bluuebunny/crossref_metadata_2025 huggingface dataset:
Apicosome: newly identified cell-type-specific organelle in mouse cochlear and vestibular hair cells
Authors: Li Xiaofen , Zhao Qirui , Yu Xiaojie , Cao Wenhan , Zhang Yingyi , Feng Wanying , Jiang Liwen , He David Z. , Qi Robert Z. , Huang Pingbo
Published: July 2022

Cochlear and vestibular hair cells in the inner ear are highly specialized sensory receptors for sound waves and acceleration of body movements; these cells can perform their specialized functions because of their distinctive morphology and some unique organelles that they harbor. Here, we report a serendipitous identification in the mouse of a hair-cell-specific organelle, which we name “apicosome.” The apicosome was recognized by anti-FLRT1 antibodies but contains no FLRT1, and the organelle presents several distinctive characteristics: (1) the apicosome typically appears as a single entity (∼500 nm in diameter), but occasionally as two entities, in hair cells; (2) it first appears in the subapical region at the neural side at embryonic day (E) 17–18 in cochlear hair cells, subsequently descends to the perinuclear region during the first postnatal week, and completely disappears around postnatal day (P) 10; (3) in vestibular hair cells, it can be detected in the subapical region of neonatal (P3) cells and persists in adult hair cells although it becomes smaller and more distant from the subapical region; (4) the timing of apicosome translocation and disappearance during development is correlated in kinocilium maintenance; (5) the organelle is potentially associated with microtubules; and (6) the appearance of the apicosome is irregular in supernumerary hair cells and this is likely linked to anomalous lateral inhibition. Thus, our study identifies a previously undescribed organelle in sensory hair cells and lays the foundation for further characterization of this specialized structure potentially linked to hair-cell development and morphogenesis.
Metabolic Reprogramming in Rheumatoid Arthritis Synovial Fibroblasts: a Hybrid Modeling Approach
Authors: Aghakhani Sahar , Soliman Sylvain , Niarakis Anna
Published: July 2022

Rheumatoid Arthritis (RA) is an autoimmune disease characterized by a highly invasive pannus formation consisting mainly of synovial fibroblasts (RASFs). This pannus leads to cartilage, bone and soft tissue destruction in the affected joint. RASFs’ activation is associated with metabolic alterations resulting from dysregulation of extracellular signals transduction and gene regulation machinery. Deciphering the intricate mechanisms at the origin of this metabolic reprogramming may provide significant insight into RASFs’ involvement in RA’s pathogenesis and offer new therapeutic strategies. Qualitative and quantitative dynamic modeling can address some of these features, but hybrid models represent a real asset in their ability to span multiple layers of biological machinery. This work presents the first hybrid RASF model: the combination of a cell-specific qualitative regulatory network with a global metabolic network. The automated framework for hybrid modeling exploits the regulatory network’s trap-spaces as additional constraints on the metabolic networks. Subsequent flux balance analysis allows assessment of RASFs’ regulatory outcomes’ impact on their metabolic flux distribution. The hybrid RASF model simulates the experimentally observed metabolic reprogramming induced by signaling and gene regulation in RASFs. Simulations also enable further hypotheses on the potential reverse Warburg effect in RA. RASFs may undergo metabolic reprogramming to turn into “metabolic factories”, producing high levels of energy-rich fuels and nutrients for neighboring demanding cells through the crucial role of HIF1.

Author Summary

We successfully built the first large-scale hybrid dynamical model for human Rheumatoid Arthritis Synovial Fibroblasts (RASFs) including signaling, gene regulation and metabolism. We used a state-of-the-art molecular map for upstream signaling and gene regulation, the tool CaSQ to infer a large-scale Boolean model, and a genome-scale metabolic model. Trap-spaces of the Boolean asynchronous model were used to infer additional metabolic constraints on the metabolic network for subsequent flux balance analysis. This method allowed us to study the impact of various regulatory initial conditions on RASFs’ metabolic fluxes distribution. Our model successfully reproduces the metabolic reprogramming of RASFs which shift their ATP production from oxidative pathways to glycolysis, highlighting the key role of HIF1 in this process. Our findings allow us to hypothesize a reverse Warburg relationship occurring between RASFs and other RA joint cells. Similarly to tumor microenvironment’s fibroblasts, RASFs would undergo a metabolic switch and reprogram their metabolism to adapt to their hypoxic environment and provide crucial metabolic intermediates to neighboring cells to sustain their inflammatory activity.
Identification of Antigenic Regions Responsible for inducing Type 1 diabetes mellitus
Authors: Kumar Nishant , Patiyal Sumeet , Choudhury Shubham , Tomer Ritu , Dhall Anjali , Raghava Gajendra P. S.
Published: July 2022

There are a number of antigens that induce autoimmune response against β-cells, leading to Type 1 diabetes mellitus (T1DM). Recently several antigen-specific immunotherapies have been developed to treat T1DM. Thus identification of T1DM associated peptides with antigenic regions or epitopes is important for peptide based-therapeutics (e.g., immunotherapeutic). In this study, for the first time an attempt has been made to develop a method for predicting, designing and scanning of T1DM associated peptides with high precision. We analyzed 815 T1DM associated peptides and observed that these peptides are not associated with a specific class of HLA alleles. Thus, HLA binder prediction methods are not suitable for predicting T1DM associated peptides. Firstly, we developed a similarity/alignment based method using BLAST and achieved a high probability of correct hits with poor coverage. Secondly, we developed an alignment free method using machine learning techniques and got maximum AUROC 0.89 using dipeptide composition. Finally, we developed a hybrid method that combines the strength of both alignment free and alignment based methods and achieve maximum AUROC 0.95 with MCC 0.81 on independent dataset. We developed a webserver “DMPPred” and standalone server, for predicting, designing and scanning of T1DM associated peptides (https://webs.iiitd.edu.in/raghava/dmppred/).

Key Points

Prediction of peptides responsible for inducing immune system against β-cells

Compilation and analysis of Type 1 diabetes associated HLA binders

BLAST based similarity search against Type 1diabetes associated peptides

Alignment free method using machine learning techniques and composition

A hybrid method using alignment free and alignment based approach

Author’s Biography

Nishant Kumar is currently working as Ph.D. in Computational biology from Department of Computational Biology, Indraprastha Institute of Information Technology, New Delhi, India

Sumeet Patiyal is currently working as Ph.D. in Computational biology from Department of Computational Biology, Indraprastha Institute of Information Technology, New Delhi, India

Shubham Choudhury is currently working as Ph.D. in Computational biology from Department of Computational Biology, Indraprastha Institute of Information Technology, New Delhi, India

Ritu Tomer is currently working as Ph.D. in Computational biology from Department of Computational Biology, Indraprastha Institute of Information Technology, New Delhi, India

Anjali Dhall is currently working as Ph.D. in Computational Biology from Department of Computational Biology, Indraprastha Institute of Information Technology, New Delhi, India.

Gajendra P. S. Raghava is currently working as Professor and Head of Department of Computational Biology, Indraprastha Institute of Information Technology, New Delhi, India.
Cannabinoids Activate the Insulin Pathway to Modulate Mobilization of Cholesterol in C. elegans
Authors: Hernandez-Cravero Bruno , Gallino Sofia , Florman Jeremy , Vranych Cecilia , Diaz Philippe , Belén Elgoyhen Ana , Alkema Mark J. , de Mendoza Diego
Published: July 2022

The nematode Caenorhabditis elegans requires exogenous cholesterol to survive and its depletion leads to early development arrest. Thus, tight regulation of cholesterol storage and distribution within the organism is critical. Previously, we demonstrated that the endocannabinoid (eCB) 2-arachidonoylglycerol (2-AG) plays a key role in C. elegans modulating sterol mobilization, but the mechanism is unknown. Here we show that mutations in the ocr-2 and osm-9 genes coding for transient receptors potential V (TRPV) ion channels, dramatically reduces the effect of 2-AG in cholesterol mobilization. Through genetic analysis combined with the rescuing of larval arrest induced by sterol starvation we found that the insulin/IGF-1signaling (IIS) pathway and UNC-31/CAPS, a calcium-activated regulator of neural dense-core vesicles release, are essential for 2-AG-mediated stimulation of cholesterol mobilization. These findings indicate that 2-AG-dependent cholesterol trafficking requires the release of insulin peptides and signaling through the DAF-2 insulin receptor. These results suggest that 2-AG acts as an endogenous modulator of TRPV signal transduction to control intracellular sterol traffic through modulation of the IGF-1 signaling pathway.

Author summary

Although cannabis extracts have been used in folklore medicine for centuries, the past few years have seen an increased interest in the medicinal uses of cannabinoids, the bioactive components of the cannabis plant, for treatment of many diseases of the nervous system. However, the human body naturally produces endocannabinoids that are similar to the cannabinoids present in Cannabis sativa. Our goal is to understand how endocannabinoids maintain cholesterol homeostasis in animals, underscoring the importance of cholesterol balance for healthy life. Both cholesterol excess and cholesterol deficiency can have detrimental effects on health, and a myriad of regulatory processes have thus evolved to control the metabolic pathways of sterol metabolism. The nematode C. elegans is auxotroph for sterols, that is; contrary to mammals they cannot synthesize sterols, therefore, dietary supply is essential for survival. The aim of our study was to elucidate the mechanism by which endocannabinoids abolish larval arrest of C. elegans induced by cholesterol depletion. We discovered that endocannabinoids stimulate the insulin pathway, which affects development, reproduction and life span, to modulate mobilization of cholesterol in C. elegans. Our studies have important implications for a better understanding of human pathological conditions associated with impaired cholesterol homeostasis.
Synthesis and characterization of poly(propylene imine) dendrimers, as nanocarriers of Benznidazole: an in vitro controlled release assay
Authors: Ordoñez-Benavides Jenny , Andrade-Caicedo Henry
Published: July 2022

Background

American trypanosomiasis, or Chagas disease, is the result of an infection caused by the Trypanosoma cruzi parasite. The disease is endemic in Latin America, where the main clinical manifestation and cause of death of Chagas patients is cardiomyopathy. The current approved treatment for this disease is based on the use of the nitroheterocyclic compound, Benznidazole. The drug is administered in high doses and for prolonged periods, which causes serious adverse effects, eventually leading to treatment discontinuation. In addition, it has only shown efficacy in the acute phase of the disease. Benznidazole has low solubility, low permeability, low bioavailability and high toxicity in the body. These physicochemical characteristics can be improved by using dendritic structures that serve as nanocarriers.

Methods

In this research, poly(propylene imine) PPI dendrimers in generations 4.0 G and 5.0 G were synthesized and characterized. We performed the synthesis by divergent approach. We encapsulated Benznidazole using the equilibrium dialysis method, and we evaluated the loading efficiency and the concentration of the released drug by high-performance liquid chromatography (HPLC).

Results

Preliminary results showed a drug loading efficiency on the dendrimer of 78% and an entrapment percentage of 99.6%. The release kinetics showed a controlled and sustained release over time compared to dendrimer-free Benznidazole.

Conclusions

The PPI 5.0 G - Benznidazole dendrimer system could be considered as an alternative to be evaluated in vitro and in vivo, as an alternative to conventional treatment of Chagas disease. The next stage of the experimental work consists of standardizing an infection model of H9C2 cardiomyocytes with Colombian strains of Trypanosoma cruzi, in order to evaluate the effect of the encapsulated drug on nanocarriers.
Visuomotor phase-locked loop reproduces elliptic hand trajectories across different rhythms
Authors: Matić Adam
Published: July 2022

A well-known phenomenon in human hand movement is the correlation between speed and curvature, also known as the speed-curvature power law (V≈kCβ). In drawing elliptic shapes, the exponent is often found to be β ≈ −1/3, however it is not clear why the power law appears and why the exponent is near −1/3. More fundamentally, it is not clear how do people track elliptic targets. In answering these questions, I’ve analyzed trajectories of participants’ cursors while they tracked visual targets moving along elliptical paths, across different target speed profiles and cycle frequencies. The speed-curvature power law emerged when drawing ellipses at about 1 Hz or faster, regardless of the target speed profile, and it did not emerge for lower frequency movements. Analysis of the position frequency spectrum shows that the target-cursor trajectory transformation may be seen as a low-pass filter. Comparison of different hypothetical salient features of the visual field shows that phase difference (angular difference between the cursor and the target) and size difference (difference in the sizes of the elliptic paths) are the features most likely used in the task. The next experiment confirmed that phase and size difference could be controlled variables because participants kept them stable even under direct pseudorandom disturbances. A numerical model simulating the sensorimotor processes of the participant, similar to a phase-locked loop, using the visual features of phase and size difference as controlled variables, performed the same target tracking tasks as the participants. When fitted, the model closely replicated position and speed profiles of the participants across all trials, as well as the emergence of the power law at high frequencies. The model also reproduced the trajectories of participants in the experiment with direct pseudorandom disturbances. In conclusion (1) the speed-curvature power law emerges as a side effect of movement system properties, namely low-pass filtering in the sensorimotor loop; (2) people could be tracking elliptical targets by varying the frequency and amplitude of an internal pattern generator until the produced phase and shape size match the target’s phase and shape size. The model generates new hypotheses about the neural mechanisms of rhythmic movement control.
Behaviour of pyrethroid resistant Anopheles gambiae at the interface of two dual active-ingredient bed nets, assessed by room-scale infrared video tracking
Authors: Gleave K. , Guy A. , Mechan F. , Emery M. , Murphy A. , Voloshin V. , Towers C. E. , Towers D. , Ranson H. , Foster G. M. , McCall P. J.
Published: July 2022

Background

The success of Insecticide Treated Bednets (ITNs) for malaria vector control in Africa relies on the behaviour of the major malaria vectors, Anopheles species. Research into mosquito behavioural traits influencing the performance of ITNs has focused largely on time or location of biting. Here we investigated less tractable behaviours including timings of net contact, willingness to refeed and longevity post exposure to two next-generation nets, PermaNet® 3.0 (P3) and Interceptor® G2 (IG2) in comparison with a standard pyrethroid only net (Olyset (OL)) and an untreated net.

Methods

Susceptible and resistant Anopheles gambiae mosquitoes were exposed to the nets with a human volunteer host in a room scale assay. Mosquito movements were tracked for two hours using an infrared video system, collecting flight trajectory, spatial position and net contact data. Post-assay, mosquitoes were monitored for a range of sublethal insecticide effects.

Results

OL, P3 and IG2 all killed over 90% of susceptible mosquitoes 24 hours after exposure, but this effect was not seen with resistant mosquitoes where mortality ranged from 16% to 72%. Total mosquito activity was higher around untreated nets than ITNs. There was no difference in total activity, the number, or duration, of net contact, between any mosquito strain, with similar behaviours recorded in susceptible and resistant strains at all ITNs. Net contact was focussed predominantly on the roof for all bednets. We observed a steep decay in activity for both susceptible strains when P3 and OL were present and with IG2 for one of the two susceptible strains. All treated nets reduced the willingness of resistant strains to re-feed when offered blood one-hour post-exposure, with a more pronounced effect seen with P3 and OL than IG2.

Conclusion

Results indicate that the effects of ITNs on mosquito behaviour are consistent, with no major differences in responses between strains of different pyrethroid susceptibility.
Structural characterization of functionally important chloride binding sites in the marine Vibrio alkaline phosphatase
Authors: Markússon Sigurbjörn , Hjörleifsson Jens G. , Kursula Petri , Ásgeirsson Bjarni
Published: July 2022

Enzyme stability and function can be affected by various environmental factors, such as temperature, pH and ionic strength. Enzymes that are located outside the relatively unchanging environment of the cytosol, such as those residing in the periplasmic space of bacteria or extracellularly secreted, are challenged by more fluctuations in the aqueous medium. Bacterial alkaline phosphatases (APs) are generally affected by ionic strength of the medium, but this varies substantially between species. An AP from the marine bacterium Vibrio splendidus (VAP) shows complex pH-dependent activation and stabilization in the 0 – 1.0 M range of halogen salts and has been hypothesized to specifically bind chloride anions. Here, using X-ray crystallography and anomalous scattering, we have located two chloride binding sites in the structure of VAP, one in the active site and another one at a peripheral site. Further characterization of the binding sites using site-directed mutagenesis and small angle X-ray scattering (SAXS) showed that upon binding of chloride to the peripheral site, structural dynamics decreased locally, resulting in thermal stabilization of the VAP active conformation. Binding of the chloride ion in the active site did not displace the bound inorganic phosphate product, but it may promote product release by facilitating rotational stabilization of the substrate-binding Arg129. Overall, these results reveal the complex nature and dynamics of chloride binding to enzymes through long-range modulation of electronic potential in the vicinity of the active site, resulting in increased catalytic efficiency and stability.
Infrared Spectroscopy for Structure Analysis of Protein Inclusion Bodies
Authors: Schwaighofer Andreas , Lendl Bernhard
Published: July 2022

Infrared (IR) spectroscopy is a widely used technique for evaluation of protein secondary structure. In this chapter, we focus on the application of this analytical technique for analysis of inclusion bodies. After a general introduction to protein analysis by IR spectroscopy different approaches for spectra acquisition, data processing and secondary structure evaluation are presented.
Structural basis for differential targeting properties of small RNAs in plants and animals
Authors: Xiao Yao , MacRae Ian J.
Published: July 2022

microRNAs (miRNAs) regulate gene expression in plants and animals. Animals use miRNAs to sculpt the transcriptome, with each miRNA modestly repressing hundreds of otherwise unrelated targets. By contrast, each plant miRNA potently silences a small number of physiologically related targets. Here, we show that this major functional distinction depends on a minor structural difference between plant and animal Argonaute (AGO) proteins. A 7-amino-acid loop in the PIWI domain of Arabidopsis Argonaute10 (AtAGO10) reduces the affinity of the miRNA seed region for target RNAs. Swapping the PIWI-loop from human Argonaute2 (HsAGO2) into AtAGO10 increases seed strength and target-binding promiscuity, resembling animal miRNA targeting. Conversely, swapping the plant PIWI-loop into HsAGO2 increases targeting stringency and elevates target cleavage rates. The loop-swapped HsAGO2-siRNA complex silences targets more potently, with reduced off-targeting, than wild-type HsAGO2 and small interfering RNA (siRNA) duplexes in mammalian cells. Thus, tiny structural differences can tune the targeting properties of AGO proteins for distinct biological roles and HsAGO2 can be engineered for potential applications in siRNA therapeutics.
Light-up split Broccoli aptamer as a versatile tool for RNA assembly monitoring in cell-free TX-TL system, hybrid RNA/DNA origami tagging and DNA biosensing
Authors: Torelli Emanuela , Shirt-Ediss Benjamin , Navarro Silvia A. , Manzano Marisa , Vizzini Priya , Krasnogor Natalio
Published: July 2022

Binary light-up aptamers are intriguing and emerging tools with potential in different fields. Herein, we demonstrate the versatility of a split Broccoli aptamer system able to turn on the fluorescence signal only in the presence of a complementary sequence.

First, an RNA three-way junction harbouring the split system was assembled in an E. coli based cell-free TX-TL system where the folding of the functional aptamer is demonstrated. Then, the same strategy is introduced into a ‘bio-orthogonal’ hybrid RNA/DNA rectangle origami characterized by atomic force microscopy: the activation of the split system through the origami self-assembly is demonstrated. Finally, our system is successfully used to detect femtomoles of a Campylobacter spp. DNA target sequence.

Potential applications of our system include real-time monitoring of the self-assembly of nucleic acid-based devices in vivo and of intracellular delivery of therapeutic nanostructures, as well as in vitro and in vivo detection of different DNA/RNA target.
Negative Electrospray Supercharging Mechanisms of Oligonucleotide Single Strands and G-quadruplexes
Authors: Ghosh Debasmita , Rosu Frédéric , Gabelica Valérie
Published: July 2022

When sprayed from physiological ionic strength, nucleic acids typically end up with low levels of charging and in compact conformations. Increasing the electrospray negative charging of nucleic acids while preserving the native non-covalent interactions can help distinguish solution folds by ion mobility mass spectrometry. To get fundamental insight into the supercharging mechanisms of nucleic acids in the negative mode, we studied model G-quadruplex structures and single strand controls in 100 mM ammonium acetate. We found that adding 0.4% propylene carbonate, 0.4% sulfolane or 0.1% m-NBA induces native supercharging through the charged residue mechanism. However, although 0.4% m-NBA shows the highest supercharging ability, it induces unwanted unfolding of solution-folded G-quadruplexes. The supercharging effect resembles the effect of lowering the ionic strength, which could be explained by partial neutralization of the ampholytes when droplets become more concentrated in their non-aqueous components. The supercharging ability ranks: PC < sulfolane < m-NBA. m-NBA adducts to G-quadruplexes with high charge states confirms that the supercharging agent interacts directly with DNA. Surprisingly, in presence of supercharging agents, more negative charge states also bear more alkali metal ion adducts. This suggests that native supercharging results from larger droplets evaporating to the charged residue, leading to higher concentration of both the supercharging agent and of alkali counterions. However, when negative charge carriers from the electrolyte become too rare, chain ejection accompanied by denaturation, and hence non-native supercharging, can become predominant.
Phylogenomics oftrans-Andean tetras of the genusHyphessobryconDurbin 1908 and colonization patterns of Middle America
Authors: Elías Diego J. , McMahan Caleb D. , Alda Fernando , García-Alzate Carlos , Hart Pamela B. , Chakrabarty Prosanta
Published: July 2022

Hyphessobryconis one of the most species rich and widely distributed genera in the family Characidae, with more than 160 species ranging from Veracruz, Mexico to Argentina. The majority of the diversity ofHyphessobryconshows acis-Andean distribution; only nine species aretrans-Andean includingH. compressus(Meek 1908). It is well established thatHyphessobryconis not monophyletic but has been suggested that natural groups can be identified within the largerHyphessobryconspecies group. In this study, we test the monophyly oftrans-Andean species ofHyphessobryconand investigate the placement ofH. compressus. We have inferred the first phylogenomic hypothesis oftrans-AndeanHyphessobryconthat includes nearly complete taxonomic sampling (eight of nine valid species) using ultraconserved elements (UCEs). We analyzed 75% (1682 UCEs), 90% (1258 UCEs), and 95% (838 UCEs) complete data matrices, and inferred phylogenomic hypotheses under concatenation and coalescent approaches. In all cases, we recovered the monophyly oftrans-AndeanHyphessobryconinclusive ofH. compressus, and strong support for three species groups and evidence of cryptic diversity within the widespreadH. compressusandH. condotensis. We used our phylogenomic hypothesis to investigate the biogeographic history ofHyphessobryconin Middle America. Our ancestral range estimation analysis suggests a single event ofcis- totrans-Andean colonization followed by stepwise colonization from the Pacific slope of northwestern South America (Chocó block) to northern Middle America (Maya block). Our work supports the recognition of thetrans-Andean species asHyphessobryconsensu stricto and provides a robust evolutionary template to examine morphological characters that will allow us to better understand the diversity ofHyphessobryconin Middle America.
DNA methyltransferase enhanced Fusobacterium nucleatum genetics
Authors: Umaña Ariana , Nguyen Tam T.D. , Sanders Blake E. , Williams Kevin J. , Wozniak Bryce , Slade Daniel J.
Published: July 2022

Bacterial restriction-modification (R-M) systems are a first line immune defense against foreign DNA from viruses and other bacteria. While R-M systems are critical in maintaining genome integrity, R-M nucleases unfortunately present significant barriers to targeted genetic modification. Bacteria of the genus Fusobacterium are oral, Gram-negative, anaerobic, opportunistic pathogens that are implicated in the progression and severity of multiple cancers and tissue infections, yet our understanding of their direct roles in disease have been severely hindered by their genetic recalcitrance. Here, we demonstrate a path to overcome these barriers in Fusobacterium by using native DNA methylation as a host mimicry strategy to bypass R-M system cleavage of user introduced plasmid DNA. We report the identification, characterization, and successful use of Fusobacterium nucleatum (Fn) Type II and III DNA methyltransferase (DMTase) enzymes to produce a multi-fold increase in gene knockout efficiency in the strain Fusobacterium nucleatum subsp. nucleatum 23726 (Fnn 23726), as well as the first efficient gene knockouts and complementations in Fnn 25586. We show plasmid protection can be accomplished in vitro with purified enzymes, as well as in vivo in an E. coli host that constitutively expresses Fnn DMTase enzymes. By characterizing specific DMTases that are critical for bypassing R-M systems, we have enhanced our understanding of potential enzyme combinations, with the goal of expanding these studies to genetically modify clinical isolates of Fusobacterium that have thus far been inaccessible to molecular characterization. This proof-of-concept study provides a roadmap to guide molecular microbiology efforts of the scientific community to facilitate the discovery of new Fusobacterium virulence genes, thereby leading to a new era of characterizing how an oral opportunistic pathogen contributes to an array of human infections and diseases.

IMPORTANCE

Fusobacterium nucleatum is an oral opportunistic pathogen associated with diseases including cancer and preterm birth. Our understanding of how this bacterium modulates human disease has been hindered by a lack of genetic systems. Here we show that F. nucleatum DNA methyltransferase modified plasmid DNA overcomes the transformation barrier and allows the development of genetic systems in previously inaccessible strains. We present a strategy that can be expanded to enable the genetic modification of clinical isolates, thereby fostering investigational studies to uncover novel host-pathogen interactions in Fusobacterium.
Gabapentin Disrupts Binding of Perlecan to the α2δ1Voltage Sensitive Calcium Channel Subunit and Impairs Skeletal Mechanosensation
Authors: Reyes Fernandez Perla C. , Wright Christian S. , Masterson Adrianna N. , Yi Xin , Tellman Tristen V. , Bonteanu Andrei , Rust Katie , Noonan Megan L. , White Kenneth E. , Lewis Karl J. , Sankar Uma , Hum Julia M. , Bix Gregory , Wu Danielle , Robling Alexander G. , Sardar Rajesh , Farach-Carson Mary C. , Thompson William R.
Published: July 2022

Our understanding of how osteocytes, the principal mechanosensors within bone, sense and perceive force remains unclear. Previous work identified “tethering elements” (TEs) spanning the pericellular space of osteocytes and transmitting mechanical information into biochemical signals. While we identified the heparan sulfate proteoglycan perlecan (PLN) as a component of these TEs, PLN must attach to the cell surface to induce biochemical responses. As voltage-sensitive calcium channels (VSCCs) are critical for bone mechanotransduction, we hypothesized that PLN binds the extracellular α2δ1subunit of VSCCs to couple the bone matrix to the osteocyte membrane. Here, we showed co-localization of PLN and α2δ1along osteocyte dendritic processes. Additionally, we quantified the molecular interactions between α2δ1and PLN domains and demonstrated for the first time that α2δ1strongly associates with PLN via its domain III. Furthermore, α2δ1is the binding site for the commonly used pain drug, gabapentin (GBP), which is associated with adverse skeletal effects when used chronically. We found that GBP disrupts PLN::α2δ1bindingin vitro, and GBP treatmentin vivoresults in impaired bone mechanosensation. Our work identified a novel mechanosensory complex within osteocytes composed of PLN and α2δ1, necessary for bone force transmission and sensitive to the drug GBP. This work provides insights into the mechanisms underlying mechanotransduction and will inform future studies to understand the mechanisms responsible for the negative effects of GBP on bone.
Actively replicating gut bacteria identified by 5-ethynyl-2’-deoxyuridine (EdU) click chemistry and cell sorting
Authors: Beauchemin Eve T. , Hunter Claire , Maurice Corinne F.
Published: July 2022

The composition of the intestinal bacterial community is well described, but recent research suggests that the metabolism of these bacteria plays a larger role in health than which species are present. One fundamental aspect of gut bacterial metabolism that remains understudied is bacterial replication. Indeed, there exist few techniques which can identify actively replicating gut bacteria. In this study, we aimed to address this gap by adapting 5-ethynyl-2’-deoxyuridine (EdU) click chemistry (EdU-click), a metabolic labeling method, coupled with fluorescence-activated cell sorting and sequencing (FACS-Seq) to characterize replicating gut bacteria. We first used EdU-click with human gut bacterial isolates and show that many of them are amenable to this technique. We then optimized EdU-click and FACS-Seq for murine fecal bacteria and reveal thatPrevotellaUCG-001 andIleibacteriumare enriched in the replicating fraction. Finally, we labelled the actively replicating murine gut bacteria during exposure to cell wall-specific antibioticsin vitro. We show that regardless of the antibiotic used, the actively replicating bacteria largely consist ofIleibacterium, suggesting the resistance of this taxon to perturbations. Overall, we demonstrate how combining EdU-click and FACSeq can identify the actively replicating gut bacteria and their link with the composition of the whole community in both homeostatic and perturbed conditions. This technique will be instrumental in elucidatingin situbacterial replication dynamics in a variety of other ecological states, including colonization and species invasion, as well as for investigating the relationship between the replication and abundance of bacteria in complex communities.
Developing cyanobacterial quorum sensing toolkits: towards interspecies coordination in mixed autotroph/heterotroph communities
Authors: Kokarakis Emmanuel J. , Rillema Rees , Ducat Daniel C. , Sakkos Jonathan K.
Published: July 2022

1.

There has been substantial recent interest in the promise of sustainable, light-driven bioproduction using cyanobacteria, including developing efforts for microbial bioproduction using mixed autotroph/heterotroph communities, which could provide useful properties, such as division of metabolic labor. However, building stable mixed-species communities of sufficient productivity remains a challenge, partly due to the lack of strategies for synchronizing and coordinating biological activities across different species. To address this obstacle, we developed an inter-species communication system using quorum sensing (QS) modules derived from well-studied pathways in heterotrophic microbes. In the model cyanobacterium, Synechococcus elongatus PCC 7942 (S. elongatus), we designed, integrated, and characterized genetic circuits that detect acyl-homoserine lactones (AHLs), diffusible signals utilized in many QS pathways. We showed that these receiver modules sense exogenously supplied AHL molecules and activate gene expression in a dose-dependent manner. We characterized these AHL receiver circuits in parallel in Escherichia coli W (E. coli W) to dissect species-specific properties, finding broad agreement, albeit with increased basal expression in S. elongatus. Our engineered “sender” E. coli strains accumulated biologically synthesized AHLs within the supernatant and activated receiver strains similarly to exogenous AHL activation. Our results will bolster the design of sophisticated genetic circuits in cyanobacterial/heterotroph consortia and the engineering of QS-like behaviors across cyanobacterial populations.

Highlights

Designed, built, and tested an inter-species quorum sensing-based communication system.

These genetic circuits can sense and respond to exogenous and secreted signals.

Circuit function in S. elongatus was comparable to E. coli, albeit with increased basal expression and lower induction ratios

Demonstrated inter-species communication in direct co-cultivation

First demonstration of inducible promoters and cross-species gene regulation in S. elongatus based on quorum sensing
Estimated Monkeypox Susceptible MSM Population in North Carolina
Authors: DeWitt Michael E. , Sampson Mindy M. , Fairman Robert T. , McNeil Candice J. , Polk Christopher , Passaretti Catherine L. , Sanders John W.
Published: July 2022

Using NHANES survey data we estimate that there are nearly 65,100 North Carolina residents who identify as men who have sex with men (MSM). Among those men, it is estimated nearly 15,700 have had at least one new sexual partner in the last year and represent the highest risk for infection and onward transmission of monkeypox. Vaccination strategies should consider vaccinating with highest priority those who are highly sexually active men who have sex with men as these sexual networks have the capacity to drive the monkeypox epidemic. Estimates of the number of MSM by county as well as the estimates of highly sexually active MSM are provided by North Carolina county in order to inform vaccination quantities and priorities given the current limited supply of vaccines.
PHIP variants associated with Chung–Jansen syndrome disrupt replication fork stability and genome integrity
Authors: Tirado-Class Neysha , Hathaway Caitlin , Chung Wendy K. , Dungrawala Huzefa
Published: July 2022

Chung–Jansen syndrome (CJS) is a rare, autosomal dominant disorder characterized by developmental delay, intellectual disability/cognitive impairment, behavioral challenges, obesity, and dysmorphic features. CJS is associated with heterozygous variants in PHIP (Pleckstrin-Homology Interacting Protein), a gene that encodes one of several substrate receptors for Cullin4-RING (CRL4) E3 ubiquitin ligase complex. Full-length PHIP, also called DCAF14, was recently identified to function as a replication stress response protein. Herein, we report the identification of two PHIP missense variants identified by exome sequencing in unrelated individuals with CJS. The variants p.D488V and p.E963G occur in different functional elements of DCAF14-WD40 repeat domain and pleckstrin homology-binding region (PBR), respectively. Using DNA fiber assays, we reveal that cells expressing either variant exhibit defective replication fork progression in conditions of replication stress. Furthermore, unlike wild-type DCAF14, both variants fail to accomplish DNA replication after exposure to genotoxic stress indicating a critical role of DCAF14 in protecting stalled replication forks. Thus, we have identified replication defects associated with CJS variants and predict replication-associated genome instability with CJS syndrome.
Development of a framework for utilising Lean Six Sigma’s intangible impacts in creating organisational innovation climate
Authors: Alblooshi Mohamed , Shamsuzzaman Mohammad , Karim Azharul , Haridy Salah , Shamsuzzoha Ahm , Badar M. Affan
Published: July 2022

Purpose

The purpose of this paper is to develop a framework that illustrates the role of Lean Six Sigma (LSS) in creating organisational innovation climate by investigating the relationship between LSS’s intangible impacts and organisational innovation climate factors.

Design/methodology/approach

A self-administrated survey questionnaire was distributed among 145 public sector officials to get their opinions on the relationship between various observable elements of LSS’s intangible impacts and organisational innovation climate factors, where a response rate of 73.8% was achieved. The collected data were demographically, descriptively and statistically analysed. Accordingly, a house-of-pillars-based framework that illustrates the role of LSS’s intangible impacts in creating innovation climate in an organisation was developed.

Findings

Results from this study indicated that LSS’s intangible impacts on organisational structure and hierarchy, culture, change adaptability, utilisation of staff and staff’s behavioural aspects are positively related to many of organisational innovation climate factors such as trust and openness, challenge and involvement, support for ideas and freedom and autonomy.

Research limitations/implications

The findings of this study are based on the data collected from public sector organisations in the UAE and are supported by relevant literature. However, this study can provide useful guidance for further research for the generalisation of the results to wider scopes in terms of sectors and geographical domains.

Practical implications

The findings of this study will provide UAE public sector officials with a clear roadmap on how to use LSS for promoting innovation and fostering its implementation in practice. This study will also encourage professionals in public sectors to integrate LSS into their innovation strategies to enhance organisational innovativeness and improve service quality.

Originality/value

It is one of the first studies that explores LSS’s intangible impacts and assesses their relationship with organisational innovation climate factors. Hence, this study offers valuable insights for both academics and practitioners and is expected to lay a foundation for a better understanding of how LSS’s intangible impacts can be used in creating organisational innovation climate.
Staying, switching, and multiplatforming of user-generated content activities: a 12-year panel study
Authors: Hou Lei , Guan Lu , Zhou Yixin , Shen Anqi , Wang Wei , Luo Ang , Lu Heng , Zhu Jonathan J.H.
Published: July 2022

Purpose

User-generated content (UGC) refers to semantic and behavioral traces created by users on various social media platforms. While several waves of platforms have come and gone, the long-term sustainability of UGC activities has become a critical question that bears significance for theoretical understanding and social media practices.

Design/methodology/approach

Based on a large and lengthy dataset of both blogging and microblogging activities of the same set of users, a multistate survival analysis was applied to explore the patterns of users' staying, switching and multiplatforming behaviors, as well as the underlying driving factors.

Findings

UGC activities are generally unsustainable in the long run, and natural attrition is the primary reason, rather than competitive switching to new platforms. The availability of leisure time, expected gratification and previous experiences drive users' sustainability.

Originality/value

The authors adopted actual behavioral data from two generations of platforms instead of survey data on users' switching intentions. Four types of users are defined: loyal, switcher, multiplatformer and dropout. As measured by the transitions among the four states, the different sustainability behaviors are thereby studied via an integrated framework. These two originalities bridge gaps in the literature and offer new insights into exploring user sustainability in social media.
Promoting transformative encounters in libraries and archives
Authors: Lowe Carli V.
Published: July 2022

Purpose

In several existing studies of Information Encountering (IE), a recurring sub-phenomenon of serendipity arises that indicates the potential for certain unexpected encounters with information to be transformative. The author labels this sub-phenomenon Transformative Information Encountering (TIE), deriving its definition from an application of Transformative Education (TE) theory to existing understandings of IE. This paper aims to discuss the potential for librarians and archivists to promote TIE through everyday practices.

Design/methodology/approach

After defining and identifying TIE in existing studies of IE, this article will put models of IE in conversation with theories of TE and propose ways in which TIE may arise in the everyday work of librarians and archivists.

Findings

In TE theory, there are three phases of the process of critical premise reflection that may be especially relevant to the work of libraries and archives. These are a disorienting dilemma (phase 1); recognition that the process of transformation is shared (phase 4); and acquiring knowledge and skills (phase 7). Each of these aligns with aspects of IE models.

Practical implications

Understanding how TIE might inform everyday Library and Information Science (LIS) work may increase the positive impact cultural institutions have on the communities they serve.

Originality/value

While several IE studies have suggested the existence of TIE as a sub-phenomenon, none thus far have attempted to define it or apply an understanding of it to LIS work.
Conceptual design of a small production plant for eco-friendly packaging
Authors: Fiorineschi Lorenzo , Conti Leonardo , Rossi Giuseppe , Rotini Federico
Published: July 2022

Purpose

This paper aims to present the application of a tailored systematic engineering design procedure to the concept design of a small production plant for compostable packaging made by straw fibres and bioplastic. In particular, the obtained boxes are intended to be used for wine bottles.

Design/methodology/approach

A systematic procedure has been adopted, which underpins on a comprehensive analysis of the design requirements and the function modelling of the process. By considering well-known models of the engineering design process, the work focuses on the early design stages that precede the embodiment design of the whole components of the plant.

Findings

The followed design approach allowed to preliminarily evaluate different alternatives of the process from a functional point of view, thus allowing to identify the preferred conceptual process solution. Based on the identified functional sequence, a first evaluation of the potential productivity and the required human resources has been performed.

Research limitations/implications

The procedure shown in this work has been applied only for the considered case of compostable packaging, and other applications are needed to optimize it. Nevertheless, the adopted systematic approach can be adapted for any context where it is necessary to conceive a new production plant for artefacts made by innovative materials.

Originality/value

The work presented in this paper represents one of the few practical examples available in the literature where systematic conceptual design procedures are presented. More specifically, to the best of the authors’ knowledge, this is the very first application of systematic design methods to compostable packaging production.
Exploring locally-produced design solutions for thermal comfort: a socio-technical assessment
Authors: Peker Ender
Published: July 2022

Purpose

The purpose of this paper is to explore the role of locally-produced architectural design solutions for the provision of thermal comfort in the vernacular settlements of Mardin, Turkey.

Design/methodology/approach

With an aim of extracting clues of climate responsive design, the paper develops a socio-technical assessment methodology and presents a comparative inquiry between the vernacular and contemporary built environments of Mardin.

Findings

Findings display that the capacity of vernacular architecture in providing a more climate responsive living environment than contemporary one is in fact correlated with the design of living spaces in harmony with the local climatic conditions as well as how inhabitants traditionally use and behave in designed space.

Originality/value

The paper argues for a need for (re)conceptualization of thermal comfort within and through the production of housing, as well as by taking into account the ways in which end-users interact, adapt and sustain end-users' everyday life in accordance with the local climatic characteristics.