bluuebunny/crossref_metadata_2025 huggingface dataset:
Enhanced meningeal lymphatic drainage ameliorates neuroinflammation and hepatic encephalopathy in cirrhotic rats
Authors: Hsu Shao-Jung , Zhang Chihao , Jeong Jain , Lee Seong-il , McConnell Matthew , Utsumi Teruo , Iwakiri Yasuko
Published: May 2020

Background and aims

Hepatic encephalopathy (HE) is a serious neurological complication in patients with liver cirrhosis. Nothing is known about the role of the meningeal lymphatic system in HE. We tested our hypothesis that enhancement of meningeal lymphatic drainage could decrease neuroinflammation and ameliorate HE.

Methods

A 4-week bile duct ligation (BDL) model was used to develop cirrhosis with HE in rats. Brain inflammation in patients with HE was evaluated using archived GSE41919. Motor function of rats was assessed by the rotarod test. AAV8-VEGF-C was injected into the cisterna magna of BDL rats one day after surgery to induce meningeal lymphangiogenesis.

Results

Cirrhotic rats with HE showed significantly increased microglia activation in the middle region of the cortex (p<0.001) as well as increased neuroinflammation as indicated by significant increases in IL-1β, INFγ, TNFα and Iba1 expression in at least one of the three regions of the cortex. Motor function was also impaired in rats with HE (p<0.05). Human brains of cirrhotic patients with HE also exhibited upregulation of pro-inflammatory genes (NF-κβ, Iba1, TNFα and IL-1β) (n=6). AAV8-VEGF-C injection significantly increased meningeal lymphangiogenesis (p=0.035) and tracer dye uptake in the anterior and middle regions of the cortex (p=0.006 & 0.003, respectively), their corresponding meninges (p=0.086 & 0.006, respectively) and the draining lymph nodes (p=0.02). Further, AAV8-VEGF-C decreased microglia activation (p<0.001) and neuroinflammation, and ameliorated motor dysfunction (p=0.024).

Conclusion

Promoting meningeal lymphatic drainage and enhancing waste clearance improves HE. Manipulation of meningeal lymphangiogenesis could be a new therapeutic strategy for the treatment of HE.
Gliding motility ofPlasmodiummerozoites
Authors: Yahata Kazuhide , Hart Melissa N. , Davies Heledd , Asada Masahito , Wassmer Samuel C. , Templeton Thomas J. , Treeck Moritz , Moon Robert W. , Kaneko Osamu
Published: May 2020

Plasmodiummalaria parasites are obligate intracellular protozoans that use a unique form of locomotion, termed gliding motility, to move through host tissues and invade cells. The process is substrate-dependent and powered by an actomyosin motor that drives the posterior translocation of extracellular adhesins which in turn propel the parasite forward. Gliding motility is essential for tissue translocation in the sporozoite and ookinete stages; however, the short-lived erythrocyte-invading merozoite stage has never been observed to undergo gliding movement. Here we showPlasmodiummerozoites possess the ability to undergo gliding motility and that this mechanism is likely an important precursor step for successful parasite invasion. We demonstrate that two human infective species,P. falciparumandP. knowlesi, have distinct merozoite motility profiles which may reflect distinct invasion strategies. Additionally, we develop and validate a higher throughput assay to evaluate the effects of genetic and pharmacological perturbations on both the molecular motor and complex signaling cascade that regulates motility in merozoites. The discovery of merozoite motility provides a new model to study the glideosome and may facilitate the pursuit of new targets for malaria treatment.

Significance statement

Plasmodiummalaria parasites use a unique substrate-dependent locomotion termed gliding motility to translocate through tissues and invade cells. Dogma has suggested that the small labile invasive stages that invade erythrocytes, merozoites, use this motility solely to penetrate target erythrocytes. Here we reveal that merozoites use gliding motility for translocation across host cells prior to invasion. This forms an important pre-invasion step that is powered by a conserved actomyosin motor and is regulated by a complex signaling pathway. This work fundamentally changes our understanding of the role of gliding motility and invasion in the blood and will have a significant impact on our understanding of blood stage host-pathogen interactions, parasite biology, and could have implications for vaccine development.
DNA methylation covariation in human whole blood and sperm: implications for studies of intergenerational epigenetic effects
Authors: Åsenius Fredrika , Gorrie-Stone Tyler J. , Brew Ama , Panchbaya Yasmin , Williamson Elizabeth , Schalkwyk Leonard C. , Rakyan Vardhman K. , Holland Michelle L. , Marzi Sarah J. , Williams David J.
Published: May 2020

Background

Epidemiological studies suggest that paternal obesity may increase the risk of fathering small for gestational age offspring. Studies in non-human mammals suggest that such associations could be mediated by DNA methylation changes in spermatozoa that influence offspring development in utero. Human obesity is associated with differential DNA methylation in peripheral blood. It is unclear, however, whether this differential DNA methylation is reflected in spermatozoa. We profiled genome-wide DNA methylation using the Illumina MethylationEPIC array in matched human blood and sperm from lean (discovery n=47; replication n=21) and obese (n=22) males to analyse tissue covariation of DNA methylation, and identify whether this covariation is influenced by obesity.

Results

DNA methylation signatures of human blood and spermatozoa are highly discordant, and methylation levels are correlated at only a minority of CpG sites (∼1%). While at the majority of these sites, DNA methylation appears to be influenced by genetic variation, obesity-associated DNA methylation in blood was not generally reflected in spermatozoa, and obesity did not influence covariation patterns. However, one cross-tissue obesity-specific hypermethylated site (cg19357369; chr4:2429884;P=8.95 × 10−8; beta=0.02) was identified, warranting replication and further investigation. When compared to a wide range of human somatic tissue samples (n=5,917), spermatozoa displayed differential DNA methylation in pathways enriched in transcriptional regulation.

Conclusions

Human sperm displays a unique DNA methylation profile that is highly discordant to, and practically uncorrelated with, that of matched peripheral blood. Obesity only nominally influences sperm DNA methylation, making it an unlikely mediator of intergenerational effects of metabolic traits.
A statistical framework for assessing pharmacological response and biomarkers using uncertainty estimates
Authors: Wang Dennis , Hensman James , Kutkaite Ginte , Toh Tzen S. ,   GDSC Screening Team, Dry Jonathan R , Saez-Rodriguez Julio , Garnett Mathew J. , Menden Michael P. , Dondelinger Frank
Published: May 2020

Drug high-throughput screenings across large molecular-characterised cancer cell line panels enable the discovery of biomarkers, and thereby, cancer precision medicine. The ability to experimentally generate drug response data has accelerated. However, this data is typically quantified by a summary statistic from a best-fit dose response curve, whilst neglecting the uncertainty of the curve fit and the potential variability in the raw readouts. Here, we model the experimental variance using Gaussian Processes, and subsequently, leverage this uncertainty for identifying associated biomarkers with a new statistical framework based on Bayesian testing. Applied to the Genomics of Drug Sensitivity in Cancer,in vitroscreening data on 265 compounds across 1,074 cell lines, our uncertainty models identified 24 clinically established drug response biomarkers, and in addition provided evidence for 6 novel biomarkers. We validated our uncertainty estimates with an additional drug screen of 26 drugs, 10 cell lines with 8 to 9 replicates. Our method is applicable to drug high-throughput screens without replicates, and enables robust biomarker discovery for new cancer therapies.
Functional profiling of COVID-19 respiratory tract microbiomes
Authors: Haiminen Niina , Utro Filippo , Seabolt Ed , Parida Laxmi
Published: May 2020

In response to the ongoing global pandemic, progress has been made in understanding the molecular-level host interactions of the new coronavirus SARS-CoV-2 responsible for COVID-19. However, when the virus enters the body it interacts not only with the host but also with the micro-organisms already inhabiting the host. Understanding the virus-host-microbiome interactions can yield additional insights into the biological processes perturbed by viral invasion. With this aim we carry out a functional analysis of previously published RNA sequencing data of bronchoalveolar lavage fluid from eight COVID-19 patients, twenty-five community-acquired pneumonia patients, and twenty healthy controls. The resulting microbiome functional profiles and their top differentiating features clearly separate the cohorts. By examining the functional features in connection with their associated metabolic pathways, differentially abundant pathways are indicated, compared to both the community-acquired pneumonia and healthy cohorts. From this analysis, distinguishing signatures in COVID-19 respiratory tract microbiomes are identified, including decreased lipid and glycan metabolism pathways, and increased carbohydrate metabolism pathways. Here we present a framework for comparative functional analysis of microbiomes, the results from which can lead to new hypotheses on the host-microbiome interactions in healthy versus afflicted cohorts. The findings from this analysis call for further research on microbial functions and host-microbiome interactions during SARS-CoV-2 infection.
Obesity-induced astrocyte dysfunction impairs heterosynaptic plasticity in the orbitofrontal cortex
Authors: Lau Benjamin K. , Murphy-Royal Ciaran , Kaur Manpreet , Qiao Min , Bains Jaideep S. , Gordon Grant R. , Borgland Stephanie L.
Published: May 2020

Summary

Overconsumption of highly palatable, energy dense food is considered a key driver of the obesity pandemic. The orbitofrontal cortex (OFC) is critical for reward valuation of gustatory signals, yet how the OFC adapts to obesogenic diets is poorly understood. Here we show that extended access to a cafeteria diet impairs astrocyte glutamate clearance, which leads to a heterosynaptic depression of GABA transmission onto pyramidal neurons of the OFC. This decrease in GABA tone is due to an increase in extrasynaptic glutamate, which acts via metabotropic glutamate receptors to liberate endocannabinoids. This impaired the induction of endocannabinoid-mediated long-term plasticity. In obese rats, this cascade of synaptic impairments was rescued by restoring astrocyte glutamate transport with the nutritional supplement, N-acetylcysteine. Together, our findings indicate that obesity targets astrocytes to disrupt the delicate balance between excitatory and inhibitory transmission in the OFC.

Highlights

Diet-induced obesity induces hypertrophy of astrocytes and impairs their ability to transport glutamate.

Failure of astrocytes to clear extrasynaptic glutamate drives endocannabinoid-mediated inhibitory long-term depression of principal output neurons in the OFC.

Astrocytic glutamate transporter function is restored with NAC, which rescues the synaptic deficits.
Long-TUC-seq is a robust method for quantification of metabolically labeled full-length isoforms
Authors: Rahmanian Sorena , Balderrama-Gutierrez Gabriela , Wyman Dana , McGill Cassandra Joan , Nguyen Kim , Spitale Robert , Mortazavi Ali
Published: May 2020

The steady state expression of each gene is the result of a dynamic transcription and degradation of that gene. While regular RNA-seq methods only measure steady state expression levels, RNA-seq of metabolically labeled RNA identifies transcripts that were transcribed during the window of metabolic labeling. Whereas short-read RNA sequencing can identify metabolically labeled RNA at the gene level, long-read sequencing provides much better resolution of isoform-level transcription. Here we combine thiouridine-to-cytosine conversion (TUC) with PacBio long-read sequencing to study the dynamics of mRNA transcription in the GM12878 cell line. We show that using long-TUC-seq, we can detect metabolically labeled mRNA of distinct isoforms more reliably than using short reads. Long-TUC-seq holds the promise of capturing isoform dynamics robustly and without the need for enrichment.
Culturing of ‘Unculturable’ Subsurface Microbes: Natural Organic Carbon Source Fuels the Growth of Diverse and Distinct Bacteria from Groundwater
Authors: Wu Xiaoqin , Spencer Sarah , Gushgari-Doyle Sara , Yee Mon Oo , Voriskova Jana , Li Yifan , Alm Eric J. , Chakraborty Romy
Published: May 2020

The recovery and cultivation of diverse field-related microorganisms from the terrestrial subsurface environment remains a challenge despite recent advances in modern molecular technology. Here we applied natural organic carbon (C), i.e., sediment-derived natural organic matter (NOM) and bacterial cell lysate, to groundwater microbial communities for a 30-day enrichment incubation, followed by conventional direct-plating for isolation. The groundwater was collected from a background well at the Oak Ridge Reservation Field Research Center, Tennessee. As a comparison, we also included enrichments amended with simple organic C sources, including glucose, acetate, benzoate, oleic acid, cellulose, and mixed vitamins. Our results demonstrate that complex natural organic C sources are more effective in enriching diverse bacterial species from groundwater than simple organic C sources. Microcosms amended with simple organic C (glucose, acetate, benzoate, or oleic acid) show significantly lower biodiversity than unamended control and are dominated by only few phyla such asProteobacteriaandBacteroidetes. In contrast, microcosms amended with complex natural organic C (sediment NOM or bacterial cell lysate) display significantly higher biodiversity, and enrich distinct species from the phyla that are poorly represented in published culture collections (e.g.,Verrucomicrobia, Planctomycetes, andArmatimonadetes). Our subsequent isolation efforts from natural organic C-amended enrichments led to 222 purified bacterial isolates representing 5 phyla, 16 orders, and 54 distinct species including candidate novel, rarely cultivated, and undescribed organisms.

Importance

Innovative strategies for recovering bacterial strains representing the true diversity of microbial communities in the terrestrial subsurface would significantly advance the understanding of ecologically critical taxa residing in these ecosystems. In this study, we demonstrate that complex natural organic C that mimic the naturally available resources for microbes encourages the growth of diverse bacteria much more robustly than traditional simplistic organic C sources. Results from this study will substantially advance and improve the design of strategies to effectively cultivate and isolate diverse and novel subsurface microorganisms in the laboratory. Obtaining axenic cultures of the ‘once-unculturable’ microorganisms will greatly enhance our understanding of microbial physiology, function, and roles in different biogeochemical niches in terrestrial subsurface ecosystems.
The mitotic protein NuMA plays a spindle-independent role in nuclear formation and mechanics
Authors: Serra-Marques Andrea , Houtekamer Ronja , Hintzen Dorine , Canty John T. , Yildiz Ahmet , Dumont Sophie
Published: May 2020

Eukaryotic cells typically form a single, round nucleus after mitosis, and failures to do so can compromise genomic integrity. How mammalian cells form such a nucleus remains incompletely understood. NuMA is a spindle protein whose disruption results in nuclear fragmentation. What role NuMA plays in nuclear integrity, or whether its perceived role stems from its spindle function, is unclear. Here, we use live imaging to demonstrate that NuMA plays a spindle-independent role in forming a single, round nucleus. NuMA keeps the decondensing chromosome mass compact at mitotic exit, and promotes a mechanically robust nucleus. NuMA’s C-terminus binds DNA in vitro and chromosomes in interphase, while its coiled-coil acts as a regulatory and structural hub: it prevents NuMA from binding chromosomes at mitosis, regulates its nuclear mobility and is essential for nuclear formation. Thus, NuMA plays a long-range structural role in building and maintaining an intact nucleus, as it does for the spindle, playing a protective role over the cell cycle.
Mutational landscape and in silico structure models of SARS-CoV-2 Spike Receptor Binding Domain reveal key molecular determinants for virus-host interaction
Authors: Nelson-Sathi Shijulal , Umasankar PK , Sreekumar E , Radhakrishnan Nair R , Joseph Iype , Nori Sai Ravi Chandra , Philip Jamiema Sara , Prasad Roshny , Navyasree KV , Ramesh Shikha , Pillai Heera , Ghosh Sanu , Santosh Kumar TR , Radhakrishna Pillai M.
Published: May 2020

Protein-protein interactions between virus and host are crucial for infection. SARS-CoV-2, the causative agent of COVID-19 pandemic is an RNA virus prone to mutations. Formation of a stable binding interface between the Spike (S) protein Receptor Binding Domain (RBD) of SARS-CoV-2 and Angiotensin-Converting Enzyme 2 (ACE2) of host actuates viral entry. Yet, how this binding interface evolves as virus acquires mutations during pandemic remains elusive. Here, using a high fidelity bioinformatics pipeline, we analysed 31,403 SARS-CoV-2 genomes across the globe, and identified 444 non-synonymous mutations that cause 49 distinct amino acid substitutions in the RBD. Molecular phylogenetic analysis suggested independent emergence of these RBD mutants during pandemic. In silico structure modelling of interfaces induced by mutations on residues which directly engage ACE2 or lie in the near vicinity revealed molecular rearrangements and binding energies unique to each RBD mutant. Comparative structure analysis using binding interface from mouse that prevents SARS-CoV-2 entry uncovered minimal molecular determinants in RBD necessary for the formation of stable interface. We identified that interfacial interaction involving amino acid residues N487 and G496 on either ends of the binding scaffold are indispensable to anchor RBD and are well conserved in all SARS-like corona viruses. All other interactions appear to be required to locally remodel binding interface with varying affinities and thus may decide extent of viral transmission and disease outcome. Together, our findings propose the modalities and variations in RBD-ACE2 interface formation exploited by SARS-CoV-2 for endurance.

Importance

COVID-19, so far the worst hit pandemic to mankind, started in January 2020 and is still prevailing globally. Our study identified key molecular arrangements in RBD-ACE2 interface that help virus to tolerate mutations and prevail. In addition, RBD mutations identified in this study can serve as a molecular directory for experimental biologists to perform functional validation experiments. The minimal molecular requirements for the formation of RBD-ACE2 interface predicted using in silico structure models may help precisely design neutralizing antibodies, vaccines and therapeutics. Our study also proposes the significance of understanding evolution of protein interfaces during pandemic.
Oral pyruvate prevents glaucomatous neurodegeneration
Authors: Williams Pete A , Harder Jeffrey M , Guymer Chelsea , Wood John P M , Daskalaki Evangelia , Chidlow Glyn , Cardozo Brynn H , Foxworth Nicole E , Cochran Kelly E , Ouellette Tionna B , Wheelock Craig E , Casson Robert J , John Simon W M
Published: May 2020

Intraocular pressure-sensitive retinal ganglion cell degeneration is a hallmark of glaucoma, the leading cause of irreversible blindness. Converging evidence indicates that age-related bioenergetic insufficiency increases the vulnerability of retinal ganglion cells to intraocular pressure. To investigate further, we used metabolomics and RNA-sequencing to examine early glaucoma in DBA/2J mice. We demonstrate an intraocular pressure-dependent decline in retinal pyruvate levels coupled to dysregulated glucose metabolism prior to detectable optic nerve degeneration. Oral supplementation of pyruvate strongly protected from neurodegeneration in pre-clinical models of glaucoma. We detected mTOR activation at the mechanistic nexus of neurodegeneration and metabolism. Rapamycin-induced inhibition of mTOR robustly prevented glaucomatous neurodegeneration. Bioenergetic enhancement, in combination with intraocular pressure reduction, therefore provides a readily translatable strategy that warrants investigation in clinical trials.

Funding

Vetenskapsrådet 2018-02124 and StratNeuro StartUp grant (PAW). Pete Williams is supported by the Karolinska Institutet in the form of a Board of Research Faculty Funded Career Position and by St. Erik Eye Hospital philanthropic donations. EY011721 and the Barbra and Joseph Cohen Foundation and startup funds from Columbia University (SWMJ). Simon John is an Investigator of HHMI.
Effective Dynamics of Nucleosome Configurations at the YeastPHO5Promoter
Authors: Wolff Michael Roland , Schmid Andrea , Korber Philipp , Gerland Ulrich
Published: May 2020

Chromatin dynamics are mediated by remodeling enzymes and play crucial roles in gene regulation, as established in a paradigmatic model, the yeastPHO5promoter. However, effective nucleosome dynamics, i.e. trajectories of promoter nucleosome configurations, remain elusive. Here, we infer such dynamics from the integration of published single-molecule data that capture multi-nucleosome configurations for repressed to fully activePHO5promoter states with other existing histone turnover and new chromatin accessibility data. We devised and systematically investigated a new class of “regulated on-off-slide” models simulating global and local nucleosome (dis)assembly and sliding. Only seven of 68 145 models agreed well with all data. All seven models involve sliding and the known central role of the N-2 nucleosome, but regulate promoter state transitions by modulating just one assembly rather than disassembly process. This is consistent with but challenges common interpretations of previous observations at thePHO5promoter and suggests chromatin opening by binding competitions.
Illuminating chromatin compaction in live cells and fixed tissues using SiR-DNA fluorescence lifetime
Authors: Hockings Colin , Poudel Chetan , Feeney Kevin A. , Novo Clara L. , Hamouda Mehdi S. , Mela Ioanna , Fernandez-Antoran David , Vallejo-Ramirez Pedro P. , Rugg-Gunn Peter J. , Chalut Kevin , Kaminski Clemens F. , Kaminski-Schierle Gabriele
Published: May 2020

The global compaction state of chromatin in a nucleus is an important component of cell identity that has been difficult to measure. We have developed a quantitative method to measure the chromatin compaction state in both live and fixed cells, without the need for genetic modification, using the fluorescence lifetime of SiR-DNA dye. After optimising this method using live cancer cell lines treated to induce chromatin compaction or decompaction, we observed chromatin compaction in differentiating epithelial cells in fixed tissue sections, as well as local decompaction foci that may represent transcription factories. In addition, we shed new light on chromatin decompaction during embryonic stem cell transition out of their naïve pluripotent state. This method will be useful to studies of nuclear architecture, and may be easy, cheap, and accessible enough to serve as a general assay of ‘stem-ness’.
A simple description of biological transmembrane potential from simple biophysical considerations and without equivalent circuits
Authors: Herrera-Valdez Marco Arieli
Published: May 2020

Biological membranes mediate different physiological processes necessary for life, including but not limited to electrical signaling, volume regulation, and other different forms of communication within and between cells. Ion movement is one of the physical processes underlying many of such processes. In turn, the difference between the electrical potentials inside and outside the cell called transmembrane potential, or membrane potential for short, is one of the key biophysical variables affecting ion movement. Most of the equations available to describe the change in membrane potential are based on analogies with resistive-capacitative electrical circuits. These equivalent circuit models were originally proposed in seminal studies dating back to 1872, and possibly earlier, and assume resistance and capacitance as measures of the permeable and the impermeable properties of the membrane, respectively. These models have been successful in shedding light on our understanding of electrical activity in cells, especially at times when the basic structure, biochemistry and biophysics of biological membrane systems were not well known. However, the parts in the ohmic circuits from which equations are derived, are not quite like the biological elements present in the spaces around and within biological membranes. Using current, basic knowledge about the structure, biophysics, and biochemical properties of biological membrane systems, it is possible to derive a simple, equation describing local changes in the transmembrane potential that is not based on electrical circuit analogies. The classical model for the membrane potential based on an equivalent RC-circuit is recovered as a particular case from the general derivation presented here, and concepts like the membrane capacitance can be explained as particular cases of the new equation. Modeling examples are presented to illustrate the use of the derivation, and the effects of changes in the voltage dependence of charge aggregation around the membrane on the timing and shape of neuronal action potentials.
Reference genome for the highly transformableSetaria viridiscultivar ME034V
Authors: Thielen Peter M. , Pendleton Amanda L. , Player Robert A. , Bowden Kenneth V. , Lawton Thomas J. , Wisecaver Jennifer H.
Published: May 2020

Setaria viridis(green foxtail) is an important model system for improving cereal crops due to its diploid genome, ease of cultivation, and use of C4photosynthesis. TheS. viridiscultivar ME034V is exceptionally transformable, but the lack of a sequenced genome for this cultivar has limited its utility. We present a 397 Mb highly contiguousde novoassembly of ME034V using ultra-long nanopore sequencing technology (read N50=41kb). We estimate that this genome is largely complete based on our updated k-mer based genome size estimate of 401 Mb forS. viridis. Genome annotation identified 37,908 protein-coding genes and >300k repetitive elements comprising 46% of the genome. We compared the ME034V assembly with two other previously sequencedSetariagenomes as well as to a diversity panel of 235S. viridiscultivars. We found the genome assemblies to be largely syntenic, but numerous unique polymorphic structural variants were discovered. Several ME034V deletions may be associated with recent retrotransposition ofcopiaandgypsyLTR repeat families, as evidenced by their low genotype frequencies in the sampled population. Lastly, we performed a phylogenomic analysis to identify gene families that have expanded inSetaria, including those involved in specialized metabolism and plant defense response. The high continuity of the ME034V genome assembly validates the utility of ultra-long DNA sequencing to improve genetic resources for emerging model organisms. Structural variation present inSetariaillustrates the importance of obtaining the proper genome reference for genetic experiments. Thus, we anticipate that the ME034V genome will be of significant utility for theSetariaresearch community.
Structural and functional alterations associated with the LRRK2 G2019S mutation revealed in structured human neural networks
Authors: Valderhaug Vibeke Devold , Ramstad Ola Huse , van de Wijdeven Rosanne , Heiney Kristine , Nichele Stefano , Sandvig Axel , Sandvig Ioanna
Published: May 2020

Mutations in the LRRK2 gene have been widely linked to Parkinson’s disease. The G2019S variant has been shown to contribute uniquely to both familial and sporadic forms of the disease. LRRK2-related mutations have been extensively studied, yet the wide variety of cellular and network events directly or indirectly related to these mutations remain poorly understood. In this study, we structured multi-nodal human neural networks carrying the G2019S mutation using custom-designed microfluidic chips coupled to microelectrode-arrays. By applying live imaging approaches, immunocytochemistry and computational modelling, we have revealed alterations in both the structure and function of the resulting neural networks when compared to controls. We provide first evidence of increased neuritic density associated with the G2019S LRRK2 mutation, while previous studies have found either a strong decrease, or no change, compared to controls. Additionally, we corroborate previous findings regarding increased baseline network activity compared to control neural networks. Furthermore, we can reveal additional network alterations attributable to the specific mutation by selectively inducing transient overexcitation to confined parts of the structured multi-nodal networks. These alterations, which we were able to capture both at the micro- and mesoscale manifested as differences in relative network activity and correlation, as well as in mitochondria activation, neuritic remodelling, and synaptic alterations. Our study thus provides important new insights into early signs of neural network pathology significantly expanding upon the current knowledge relating to the G2019S Parkinson’s disease mutation.
Low protein intake compromises the recovery of lactation-induced bone loss in female mouse dams without affecting skeletal muscles
Authors: Kanakis Ioannis , Alameddine Moussira , Scalabrin Mattia , van ‘t Hof Rob J. , Liloglou Triantafillos , Ozanne Susan E. , Goljanek-Whysall Katarzyna , Vasilaki Aphrodite
Published: May 2020

Lactation-induced bone loss occurs due to high calcium requirements for fetal growth but skeletal recovery is normally achieved promptly post-weaning. Dietary protein is vital for fetus and mother but the effects of protein undernutrition on the maternal skeleton and skeletal muscles is largely unknown. We used mouse dams fed with normal (N, 20%) or low (L, 8%) protein diet during gestation and lactation and maintained on the same diets (NN, LL) or switched from low to normal (LN) during a 28d skeletal restoration period post lactation. Skeletal muscle morphology and neuromuscular junction integrity was not different between any of the groups. However, dams fed the low protein diet showed extensive bone loss by the end of lactation, followed by full skeletal recovery in NN dams, partial recovery in LN and poor bone recovery in LL dams. Primary osteoblasts from low protein diet fed mice showed decreasedin vitrobone formation and decreased osteogenic marker gene expression; promoter methylation analysis by pyrosequencing showed no differences inBmpr1a, Ptch1, Sirt1, OsxandIgf1rosteoregulators, while miR-26a, -34a and -125b expression was found altered in low protein fed mice. Therefore, normal protein diet is indispensable for maternal musculoskeletal health during the reproductive period.
Rab18 Binds PLIN2 and ACSL3 to Mediate Lipid Droplet Dynamics
Authors: Deng Yaqin , Zhou Chang , Hammad Mirza Ahmed , Bamigbade Adekunle T. , Xu Shimeng , Zhang Shuyan , Liu Pingsheng
Published: May 2020

Rab18 has been linked to lipid metabolism and metabolic diseases in different model systems, but the mechanism of Rab18-mediated lipid droplet (LD) dynamics in muscle cells remains elusive. Here, we report that Rab18 plays an essential role in oleic acid (OA)-induced LD growth and formation in mouse myoblast cell line C2C12. Rab18 was translocated from endoplasmic reticulum (ER) to LDs during the LD growth induced by OA in C2C12 cells, which was directly regulated by perilipin 2 (PLIN2), a LD resident protein. LD-associated Rab18 bound with the C terminus of PLIN2, and the LD localization of Rab18 was diminished after PLIN2 deletion. Moreover, loss of function of Rab18 led to less triacylglycerol (TAG) accumulation and fewer but larger LD formation. In contrast, expression of wild type Rab18 and a constitutively active Rab18 (Q67L) mutant resulted in elevated TAG content and LD number. Furthermore, LD-associated Rab18 interacted with acyl-CoA synthetase long-chain family member 3 (ACSL3) and in turn, promoted the LD localization of ACSL3, which may play an important role in the accumulation of TAG induced by OA. These data showed that Rab18 was recruited to LD after OA treatment, and formed a complex with PLIN2 and ACSL3, which contributes to TAG accumulating and LD growth.
Unsupervised Idealization of Nano-Electronic Sensors Recordings with Concept Drifts: A Compressive Feature Learning Approach for Non-Stationary Single-Molecule Data Analysis
Authors: Ouqamra Mohamed
Published: May 2020

Single-molecule nanocircuits based on field-effect transistors (smFETs) are emerging and promising nano-bioelectronic sensors for the functional detection of molecular dynamics involved in biochemical transformations, in particular for applications in cancer thanks to a potentially better understanding of some hidden and complex molecular interactions. In fact, functionalized carbon nanotubes have been recently exploited to probe molecular events occurring at a single molecule scale with ultra high sensitivity and specificity, such as nucleic acids hybridization, enzyme folding in catalysis reactions, or protein-nucleic acids interactions. Extracting the kinetics and thermodynamics from such single-molecule dynamics implies robust analytic tools that can handle the complexity of the sensed reaction system changing between transient and steady-state molecular conformations, but also some challenging signal specificities, such as the multi-source composition of the recorded signals, the mixed and high-level noises, and the sensor baseline drift, leading to non-stationary time series. We present a new smFET data analysis framework, based on a compressive feature learning scheme to optimize unsupervised idealization of smFET traces, by a precise and accurate molecular events detection and states characterization algorithm, tailored for non-stationary signals at high sampling rate and long acquisition periods, without any prior knowledge on the data generating process nor signal pre-filtering. Experimental results show the accuracy and robustness of our trace idealization algorithm to stochastic state-space models, and better performances than commonly used hidden Markov models.
An artificial intelligence system reveals liquiritin inhibits SARS-CoV-2 by mimicking type I interferon
Authors: Zhu Jie , Deng Yong-Qiang , Wang Xin , Li Xiao-Feng , Zhang Na-Na , Liu Zurui , Zhang Bowen , Qin Cheng-Feng , Xie Zhengwei
Published: May 2020

The pandemic COVID-19 has spread to all over the world and greatly threatens safety and health of people. COVID-19 is highly infectious and with high mortality rate. As no effective antiviral treatment is currently available, new drugs are urgently needed. We employed transcriptional analysis to uncover potential antiviral drugs from natural products or FDA approved drugs. We found liquiritin significantly inhibit replication of SARS-CoV-2 in Vero E6 cells with EC50= 2.39 μM. Mechanistically, we found liquiritin exerts anti-viral function by mimicking type I interferon. Upregulated genes induced by liquiritin are enriched in GO categories including type I interferon signaling pathway, negative regulation of viral genome replication and etc. In toxicity experiment, no death was observed when treated at dose of 300 mg/kg for a week in ICR mice. All the organ indexes but liver and serum biochemical indexes were normal after treatment. Liquiritin is abundant in licorice tablet (~0.2% by mass), a traditional Chinese medicine. Together, we recommend liquiritin as a competitive candidate for treating COVID-19. We also expect liquiritin to have a broad and potent antiviral function to other viral pathogens, like HBV, HIV and etc.
Mutually opposing activity of PIN7 splicing isoforms is required for auxin-mediated tropic responses inArabidopsis thaliana
Authors: Kashkan Ivan , Hrtyan Mónika , Retzer Katarzyna , Humpolíčková Jana , Jayasree Aswathy , Filepová Roberta , Vondráková Zuzana , Simon Sibu , Rombaut Debbie , Jacobs Thomas B. , Frilander Mikko J. , Hejátko Jan , Friml Jiří , Petrášek Jan , Růžička Kamil
Published: May 2020

Advanced transcriptome sequencing has uncovered that the majority of eukaryotic genes undergo alternative splicing (AS). Nonetheless, little effort has been dedicated to investigating the functional relevance of particular splicing events, even those in the key developmental and hormonal regulators. Here we reveal, in the plant modelArabidopsis thaliana, that thePIN7gene, which encodes a polarly localized transporter for the phytohormone auxin, produces two evolutionary-conserved transcripts. These isoforms PIN7a and PIN7b, differing in a 4 amino acid motif, are present at nearly equal levels in most cells. Although both variants do not differ in the subcellular localization and transport auxin with similar capacity, they closely associate and mutually influence their stability within the plasma membrane. Phenotypic complementation tests reveal that the functional contribution of PIN7bper seis minor but it markedly attenuates the prominent PIN7a activity, which is required for correct seedling apical hook formation and auxin-mediated tropic responses. These results establish alternative splicing of the PIN family as a conserved, functionally-relevant mechanism, unveiling an additional regulatory level of auxin-mediated plant development.
Identification of synaptic PPT1 substrates highlight roles of depalmitoylation in disulfide bond formation and synaptic function
Authors: Gorenberg Erica L. , Tieze Sofia Massaro , Yücel Betül , Zhao Helen R. , Chou Vicky , Wirak Gregory S. , Tomita Susumu , Lam TuKiet T. , Chandra Sreeganga S.
Published: May 2020

Loss-of-function mutations in the depalmitoylating enzyme palmitoyl protein thioesterase 1 (PPT1) cause Neuronal Ceroid Lipofuscinosis type 1 (CLN1), a devastating neurodegenerative disease. Here, we provide a resource identifying PPT1 substrates. We utilized Acyl Resin-Assisted Capture and mass spectrometry to identify proteins with increasedin vivopalmitoylation in PPT1 knockout mouse brains. We then validated putative substrates through direct depalmitoylation with recombinant PPT1. This stringent screen elucidated >100 novel PPT1 substrates at the synapse, including channels and transporters, G-protein-associated molecules, endo/exocytic components, synaptic adhesion molecules, and mitochondrial proteins. Cysteine depalmitoylation sites in transmembrane PPT1 substrates frequently participate in disulfide bonds in the mature protein. We confirmed that depalmitoylation regulates disulfide bond formation in a tertiary screen analyzing post-translational modifications. Collectively, the diverse PPT1 substrates highlight the role of PPT1 in mediating synapse functions, implicate molecular pathways in the etiology ofCLN1, and advance our basic understanding of the purpose of depalmitoylation.

Highlights

∼10% of palmitoylated proteins are palmitoyl protein thioesterase 1 (PPT1) substrates

Unbiased proteomic approaches identify 9 distinct classes of PPT1 substrates, including synaptic adhesion molecules and endocytic proteins

Protein degradation does not require depalmitoylation by PPT1

Depalmitoylation mediates disulfide bond formation in transmembrane PPT1 substrates
Revealing the high propensity of RNAs to non-specifically bind drug-like small molecules
Authors: Kelly Megan L. , Chu Chia-Chieh , Shi Honglue , Ganser Laura R. , Bogerd Hal P. , Huynh Kelly , Hou Yuze , Cullen Bryan R. , Al-Hashimi Hashim M.
Published: May 2020

Identifying small molecules that selectively bind a single RNA target while discriminating against all other cellular RNAs is an important challenge in RNA-targeted drug discovery. Much effort has been directed toward identifying drug-like small molecules that minimize electrostatic and stacking interactions that lead to non-specific binding of aminoglycosides and intercalators to a variety of RNAs. Many such compounds have been reported to bind RNAs and inhibit their cellular activities, however the ability of such compounds to discriminate against RNA stem-loops commonly found in the transcriptome has not been thoroughly assessed in all cases. Here, we examined the propensities of three drug-like compounds, previously shown to bind and inhibit the cellular activity of three distinct RNAs, to non-specifically bind two HIV-1 stem-loop RNAs: the transactivation response element (TAR) and stem IIB in the rev response element (RREIIB). All three compounds bound to TAR and RREIIBin vitro, and two inhibited TAR-dependent transactivation and RRE-dependent viral export in cell-based assays while also exhibiting substantial off-target interactions consistent with non-specific cellular activity. A survey of X-ray and NMR structures of RNA-small molecule complexes revealed that drug-like molecules form hydrogen bonds with functional groups commonly accessible in canonical stem-loop RNA motifs, much like aminoglycosides, and in contrast to ligands that specifically bind riboswitches. Our results support extending the group of non-selective RNA-binders beyond aminoglycosides and intercalators to encompass drug-like compounds with capacity for non-specific hydrogen-bonding and reinforce the importance of assaying for off-target interactions and RNA selectivityin vitroand in cells when assessing novel RNA-binders.
Spatio-temporal analyses ofOCT4expression and fate transitions in human embryonic stem cells
Authors: Orozco-Fuentes Sirio , Wadkin Laura E. , Neganova Irina , Lako Majlinda , Barrio Rafael A. , Baggaley Andrew W. , Shukurov Anvar , Parker Nicholas G.
Published: May 2020

OCT4is one of the transcription factors required to maintain an undifferentiated state in human embryonic stem cells (hESCs). Thus, it is crucial to understand howOCT4transcription is regulated both at the single-cell and colony level. Here we analyse the changes ofOCT4-mCherry intensity expression in hESCs in the presence and absence of theBMP4morphogenetic protein.

We show thatOCT4expression is dynamic, reaching a maximum response 10 h afterBMP4treatment. We obtain the stationary probability distributions that govern the hESCs transitions amongst the different cell states in the presence/absence ofBMP4and establish the times at which the hESCs, that lead to differentiated and pluripotent cells, cluster in the colony. Furthermore, by quantifying the similarities between theOCT4expression amongst neighbouring hESCs, we show that hESCs express, on average, similar values in their local neighbourhood within the first two days of the experiment and beforeBMP4treatment. These results are relevant for the development of mathematical and computational models of adherent hESC colonies.
Tcf21+mesenchymal cells contribute to testis somatic cell development, homeostasis, and regeneration
Authors: Shen Yu-chi , Larose Hailey , Shami Adrienne Niederriter , Moritz Lindsay , Manske Gabriel L. , Ma Qianyi , Zheng Xianing , Sukhwani Meena , Czerwinski Michael , Sultan Caleb , Clements Jourdan , Chen Haolin , Spence Jason R. , Orwig Kyle E. , Tallquist Michelle , Li Jun Z. , Hammoud Saher Sue
Published: May 2020

Testicular development and function relies on interactions between somatic cells and the germline, but similar to other organs, regenerative capacity decline in aging and disease. Whether the adult testis maintains a reserve progenitor population with repair or regenerative capacity remains uncertain. Here, we characterized a recently identified mouse testis interstitial population expressing the transcription factor Tcf21. We found that Tcf21+cells are bipotential somatic progenitors present in fetal testis and ovary, maintain adult testis homeostasis during aging, and act as reserve somatic progenitors following injury.In vitro, Tcf21+cells are multipotent mesenchymal progenitors which form multiple somatic lineages including Leydig and myoid cells. Additionally, Tcf21+cells resemble resident fibroblast populations reported in other organs having roles in tissue homeostasis, fibrosis, and regeneration. Our findings reveal that the testis, like other organs, maintains multipotent mesenchymal progenitors that can be leveraged in development of future therapies for hypoandrogenism and/or infertility.

Highlights

Multipotent Tcf21+MPs can differentiate into somatic testis cell types

Tcf21+cells contribute to testis and ovary somatic cells during gonadal development

Tcf21+cells replenish somatic cells of the aging testis and in response to tissue injury

Testis Tcf21 cells resemble resident fibroblast populations in multiple organs
In silicosimulations of erythrocyte aquaporins with quantitativein vitrovalidation
Authors: Chan Ruth , Falato Michael , Liang Huiyun , Chen Liao Y.
Published: May 2020

Modelling water and membrane lipids is an essential element in the computational research of biophysical/biochemical processes such as water transport across the cell membrane. In this study, we examined the accuracies of two popular water models, TIP3P and TIP4P, in the molecular dynamics simulations of erythrocyte aquaporins (AQP1 and AQP3). We modelled the erythrocyte membrane as an asymmetric lipid bilayer with appropriate lipid compositions of its inner and outer leaflet, in comparison with a symmetric lipid bilayer of a single lipid type. We computed the AQP1/3 permeabilities with the transition state theory with full correction for recrossing events. We also conducted cell swelling assays for water transport across the erythrocyte membrane. The experimental results agree with the TIP3P water-erythrocyte membrane model, in confirmation of the expected accuracy of the erythrocyte membrane model, the TIP3P water model, and the CHARMM parameters for water-protein interactions.