bluuebunny/crossref_metadata_2025 huggingface dataset:
The TMPRSS2 non-protease domains regulating SARS-CoV-2 Spike in mediated virus entry
Authors: Strobelt Romano , Adler Julia , Shaul Yosef
Published: October 2023

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) enters cells by binding to the angiotensin-converting enzyme 2 (hACE2) receptor. This process is aided by the transmembrane protease serine 2 (TMPRSS2), which enhances entry efficiency and infectiousness by cleaving the SARS-CoV-2 surface glycoprotein (Spike). The cleavage primes the Spike protein, promoting membrane fusion instead of receptor-mediated endocytosis. Despite the pivotal role played by TMPRSS2, our understanding of its non-protease distinct domains remains limited. In this report, we present evidence indicating the potential phosphorylation of a minimum of six tyrosine residues within the cytosolic tail (CT) of TMPRSS2. Through the use of TMPRSS2 CT phospho-mimetic mutants, we observed a reduction in TMPRSS2 protease activity, accompanied by a decrease in SARS-CoV-2 pseudovirus infection, which was found to occur mainly via the endosomal pathway. We expanded our investigation beyond TMPRSS2 CT and discovered the involvement of other non-protease domains in regulating infection. Our co-immunoprecipitation experiments demonstrated a strong interaction between TMPRSS2 and Spike. We revealed a 21 amino acid long TMPRSS2-Spike-binding region (TSBR) within the TMPRSS2 scavenger receptor cysteine-rich (SRCR) domain that contributes to this interaction. Our study sheds light on novel functionalities associated with TMPRSS2’s cytosolic tail and SRCR region. Both of these regions have the capability to regulate SARS-CoV-2 entry pathways. These findings contribute to a deeper understanding of the complex interplay between viral entry and host factors, opening new avenues for potential therapeutic interventions.
Experimental evolution of symbiotic microbes without their partners can imply the presence of cooperative or antagonistic adaptations
Authors: Larsen Tyler J. , Jefferson Cara , Bartley Anthony , Strassmann Joan E. , Queller David C.
Published: October 2023

Microbes adapt to the presence of other species, but the fitness consequences of specific interactions are difficult to study in their natural context. We experimentally evolved symbiotic microbes in an artificial environment without access to the partners with whom they interact in nature. As organisms will tend to lose adaptations that they do not need due to drift or pleiotropic tradeoffs, we expect normally symbiotic microbes evolved in isolation to lose adaptations to help or harm their natural partners. The direction and magnitude of such changes can suggest whether the microbes had historically been selected to help or harm one another. We apply this method to the symbiosis between the social amoebaDictyostelium discoideumand three intracellular bacterial endosymbionts,Paraburkholderia agricolaris, P. hayleyella,andP. bonniea.A minority of strains ofParaburkholderiaandD. discoideumevolved differences in their effects on one another’s fitnesses, implying the existence of adaptations to one another that were lost when no longer relevant. Our results suggest that the degree to whichD. discoideumandParaburkholderiahave adapted to help or harm one another can differ substantially between strains within each species, with some strains appearing to have a historically adversarial relationship, some strains a more benign relationship, and many strains no clear adaptations to one another at all. Our results underscore the complexity of microbial interactions in nature and suggest experimental evolution under relaxed selection is a potentially useful approach for studying adaptation in microbes.
Decision-Making Time Cells in Hippocampal Dorsal CA1
Authors: Ma M. , Simoes de Souza F. , Futia G.L. , Anderson S.R. , Riguero J. , Tollin D. , Gentile-Polese A. , Platt J.P. , Hiratani N. , Gibson E. A. , Restrepo D.
Published: October 2023

Sequential neural dynamics encoded by “time cells” play a crucial role in hippocampal function. However, the role of hippocampal sequential neural dynamics in associative learning is an open question. In this manuscript, we used two-photon Ca2+imaging of dorsal CA1 pyramidal neurons in head-fixed mice performing a go-no-go associative learning task. We found that pyramidal cells responded differentially to the rewarded or unrewarded stimuli. The stimuli were decoded accurately from the activity of the neuronal ensemble, and accuracy increased substantially as the animal learned to differentiate the stimuli. Decoding the stimulus from individual pyramidal cells that responded differentially revealed that decision-making took place at discrete times after stimulus presentation. Lick prediction decoded from the ensemble activity of cells in dCA1 correlated linearly with lick behavior indicating that sequential activity of pyramidal cells in dCA1 constitutes a temporal memory map used for decision-making in associative learning.
The protein phosphorylation landscape in photosystem I of the desert algae Chlorella sp.
Authors: Levin Guy , Yasmin Michael , Pienko Tomasz , Yehishalom Nitsan , Glaser Fabian , Schuster Gadi
Published: October 2023

The phosphorylation of photosystem II (PSII) and its antenna (LHCII) proteins has been extensively studied and its involvement in state transitions and PSII repair is well known. Yet, very little is known about the extent and functions of phosphorylation of photosystem I (PSI) and its antenna (LHCI) proteins. Here, two proteomics methods were applied to generate a detailed map of the phosphorylation sites of the PSI-LHCI proteins in Chlorella ohadii cells that were grown under low- or extreme high-light intensities (LL and HL). Furthermore, we analyzed the content of oxidized tryptophans in these cell types to estimate light-induced oxidative damage to PSI-LHCI.Our work revealed the phosphorylation of 11 out of 22 PSI-LHCI subunits. The analyses detected extensive phosphorylation of the LHCI subunits lhca6 and lhca7. Other PSI-LHCI subunits were phosphorylated to a lesser extent. Additionally, we show the accumulation of oxidatively damaged tryptophans in the psaD subunit of PSI of HL-grown C. ohadii. The significant phosphorylation of lhca6 and lhca7 suggests a physiological role during photosynthesis, possibly by altering light-harvesting characteristics and binding of other LHCI subunits. Moreover, we show that psaD is susceptible to photodamage while LHCI is protected from ROS under HL.
Experiences and impacts of side effects among contraceptive users in the UK: exploring individual narratives of contraceptive side effects
Authors: Stewart Catherine , Stevens Rose , Kennedy Fiona , Cecula Paulina , Rueda Carrasco Elena , Hall Jennifer
Published: October 2023

Purpose

While many women worldwide use contraception, there is a paucity of research on individual experiences of side effects and their impacts. To address this gap, we analysed free-text responses of contraception experiences from 337 women aged 18 to 35, based in the UK who took part in an online survey on contraception.

Materials and methods

Through a directed content analysis approach, we developed a coding framework based on existing literature and initial response review. It included six themes; method(s) of contraception, side effect(s) experienced, impact of side effect(s), timing of side effect(s), interactions with healthcare practitioners, and trial and error.

Results

Side effect experiences and impacts varied greatly between individuals and contraceptives. Most participants described negative effects, such as mental health issues and bleeding problems. Some shared positive experiences related to bleeding management and the absence of side effects. Some experienced side effects after years of use and felt unheard by practitioners.

Conclusions

This contraceptive experience variability underscores the need for further research into individual side effect variation. We advocate for a patient-centred approach to contraceptive counselling. Practitioners should play an active role in improving contraception prescription, acknowledging the diverse experiences and preferences of patients.
The hidden hand of asymptomatic infection hinders control of neglected tropical diseases
Authors: Rock Kat S. , Chapman Lloyd A.C. , Dobson Andrew P. , Adams Emily R. , Hollingsworth T. Déirdre
Published: October 2023

Neglected tropical diseases are responsible for considerable morbidity and mortality in low-income populations. International efforts have reduced their global burden, but transmission is persistent and case-finding-based interventions rarely target asymptomatic individuals. We develop a generic mathematical modelling framework for analysing the dynamics of visceral leishmaniasis in the Indian sub-continent (VL),gambiensesleeping sickness (gHAT), and Chagas disease and use it to assess the possible contribution of asymptomatics who later develop disease (pre-symptomatics) and those who do not (non-symptomatics) to the maintenance of infection. Plausible interventions, including active screening, vector control and reduced time to detection, are simulated for the three diseases. We conclude that currently available tools could bring new infections under control for gHAT and Chagas disease – albeit over a long period of time for Chagas disease – but that VL will be more difficult to control with existing interventions. gHAT protocols that allow for treatment of parasitologically positive asymptomatics result in better control of infection compared to syndromic diagnosis required for VL. This work highlights that the high asymptomatic contribution to transmission for Chagas and gHAT and the apparently high basic reproductive number of VL may greatly undermine long-term control, but that precise predictions are hampered by uncertainty in natural history.
Respiratory viral infections do not increase risk of venous thromboembolism
Authors: Korzeniewski Steven J. , Bauer Samantha J. , Kabrhel Christopher , Mark Courtney D. , Ka ming Ngai , Kelly Christopher , Camargo Carlos A. , Kline Jeffrey A.
Published: October 2023

Background

A major concern in emergency departments is whether acute respiratory illness (ARI) is associated with increased risk of venous thromboembolism (VTE).

Methods

This prospective cohort study includes ARI patients from the 91-hospital, U.S. CDC- sponsored Respiratory Virus Laboratory Emergency Department Network Surveillance (RESP-LENS) program from January 2022 to June 2023. We calculated incidence rates and used multivariable regression models to test the null hypothesis that there is no association between the results or absence of laboratory viral testing and the risk of new onset VTE within 30-days.

Findings

Out of 620,303 ARI encounters, 65% underwent laboratory viral testing; 13% tested positive for COVID and 3%-4% tested positive for influenza-A and/or RSV. The 30-day VTE incidence rate was 0.70% amongunique first patient encounters and 0.82%overall.

The highest VTE incidence rate occurred among viral test negative patient encounters (1.13% [95%CI 1.08%-1.19%]) and the lowest VTE rate occurred among patients who tested positive for COVID, influenza-A or RSV, or who did not undergo viral testing (95%CI 0.09%-0.86%). In adjusted models, only among patients receiving any ICU care was fluA associated with heightened VTE risk.

New VTE was associated with increased 30-day mortality risk (RR 2.7, 95%CI 2.3-3.2), but there was no difference in 30-day mortality risk among VTE patients grouped by the results of viral testing (interaction p > 0.05).

Conclusion

In the U.S. from January 2022 to June 2023, laboratory confirmed viral infection was not associated with increased risk of short-term VTE diagnosis or death among patients seeking emergency care.
Genetic or pharmacological inactivation of CREBBP sensitizes B-cell Acute Lymphoblastic Leukemia to Ferroptotic Cell Death upon BCL2 Inhibition
Authors: Garcia-Gimenez Alicia , Ditcham Jonathan E. , Azazi Dhoyazan M.A. , Meduri Eshwar , Asby Ryan , Sakakini Nathalie , Lopez Cecile K. , Narayan Nisha , Beinortas Tumas , Bagri Jaana , Singh Shuchi Agrawal , Giotopoulos George , Murphy Michael P. , Horton Sarah J. , Huntly Brian J.P. , Richardson Simon E.
Published: October 2023

B-cell acute lymphoblastic leukemia (B-ALL) is a leading cause of death in childhood and outcomes in adults remain dismal. There is therefore an urgent clinical need for therapies that target the highest risk cases. Mutations in the histone acetyltransferaseCREBBPassociate with high-risk features in B-ALL and have been implicated in chemoresistance. We performed a targeted drug screen in isogenic human cell lines, identifying a number of actionable small molecules that specifically targetCREBBP-mutated B-ALL. The most potent was the BCL2 inhibitor Venetoclax, which acts through a non-canonical mechanism resulting in ferroptotic cell death.CREBBP-mutated cell lines showed differences in cell-cycle, metabolism and response to oxidative stress. Lastly, we demonstrate that small-molecule inhibition of CREBBP sensitizes B-ALL cells, regardless of genotype, to Venetoclax-induced ferroptosisin-vitroandin-vivo, providing a potential novel drug combination for broader clinical translation in B-ALL.
Numerical analysis of flow anisotropy in rotated-square deterministic lateral displacement devices at moderate Reynolds number
Authors: Mallorie Calum , Vernekar Rohan , Owen Benjamin , Inglis David W. , Krüger Timm
Published: October 2023

Deterministic lateral displacement (DLD) is a microfluidic method for accurately separating particles by size or deformability. Recent efforts to operate DLD devices in the inertial, rather than in the Stokes, flow regime have been hindered by a loss of separation efficiency and difficulty predicting the separation behaviour. One factor contributing to these problems is the onset of inertia-induced flow anisotropy where the average flow direction does not align with the direction of the pressure gradient in the device. We use the lattice-Boltzmann method to simulate two-dimensional flow through a rotated-square DLD geometry with circular pillars at Reynolds number up to 100 for different gap sizes and rotation angles. We find that anisotropy in this geometry is a non-monotonous function of Reynolds number and can be positive or negative. This finding is in contradiction to the naive expectation that inertia would always drive flow along principal direction of the pillar array. Anisotropy tends to increase in magnitude with gap size and rotation angle. By analysing the traction distribution along the pillar surface, we explain how the change of the flow field upon increasing inertia leads to the observed trends of anisotropy. Our work contributes to a better understanding of the inertial flow behaviour in ordered cylindrical porous media, and might contribute to improved DLD designs for operation in the inertial regime.
Action observation reveals a network with divergent temporal and parietal lobe engagement in dogs compared to humans
Authors: Boch Magdalena , Karl Sabrina , Wagner Isabella C. , Lengersdorff Lukas L. , Huber Ludwig , Lamm Claus
Published: October 2023

Action observation is a fundamental pillar of social cognition. Neuroimaging research has revealed a human and primate action observation network (AON) encompassing fronto-temporo-parietal areas with links to a species’ imitation tendencies and relative lobe expansion. Dogs (Canis familiaris)have good action perception and imitation skills and a less expanded parietal than temporal lobe, but their AON remains unexplored. We conducted a functional MRI study with 28 dogs and 40 humans and found functionally analogous involvement of somatosensory and temporal brain areas of both species’ AONs and responses to transitive and intransitive action observation in line with their imitative skills. However, activation and task-based functional connectivity measures suggested significantly less parietal lobe involvement in dogs than in humans. These findings advance our understanding of the neural bases of action understanding and the convergent evolution of social cognition, with analogies and differences resulting from similar social environments and divergent brain expansion, respectively.
Deep mutational scanning reveals a tight correlation between protein degradation and toxicity of thousands of non-native aspartoacylase protein variants
Authors: Grønbæk-Thygesen Martin , Voutsinos Vasileios , Johansson Kristoffer E. , Schulze Thea K. , Cagiada Matteo , Pedersen Line , Clausen Lene , Nariya Snehal , Powell Rachel L. , Stein Amelie , Fowler Douglas M. , Lindorff-Larsen Kresten , Hartmann-Petersen Rasmus
Published: October 2023

When the structural stability of a protein is compromised, the protein may form non-native interactions with other cell proteins and thus becomes a hazard to the cell. To mitigate this danger, destabilized proteins are targeted by the cellular protein quality control (PQC) network, which either corrects the folding defect or targets the protein for degradation. However, the details of how the protein folding and degradation systems collaborate to combat potentially toxic non-native proteins are unknown. To address this issue, we performed systematic studies on destabilized variants of the cytosolic aspartoacylase, ASPA, where loss-of-function variants are linked to Canavan’s disease, an autosomal recessive and lethal neurological disorder, characterized by the spongy degeneration of the white matter in the brain. Using Variant Abundance by Massively Parallel sequencing (VAMP-seq), we determined the abundance of 6152 out of the 6260 (∼98%) possible single-site missense and nonsense ASPA variants in cultured human cells. The majority of the low abundance ASPA variants are degraded through the ubiquitin-proteasome system (UPS) and become toxic upon prolonged expression. Variant cellular abundance data correlates with predicted thermodynamic stability, evolutionary conservation, and separates most known disease-linked variants from benign variants. Systematic mapping of degradation signals (degrons) shows that inherent primary degrons in ASPA are located in buried regions, and reveals that the wild-type ASPA C-terminal region functions as a degron. Collectively, our data can be used to interpret Canavan’s disease variants and also offer mechanistic insight into how ASPA missense variants are targeted by the PQC system. These are essential steps towards future implementation of precision medicine for Canavan’s disease.
A novel high throughput microwell outgrowth assay for HIV infected cells
Authors: Fenton Anthony D , Archin Nancie , Turner Anne-Marie , Joseph Sarah , Moeser Matthew , Margolis David M , Browne Edward P
Published: October 2023

Although antiretroviral therapy (ART) is highly effective at suppressing HIV replication, a viral reservoir persists that can reseed infection if ART is interrupted. Curing HIV will require elimination or functional containment of this reservoir, but the size of the HIV reservoir is highly variable between individuals. To evaluate the overall size of the HIV reservoir, several assays have been developed, including PCR based assays for viral DNA, the Intact Proviral DNA Assay (IPDA), and the Quantitative Viral Outgrowth Assay (QVOA). QVOA is the gold standard assay for measuring inducible replication competent proviruses, but this assay is technically challenging and time consuming. To provide a more rapid and less laborious tool for quantifying cells infected with replication competent HIV, we developed the Microwell Outgrowth Assay (MOA), in which HIV infected CD4 T cells are cocultured with an HIV-detecting reporter cell line in a polydimethylsiloxane (PDMS)/polystyrene array of nanoliter sized wells (rafts). Transmission of HIV from infected cells to the reporter cell line induces fluorescent reporter protein expression that is detected by automated scanning across the array. We show that this assay can detect HIV infected cells with a high degree of sensitivity and precision. Using this approach, we were able to detect HIV infected cells from ART-naïve people with HIV (PWH) and from PWH on ART. Furthermore, we demonstrate that infected cells can be recovered from individual rafts and used to analyze the diversity of viral sequences. This assay may be a useful tool for quantifying and characterizing infected cells from PWH.

Author summary

Measuring the size of the HIV reservoir in people with HIV (PWH) will be important for determining the impact of HIV cure strategies. However, measuring this reservoir is challenging. We report a new method for quantifying HIV infected cells that involves culturing cells from PWH in an array of microwells with a cell line that detects HIV infection. We show that this approach can detect rare HIV infected cells and derive detailed virus sequence information for each infected cell.
Differences in white matter detected byex vivo9.4T MRI are associated with axonal changes in the R6/1 model of Huntington’s Disease
Authors: Casella C. , Kelly B. , Murillo Bartolome A. , Mills-Smith B. , Parker G.D. , Von Ruhland C. , Syed Y.A. , Dion V. , Rosser A.E. , Metzler-Baddeley C. , Jones D.K. , Lelos M.J.
Published: October 2023

White matter (WM) volume loss has been reported in people with Huntington’s disease (HD), but the cellular basis of this deficit remains to be elucidated. To address this, we assessed ex vivo WM microstructure in the transgenic R6/1 mouse model of HD with magnetic resonance imaging (MRI) and studied the neurobiological basis of the MRI brain signals with histological and electron microscopy analyses in a separate cohort of age- and sex-matched mice. Differences in the macromolecular proton fraction (MPF) from quantitative magnetization transfer (qMT) as a proxy myelin measure, and the intra-axonal signal fraction (FR) from the composite hindered and restricted model of diffusion (CHARMED) as a proxy marker of axon density, were assessed alongside diffusion tensor imaging (DTI) parameters. A tractometry approach was employed to inspect region-specific differences across the corpus callosum (CC). Furthermore, voxel-based morphometry (VBM) and tract-based spatial statistics (TBSS) were used to explore brain-wise WM macro- and microstructure abnormalities. To gain insight into disease-associated impairments in attentional and visuospatial processing, a third cohort of age-matched mice was assessed with the 5-choice serial reaction time task (5-CSRTT). We report cognitive impairments in R6/1 mice and, by evaluating MRI and light and electron microscopy results, we show that this HD mouse model presents disruptions in axonal morphology (i.e. less complex, thinner axons) and organization (i.e. more densely packed axons). Furthermore, we show that, at least early in disease progression, R6/1 mice present a reduction in the expression or content of myelin-associated proteins without significant alterations in the structure of myelin sheaths. Finally, our findings indicate that neuroinflammation-driven glial and axonal swelling might also affect this mouse model early in disease progression. Crucially, we demonstrate the potential of FR, an in vivo estimate of axon density, as a novel MRI biomarker of HD-associated changes in WM microstructure.
Drivers of thermal tolerance breadth of plants across contrasting biomes: do mean or seasonality in climate indices matter more?
Authors: Briceno Veronica F. , Arnold Pieter Andrew , Cook Alicia M. , Courtney Jones Stephanie K. , Gallagher Rachael V. , French Kris , Bravo Leon A. , Nicotra Adrienne B. , Leigh Andy
Published: October 2023

1.The Climate Variability Hypothesis (CVH) predicts that species from environments with more variable temperatures should have wide thermal tolerance breadth. We addressed this question in plants and asked which local climate predictors (including precipitation, mean and extreme temperatures and thermal variability) best predict species physiological thermal limits. 2. Measures of low (Tcrit-cold) and high (Tcrit-hot) photosystem II thermal tolerance thresholds were used to determine thermal tolerance breadth (TTB), along with ice nucleation temperature (Tnucleation, freezing tolerance) of plant species sampled from three contrasting biomes: alpine, desert and wet temperate. 3. Thermal tolerance metrics Tcrit-cold, Tnucleation, Tcrit-hot and TTB all differed among biomes. Notably, desert species had the most cold and heat tolerant leaves, and therefore the widest TTB, whereas species in alpine and temperate biomes had similar TTB. For plants in all biomes, TTB exceeded the thermal range of their local climate. 4. Overall, two Principal Component axes of local climate drivers explained substantial variation in all tolerance metrics. Extreme hot, dry climates improved freezing and heat tolerance. High thermal variability and low minimum temperatures also improved freezing tolerance, but were unrelated to heat tolerance or TTB. Species explained a significant amount of variation among all metrics, but this was not due to phylogenetic relatedness. A small portion of the variation in physiological thermal thresholds was explained by growth form. Unexplained variation could be due to microclimate-driven plasticity, leaf traits or thermoregulatory mechanisms. 5. Synthesis. Our results provide some support for the climate variability hypothesis in plants: photosynthetic thermal tolerance breadth was greatest in more thermally variable biomes. This relationship was largely driven by cold tolerance, with variation in heat tolerance explained better by mean and extreme temperatures. Therefore, we conclude that, the CVH alone is not sufficient to explain variation in plant thermal tolerance, with many aspects of climate being potentially important drivers.
Region-specific gene expression changes associated to oleoylethanolamide-induced attenuation of alcohol self-administration
Authors: González-Portilla Macarena , Montagud-Romero Sandra , Mellado Susana , Rodríguez de Fonseca Fernando , Pascual María , Rodríguez-Arias Marta
Published: October 2023

Oleoylethanolamide (OEA) is a lipid with anti-inflammatory activity that modulates multiple reward-related behaviors. Previous studies have shown that OEA treatment reduces alcohol self-administration (SA) while inhibiting alcohol-induced inflammatory signaling. Nevertheless, the specific mechanisms that OEA is targeting to achieve these effects have not been widely explored. Here, we tested the effects of OEA treatment during alcohol SA, extinction or previous to cue-induced reinstatement of alcohol seeking. In addition, we measured gene expression changes in the striatum and hippocampus of relevant receptors for alcohol consumption (DrD1, DrD2, CNR1, OPRM1) as well as immune-related proteins (IL-6, IL-1β, TLR4) and the brain-derived neurotrophic factor (BDNF). Our results confirmed that when administered contingently, systemic OEA administration reduced alcohol SA and attenuated cue-induced reinstatement. Interestingly, we also observed that OEA treatment reduced the number of sessions needed for extinction of alcohol seeking. Biochemical analyses showed region-specific OEA dopaminergic and cannabinoid receptor gene expression alterations. Also, OEA treatment modulated the long-term immune response and increased BDNF expression. These results suggest that boosting OEA levels may be an effective strategy for reducing alcohol SA and preventing relapse.
De-suppression of mesenchymal cell identities and variable phenotypic outcomes associated with knockout ofBbs1
Authors: Freke Grace , Martins Tiago , Davies Rosie , Beyer Tina , Seda Marian , Peskett Emma , Haq Naila , Prasai Avishek , Otto Georg , Srikaran Jeshmi Jeyabalan , Hernandez Victor , Diwan Gaurav , Russell Robert , Ueffing Marius , Huranova Martina , Boldt Karsten , Beales Philip L. , Jenkins Dagan
Published: October 2023

Bardet-Biedl syndrome (BBS) is an archetypal ciliopathy caused by dysfunction of primary cilia. BBS affects multiple tissues, including the kidney, eye and hypothalamic satiety response. Understanding pan-tissue mechanisms of pathogenesis versus those which are tissue specific, and gauging their associated inter-individual variation owing to genetic background and stochastic processes, is of paramount importance in syndromology. The BBSome is a membrane trafficking and intraflagellar transport (IFT) adaptor protein complex formed by 8 BBS proteins, including BBS1, which is the most commonly mutated gene in BBS. To investigate disease pathogenesis we generated a series of clonal renal collecting duct IMCD3 cell lines carrying defined biallelic nonsense or frameshift mutations inBbs1, as well as a panel of matching wild-type CRISPR control clones. Using a phenotypic screen and an unbiased multi-omics approach we note significant clonal variability for all assays, emphasising the importance of analysing panels of genetically-defined clones. Our results suggest that BBS1 is required for suppression of mesenchymal cell identities as IMCD3 cell passage number increases. This was associated with a failure to express epithelial cell markers and tight junction formation, which was variable amongst clones. Transcriptomic analysis of hypothalamic preparations from BBS mutant mice, and BBS patient fibroblasts, suggested that dysregulation of epithelial-to-mesenchymal transition (EMT) genes is a general predisposing feature of BBS across tissues. Collectively this work suggests that the dynamic stability of the BBSome is essential for suppression of mesenchymal cell identities as epithelial cells differentiate.
Recognition of the Nucleotide-Dependent Feature Facilitates the Microtubule End-Binding of MCAK
Authors: Chen Wei , Song Yin-Long , He Jian-Feng , Liang Xin
Published: October 2023

The growing plus-end is a key regulatory site for microtubule dynamics. MCAK (mitotic centromere-associated kinesin), a microtubule depolymerizing kinesin, is an end-binding regulator of catastrophe frequency. It is intriguing how MCAK specifically binds to dynamic microtubule ends. Here, we measure the end-binding kinetics of MCAK using single-molecule imaging and reveal the end-binding preference. MCAK binds to the entire GTP cap, including the EB cap and the distalmost cap. Further analysis shows that MCAK strongly binds to GTPγS microtubules, suggesting that it could recognize the nucleotide-dependent feature of microtubules. Moreover, the binding preference is independent on the nucleotide state of MCAK, and this feature facilitates the high-affinity end-binding of MCAK. Finally, we show that despite partially sharing the binding regions, MCAK and XMAP215 function in an additive manner, demonstrating a simple logic of how the end-binding regulators work in co-ordination. In all, our results provide novel insights into understanding how MCAK regulates the dynamics of microtubule ends.
Robustness and adaptability of sensorimotor skills in expert piano performance
Authors: Yasuhara Masaki , Uehara Kazumasa , Oku Takanori , Shiotani Sachiko , Nambu Isao , Furuya Shinichi
Published: October 2023

Skillful execution of sequential actions requires the delicate balance of sensorimotor control, encompassing both robustness and adaptability. Previous studies have characterized behavioral and electrophysiological responses to sensory perturbation during performance of sequential movements such as speech and singing. However, it remains unknown whether and in what manner both motor and neural responses, triggered by sensory perturbation, undergo plastic adaptation as a consequence of extensive sensorimotor experience. Here, we addressed this question by comparing effects of transiently delayed tone production on the spatiotemporal patterns of the subsequent motor actions and event-related potentials (ERPs) during fast and accurate piano performance between expert pianists and musically-untrained individuals (non-musicians). Following the delayed tone production, the inter-keystroke interval was abnormally prolonged in non-musicians but not in pianists. By contrast, the keystroke velocity following the tone delay was increased only in the pianists. A regression model further demonstrated that the change in the inter-keystroke interval following the perturbation covaried with the ERPs of the N180 and P300 components particularly at the frontal and parietal regions. In contrast, the alteration in the keystroke velocity was associated with the P300 component of the temporal region ipsilateral to the moving hand, which suggests enhancement of auditory but not somatosensory feedback gain following auditory perturbation. Together, these findings suggest that distinct neural mechanisms underlie robust and adaptive sensorimotor skills individuals with different levels of proficiency.
hiPSC-CM Electrophysiology: Impact of Temporal Changes and Study Parameters on Experimental Reproducibility
Authors: Guerrelli Devon , Pressman Jenna , Posnack Nikki
Published: October 2023

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are frequently used for preclinical cardiotoxicity testing and remain an important tool for confirming model-based predictions of drug effects in accordance with the Comprehensivein VitroProarrhythmia Assay (CiPA) initiative. Despite the considerable benefits hiPSC-CMs provide, concerns surrounding experimental reproducibility have emerged. Our study aimed to investigate the effects of temporal changes and experimental parameters on hiPSC-CM electrophysiology. hiPSC-CMs (iCell cardiomyocyte2) were cultured for 14 days and biosignals were acquired using a microelectrode array (MEA) system. Continuous recordings revealed a 22.6% increase in the beating rate and 7.7% decrease in the field potential duration (FPD) during a 20-minute equilibration period. Location specific differences across a multiwell plate were also observed, with hiPSC-CMs in the outer rows beating 8.8 beats per minute (BPM) faster than the inner rows. Cardiac endpoints were also impacted by cell culture duration; from 2-14 days the beating rate decreased (–12.7 BPM), FPD lengthened (+257 ms), and spike amplitude increased (+3.3 mV). Cell culture duration (4-10 days) also impacted hiPSC-CM drug responsiveness (E-4031, nifedipine, isoproterenol). Our study highlights multiple sources of variability that should be considered and addressed when performing hiPSC-CM MEA studies. To improve reproducibility and data interpretation, MEA-based studies should establish a standardized protocol and report key experimental conditions (e.g., culture time, equilibration time, electrical stimulation settings, report raw data values).

New & Noteworthy

We demonstrate that hiPSC-CM electrophysiology measurements are significantly impacted by slight deviations in experimental techniques including electrical stimulation protocols, equilibration time, well-to-well variability, and length of hiPSC-CM culture. Furthermore, our results indicate that hiPSC-CM drug responsiveness changes within the first two weeks following defrost.

Graphical Abstract

Image created with Biorender.com
Biocalcification in porcelaneous foraminifera
Authors: Dubicka Zofia , Tyszka Jarosław , Pałczyńska Agnieszka , Höhne Michelle , Bijma Jelle , Janse Max , Klerks Nienke , Bickmeyer Ulf
Published: October 2023

Living organisms control the formation of mineral skeletons and other structures through biomineralization. Major phylogenetic groups usually consistently follow a single biomineralization pathway. Foraminifera, which are very efficient marine calcifiers, making a substantial contribution to global carbonate production and global carbon sequestration, are regarded as an exception. This phylum has been commonly thought to follow two contrasting models of eitherin situ“mineralization of extracellular matrix” attributed to hyaline rotaliid shells, or “mineralization within intracellular vesicles” attributed to porcelaneous miliolid shells. Our previous results on rotaliids along with those on miliolids in this paper question such a wide divergence of biomineralization pathways within the same phylum of Foraminifera. We found that both groups produced calcareous shells via the intravesicular formation of unstable mineral precursors (Mg-rich amorphous calcium carbonates) supplied by endocytosed seawater and deposited at the site of new wall formation within the organic matrix. Precipitation of high-Mg calcitic mesocrystals took placein situand formed a dense, chaotic meshwork of needle-like crystallites. We did not observe deposition of calcified needles that had already precipitated in the transported vesicles, which challenges the previous model of miliolid mineralization. Hence, Foraminifera utilize less divergent calcification pathways, following the recently discovered biomineralization principles. Mesocrystalline chamber walls are therefore apparently created by accumulating and assembling particles of pre-formed liquid amorphous mineral phase within the extracellular organic matrix enclosed in a biologically controlled privileged space by active pseudopodial structures. Both calcification pathways evolved independently in the Paleozoic and are well-conserved in two clades that represent different chamber formation modes.
Tapping into the nativePseudomonasBacterial Biofilm Structure by High-Resolution 1D and 2D MAS solid-state NMR
Authors: Byeon Chang-Hyeock , Kinney Ted , Saricayir Hakan , Srinivasa Sadhana , Wells Meghan K. , Kim Wook , Akbey Ümit
Published: October 2023

We present a high-resolution 1D and 2D magic-angle spinning (MAS) solid-state NMR (ssNMR) study to characterize nativePseudomonas fluorescenscolony biofilms at natural abundance without isotope-labelling. By using a high-resolution INEPT-based 2D1H-13C ssNMR spectrum and thorough peak deconvolution approach at the 1D ssNMR spectra, approximately 80/134 (in 1D/2D) distinct biofilm chemical sites were identified. We compared CP and INEPT13C ssNMR spectra to different signals originating from the mobile and rigid fractions of the biofilm, and qualitative determined dynamical changes by comparing CP buildup behaviors. Protein and polysaccharide signals were differentiated and identified by utilizing FapC signals as a template, a biofilm forming functional amyloid fromPseudomonas. We also attempted to identify biofilm polysaccharide species by using1H/13C chemical shifts obtained from the 2D spectrum. This study marks the first demonstration of high-resolution 2D ssNMR spectroscopy for characterizing native bacterial biofilms and expands the scope of ssNMR in studying biofilms. Our experimental pipeline can be readily applied to other in vitro biofilm model systems and natural biofilms and holds the promise of making a substantial impact on biofilm research, fostering new ideas and breakthroughs to aid in the development of strategic approaches to combat infections caused by biofilm-forming bacteria.
Distinct features of ribonucleotides within genomic DNA in Aicardi-Goutières syndrome (AGS)-ortholog mutants ofSaccharomyces cerevisiae
Authors: Kundnani Deepali L. , Yang Taehwan , Gombolay Alli L. , Mukherjee Kuntal , Newnam Gary , Meers Chance , Mehta Zeel H. , Mouawad Celine , Storici Francesca
Published: October 2023

Ribonucleoside monophosphates (rNMPs) are abundantly found within genomic DNA of cells. The embedded rNMPs alter DNA properties and impact genome stability. Mutations in ribonuclease (RNase) H2, a key enzyme for rNMP removal, are associated with the Aicardi-Goutières syndrome (AGS), a severe neurological disorder. Here, we engineered two AGS-ortholog mutations inSaccharomyces cerevisiae:rnh201-G42S andrnh203-K46W. Using the ribose-seq technique and the Ribose-Map bioinformatics toolkit, we unveiled rNMP abundance, composition, hotspots, and sequence context in these yeast AGS-ortholog mutants. We found higher rNMP incorporation in the nuclear genome ofrnh201-G42S than in wild-type andrnh203-K46W-mutant cells, and an elevated rCMP content in both mutants. Moreover, we uncovered unique rNMP patterns in each mutant, highlighting a differential activity of the AGS mutants towards rNMPs embedded on the leading or on the lagging strand of DNA replication. This study guides future research on rNMP characteristics in human genomic samples carrying AGS mutations.
Building on-chip cytoskeletal circuits via branched microtubule networks
Authors: Zaferani Meisam , Song Ryungeun , Petry Sabine , Stone Howard A.
Published: October 2023

Controllable platforms to engineer robust cytoskeletal scaffolds have the potential to create novel on-chip nanotechnologies. Inspired by axons, we combined the branching microtubule (MT) nucleation pathway with microfabrication to develop “cytoskeletal circuits”. This active matter platform allows control over the adaptive self-organization of uniformly polarized MT arrays via geometric features of microstructures designed within a microfluidic confinement. We build and characterize basic elements, including turns and divisions, as well as complex regulatory elements, such as biased division and MT diodes, to construct various MT architectures on a chip. Our platform could be used in diverse applications, ranging from efficient on-chip molecular transport to mechanical nano-actuators. Further, cytoskeletal circuits can serve as a tool to study how the physical environment contributes to MT architecture in living cells.

Significance

Microtubules have essential functions within the cell, including providing a robust railroad for motor-driven cargo transport. The unique properties of microtubules have stimulated attempts to harness these characteristics for targeted delivery of molecular complexes, novel material design, and developing nanotechnologies with precision comparable to living organisms. However, Previous efforts mainly focused on microtubules with fixed length and layout and no controlled MT generation, setting a limit on designing MT architecture. In this study, we integrated nanofabrication with microtubule branching reactions borrowed directly from the cell’s toolkit to construct cytoskeletal circuits and generate microtubule architectures from scratch. That is, our system enables control over microtubule growth and autocatalytic nucleation on a microfluidic chip with micro/nanostructures patterned within.
Cytochrome P450 and Epoxide Hydrolase Metabolites in Aβ and tau-induced Neurodegeneration: Insights fromCaenorhabditis elegans
Authors: Sarparast Morteza , Hinman Jennifer , Pourmand Elham , Vonarx Derek , Ramirez Leslie , Ma Wenjuan , Liachko Nicole F. , Alan Jamie K. , Lee Kin Sing Stephen
Published: October 2023

This study aims to uncover potent cytochrome P450 (CYP) and epoxide hydrolase (EH) metabolites implicated in Aβ and/or tau-induced neurodegeneration, independent of neuroinflammation, by utilizingCaenorhabditis elegans(C. elegans) as a model organism. Our research reveals that Aβ and/or tau expression inC. elegansdisrupts the oxylipin profile, and epoxide hydrolase inhibition alleviates the ensuing neurodegeneration, likely through elevating the epoxy-to-hydroxy ratio of various CYP-EH metabolites. In addition, our results indicated that the Aβ and tau likely affect the CYP-EH metabolism of PUFA through different mechanism. These findings emphasize the intriguing relationship between lipid metabolites and neurodegenerations, in particular, those linked to Aβ and/or tau aggregation. Furthermore, our investigation sheds light on the crucial and captivating role of CYP PUFA metabolites inC. elegansphysiology, opening up possibilities for broader implications in mammalian and human contexts.

Abstract Figure
Host plant odour and sex pheromone are integral to mate finding in codling moth
Authors: Erdei Anna , Preti Michele , Bengtsson Marie , Witzgall Peter
Published: October 2023

Behaviour-modifying chemicals mediate sexual communication and host choice in insect herbivores. Sex pheromones are believed to attract insects by themselves, even though they are released into an atmosphere of plant odorants. We show for the first time that, in codling moth, feeding on apple and pear, that female pheromone is efficient for male attraction only in the presence of host plant odour. In non-host vegetation, male attraction to sex pheromone was very strongly reduced. The role of host odour in sex attraction was then substantiated by blending synthetic pheromone, codlemone, and the kairomone pear ester, a strong host plant attractant. An admixture of pear ester entirely rescued pheromone attraction in non-host vegetation. This field behavioural assay substantiates that host plant olfactory cues are integral part of sexual communication and mate recognition, which provides a mechanism for how shifts to new host plants produce new mate recognition signals.
Quantitative Profiling Method for Oxylipins in Neurodegenerative Diseases by Liquid Chromatography Coupled with Tandem Mass Spectrometry
Authors: Pourmand Elham , Zhang Fan , Sarparast Morteza , Alan Jamie K. , Lee Kin Sing Stephen
Published: October 2023

Aging is one of the major risk factors for many chronic diseases, including diabetes, neuropathy, hypertension, cancer, and neurodegenerative diseases. However, the mechanism behind aging and how aging affects a variety of disease progression remains unknown. Recent research demonstrated the cytochrome P450 (CYP)-epoxide hydrolase (EH) metabolites of polyunsaturated fatty acids (PUFAs) play a critical role in the abovementioned age-associated diseases. Therefore, aging could affect the abovementioned chronic diseases by modulating CYP-EH PUFA metabolism. Unfortunately, investigating how aging affects CYP-EH metabolism in human and mammalian models poses significant challenges.

In this regard, we will useC. elegansas a model organism to investigate the aging effects on CYP-EH metabolism of PUFA, owing to its long history of being used to study aging and its associated benefits of conducting aging research. This project will develop analytical tools to measure the endogenous levels of CYP-EH PUFA metabolites inC. elegansusing state-of-the-art ultra-performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS). These metabolites are very potent but present in low abundance. The dramatic increase in sensitivity in UPLC-MS/MS allows us to monitor these metabolites over the lifespan of C.eleganswith minimum samples. Our results show thatC. elegansproduces similar CYP PUFA metabolites to mammals and humans using our SPE-UPLC-MS/MS method. We will also show that our method successfully determined the CYP-EH PUFA metabolites profile changes induced by the inhibition ofC. elegansEH. The method developed from this project will significantly improve our understanding of the role of dietary PUFAs and associated metabolism on aging and neurodegeneration and will uncover new mechanisms of how aging affects neurodegeneration through the modulation of PUFA metabolic pathways.

Graphical abstract