded endolysins and the importance of functional domains for their catalytic activities.

Abstract Figure
Shared and unique consequences of Joubert Syndrome gene dysfunction on the zebrafish central nervous system
Authors: Noble Alexandra R. , Masek Markus , Hofmann Claudia , Cuoco Arianna , Rusterholz Tamara D. S. , Özkoc Hayriye , Greter Nadja R. , Vladimirov Nikita , Kollmorgen Sepp , Stoeckli Esther , Bachmann-Gagescu Ruxandra
Published: February 2024

Joubert Syndrome (JBTS) is a neurodevelopmental ciliopathy defined by a highly specific midbrain-hindbrain malformation, variably associated with additional neurological features. JBTS displays prominent genetic heterogeneity with >40 causative genes that encode proteins localising to the primary cilium, a sensory organelle that is essential for transduction of signalling pathways during neurodevelopment, among other vital functions. JBTS proteins localise to distinct ciliary subcompartments, suggesting diverse functions in cilium biology. Currently, there is no unifying pathomechanism to explain how dysfunction of such diverse primary cilia-related proteins results in such a highly specific brain abnormality. In order to identify the shared consequence of JBTS gene dysfunction, we carried out transcriptomic analysis using zebrafish mutants for the JBTS-causative genescc2d2auw38, cep290fh297, inpp5ezh506, talpid3i264andtogaram1zh510and the Bardet-Biedl syndrome-causative genebbs1k742.We identified no commonly dysregulated signalling pathways in these mutants and yet all mutants displayed an enrichment of altered gene sets related to central nervous system function. We found that JBTS mutants have altered primary cilia throughout the brain, however do not display abnormal brain morphology. Nonetheless, behavioural analyses revealed reduced locomotion and loss of postural control which, together with the transcriptomic results, hint at underlying abnormalities in neuronal activity and/or neuronal circuit function. These zebrafish models therefore offer the unique opportunity to study the role of primary cilia in neuronal function beyond early patterning, proliferation and differentiation.

Summary Statement

Joubert Syndrome gene dysfunction in zebrafish leads to abnormal brain cilia, altered transcription of neuron-associated genes and abnormal swimming behaviour despite normal brain morphology.
CoRAL accurately resolves extrachromosomal DNA genome structures with long-read sequencing
Authors: Zhu Kaiyuan , Jones Matthew G. , Luebeck Jens , Bu Xinxin , Yi Hyerim , Hung King L. , Wong Ivy Tsz-Lo , Zhang Shu , Mischel Paul S. , Chang Howard Y. , Bafna Vineet
Published: February 2024

Extrachromosomal DNA (ecDNA) is a central mechanism for focal oncogene amplification in cancer, occurring in approximately 15% of early stage cancers and 30% of late-stage cancers. EcDNAs drive tumor formation, evolution, and drug resistance by dynamically modulating oncogene copy-number and rewiring gene-regulatory networks. Elucidating the genomic architecture of ecDNA amplifications is critical for understanding tumor pathology and developing more effective therapies.

Paired-end short-read (Illumina) sequencing and mapping have been utilized to represent ecDNA amplifications using a breakpoint graph, where the inferred architecture of ecDNA is encoded as a cycle in the graph. Traversals of breakpoint graph have been used to successfully predict ecDNA presence in cancer samples. However, short-read technologies are intrinsically limited in the identification of breakpoints, phasing together of complex rearrangements and internal duplications, and deconvolution of cell-to-cell heterogeneity of ecDNA structures. Long-read technologies, such as from Oxford Nanopore Technologies, have the potential to improve inference as the longer reads are better at mapping structural variants and are more likely to span rearranged or duplicated regions.

Here, we propose CoRAL (CompleteReconstruction ofAmplifications withLong reads), for reconstructing ecDNA architectures using long-read data. CoRAL reconstructs likely cyclic architectures using quadratic programming that simultaneously optimizes parsimony of reconstruction, explained copy number, and consistency of long-read mapping. CoRAL substantially improves reconstructions in extensive simulations and 9 datasets from previously-characterized cell-lines as compared to previous short-read-based tools. As long-read usage becomes wide-spread, we anticipate that CoRAL will be a valuable tool for profiling the landscape and evolution of focal amplifications in tumors.
DiverseFgfr1signaling pathways and endocytic trafficking regulate early mesoderm development
Authors: Clark James F. , Soriano Philippe
Published: February 2024

The Fibroblast growth factor (FGF) pathway is a conserved signaling pathway required for embryonic development. Activated FGF receptor 1 (FGFR1) drives multiple intracellular signaling cascade pathways, including ERK/MAPK and PI3K/AKT, collectively termed canonical signaling. However, unlikeFgfr1null embryos, embryos containing hypomorphic mutations inFgfr1lacking the ability to activate canonical downstream signals are still able to develop to birth, but exhibit severe defects in all mesodermal-derived tissues. The introduction of an additional signaling mutation further reduces the activity ofFgfr1,leading to earlier lethality, reduced somitogenesis, and more severe changes in transcriptional outputs. Genes involved in migration, ECM-interaction, and phosphoinositol signaling were significantly downregulated, proteomic analysis identified changes in interactions with endocytic pathway components, and cells expressing mutant receptors show changes in endocytic trafficking. Together, we identify processes regulating early mesoderm development by mechanisms involving both canonical and non-canonicalFgfr1pathways, including direct interaction with cell adhesion components and endocytic regulation.
Exact Analysis of the Subthreshold Variability for Conductance-Based Neuronal Models with Synchronous Synaptic Inputs
Authors: Becker Logan A. , Li Baowang , Priebe Nicholas J. , Seidemann Eyal , Taillefumier Thibaud
Published: February 2024

The spiking activity of neocortical neurons exhibits a striking level of variability, even when these networks are driven by identical stimuli. The approximately Poisson firing of neurons has led to the hypothesis that these neural networks operate in the asynchronous state. In the asynchronous state, neurons fire independently from one another, so that the probability that a neuron experience synchronous synaptic inputs is exceedingly low. While the models of asynchronous neurons lead to observed spiking variability, it is not clear whether the asynchronous state can also account for the level of subthreshold membrane potential variability. We propose a new analytical framework to rigorously quantify the subthreshold variability of a single conductance-based neuron in response to synaptic inputs with prescribed degrees of synchrony. Technically, we leverage the theory of exchangeability to model input synchrony via jump-process-based synaptic drives; we then perform a moment analysis of the stationary response of a neuronal model with all-or-none conductances that neglects postspiking reset. As a result, we produce exact, interpretable closed forms for the first two stationary moments of the membrane voltage, with explicit dependence on the input synaptic numbers, strengths, and synchrony. For biophysically relevant parameters, we find that the asynchronous regime yields realistic subthreshold variability (voltage variance ≃4–9  mV2) only when driven by a restricted number of large synapses, compatible with strong thalamic drive. By contrast, we find that achieving realistic subthreshold variability with dense cortico-cortical inputs requires including weak but nonzero input synchrony, consistent with measured pairwise spiking correlations. We also show that, without synchrony, the neural variability averages out to zero for all scaling limits with vanishing synaptic weights, independent of any balanced state hypothesis. This result challenges the theoretical basis for mean-field theories of the asynchronous state.

Published by the American Physical Society
2024
Investigation of Pseudomonas aeruginosa bacteria in a number of Baghdad schools and extent of their resistance to disinfectants and sterilizers
Authors: Al-Dulaimi Shaimaa Hadi , Hassan Miyada Kh
Published: February 2024

Purpose

This study was design to investigate of P. aeruginosa, an example of Gram-negative bacteria, in seven primary and secondary schools of Baghdad city, and the effects of Ethanol and Dettol of P. aeruginosa biofilm.

Design/methodology/approach

Seventy swabs were collected from seven primary and secondary schools of Baghdad city, Iraq, during November -December 2022. Swabs were collected from classes desk, doors handles, students hands and water taps. Standard microbiological testing methods were used on the samples for isolation and identification. The ability of bacteria to form biofilm and the effects of Ethanol and Dettol on “preformed” biofilms was examined by microtiter plate with the use of an ELISA reader.

Findings

In 70 swabs from seven primary and secondary schools, growth was observed in 33 swabs as P. aeruginosa. Primary schools were higher contaminated than secondary and water taps and door handles represented the main source of this contamination. The ability of bacteria to produce biofilm was observed in 19 (57.6%) isolates and 14 (42.4%) nonbiofilm producers. As well as, Ethanol (70%) treatment of preformed biofilms led to enhance biofilm formation and revealed significantly greater staining after 4 and 24h than Dettol (3%) compared to an untreated control (tryptic soy broth (TSB) incubation).

Originality/value

Studies on P. aeruginosa in Iraqi schools are quite rare. This work is considered distinctive because it drew attention to the presence of pathogenic bacteria within primary and secondary schools, which are not considered their natural environment.
A higher education snapshot of nutrition degree programs in Iraq: implications for public health and nutrition practice
Authors: Dodge Elizabeth , Almoayad Fatmah , Mansour Miral , Sidhu Amrita , Sajja Anusha , Benajiba Nada , Aboul-Enein Basil H.
Published: February 2024

Purpose

Burdened by undernutrition, micronutrient deficiencies and overweight and obesity as a consequence of both internal conflict and the global nutrition transition, Iraq is in need of adequate public health nutrition education to mitigate nutrition-related outcomes and risks. To address nutrition-related health outcomes, trained nutrition professionals are warranted. This paper examines current nutrition-affiliated programs offered across post-secondary institutions in Iraq.

Design/methodology/approach

An electronic review of universities and colleges’ websites, department webpages and academic programs’ homepages and resources of all the private and public universities in Iraq was conducted to find programs related to nutrition, nutrition sciences and dietetics.

Findings

All identified programs belonged to the governmental sector, were administered and financed by the Iraqi Government and were under the purview of the Iraqi Ministry of Higher Education. The review highlighted a predominant focus on food sciences in agricultural departments rather than public health or clinical nutrition. Advanced education in topics such as human metabolism, medical nutrition therapy and public health nutrition are required to adequately address over- and undernutrition in Iraq.

Originality/value

The current state of public health and nutrition-related postsecondary education in Iraq warrants an increased emphasis on clinical and public health nutrition education. Despite a commendable focus on food science studies, the country’s ongoing challenges with obesity, nutrition-related noncommunicable diseases and conflict-associated food insecurity signal an urgent need for balancing this focus with grounding in postsecondary training in public health nutrition.
Market sentiment in emerging economies: evidence from the South African property market
Authors: Kwakye Benjamin , Chan Tze-Haw
Published: February 2024

Purpose

Market sentiment has shown to influence housing prices in the global north, but in emerging economies, the nexus is rare to chance on in the current state of science for policy direction. More importantly in the recent decade where policymakers are yet to conclude on the myriad of factors confronting the housing market in sub-Saharan Africa inhibiting affordability. This paper therefore examines the impact of market sentiment on house prices in South Africa.

Design/methodology/approach

The study used the Autoregressive Distributed Lag (ARDL) approach with quarterly data spanning from 2005Q1 to 2020Q4.

Findings

In all, it was established that market sentiment plays a minimal role in the property market in South Africa. But there was enough evidence of cointegration from the bound test between sentiment and house prices. Nevertheless, the lag values of sentiment pointed to a rise in house prices. Exchange rate volatilities and inflation had a statistically significant effect on prices in both the long and short term, respectively.

Research limitations/implications

Policymakers could still monitor market sentiment in the housing market due to the strong chemistry between house prices and sentiment, as evidenced from the bound test, but focus on economic fundamentals as the main policy tool for house price reduction.

Originality/value

The findings and the creation of the sentiment index make an invaluable contribution to the paper and add to the paucity of literature on the study of market sentiment in the housing market.
How generative artificial intelligence has blurred notions of authorial identity and academic norms in higher education, necessitating clear university usage policies
Authors: Duah James Ewert , McGivern Paul
Published: February 2024

Purpose

This study examines the impact of generative artificial intelligence (GenAI), particularly ChatGPT, on higher education (HE). The ease with which content can be generated using GenAI has raised concerns across academia regarding its role in academic contexts, particularly regarding summative assessments. This research makes a unique contribution to the literature by examining university student and staff perceptions of current and future issues pertaining to the role of GenAI in universities.

Design/methodology/approach

A qualitative method involving five one-to-one semi-structured interviews with four students and a lecturer explored the ethical and practical issues of GenAI text generation in academia. An inductive thematic analysis was chosen as it provided nuanced insights aligned with the study’s goals.

Findings

Use of GenAI was discussed within the context of a range of topics, including perceptions of academic misconduct, authorial integrity and issues pertaining to university policies. Participants universally defined traditional classifications of academic misconduct but were unable to provide clear definitions where the use of GenAI was included for writing summative assessments. Students showed a more open engagement with GenAI, considering it a tool for overcoming obstacles rather than a means to plagiarise. Educators were generally more cautious and less optimistic about the academic role of GenAI. Lack of clear institutional policies surrounding such tools also contributed to ethical ambiguities.

Originality/value

The study highlights diverging perspectives between students and academics, which necessitate a forum for dialogue, ensuring the need to develop clear policies to steer the integration of GenAI in a manner that is beneficial for students and academics.
Assessing the development of green innovation in China through patent evolution: the hallmark of government policy and private enterprises
Authors: Coussa Alexandre , Gugler Philippe , Reidy Jonathan
Published: February 2024

Purpose

The purpose of this paper is to develop a comprehensive overview of green innovation (GI) in China, which is carried out by reviewing the evolution of GI from 2000 to 2019, and the main type of technology, actors and localizations. When appropriate, GI is compared to non-GI.

Design/methodology/approach

The study uses patent data from the European Patent Office database (PATSTAT); these data are processed to map trends and identify the main contributors to GI and the location of such innovation. The findings are then discussed and complemented with academic literature.

Findings

Key findings reveal an increasing divergence between GI and nongreen innovation after the 2008 crisis. It is also observed that solar energy appears to be the main component of GI in China, with a shift from photovoltaic thermal energy to solar photovoltaic energy after 2008. Other areas, such as waste management, greenhouse gases capture and climate change adaptation, are less innovative. Companies play an essential role in the development of all types of innovation. In terms of location, green patents are mainly filed in China’s three main megacities. The study also highlights the significant role of the Chinese state, which led policies shaping the trajectories and forms of GI.

Originality/value

This study expands knowledge on GI in China, highlighting its main specificities and the role of key actors. It provides to the reader a comprehensive picture of China’s green policies and innovation realities. The results can therefore be used to improve the understanding of GI evolution in China and facilitate the formulation of new research questions.