bluuebunny/crossref_metadata_2025_split huggingface dataset:
Optimization of arsB (Acr3) as a positive selection marker in the cyanobacterium Synechocystis sp. PCC 6803
Authors: Tejada Irene , Florencio Francisco J. , López-Maury Luis 
Published: September 2022

AbstractThe use of cyanobacteria as bio-factories for production of numerous compounds of interest (biofuels, bioplastics …) has attracted lots of attention mainly due to their simple nutritional requirements, coupled with the decrease in atmospheric CO2 levels. However, although cyanobacteria are easily genetically manipulated, there are few genetic tools developed and, in some cases, the modifications necessary for metabolic engineering are limited for this reason. We have developed a new positive selection marker based on the arsenic resistance system of the cyanobacterium Synechocystis sp. PCC 6803. In this cyanobacterium, resistance to arsenic is mediated by the arsBHC operon in which arsB encodes an arsenite transporter whose mutation confers hypersensitivity to the presence of both arsenite and arsenate. Using arsB mutant strain (SARSB) as recipient we introduced plasmids containing both arsB and an antibiotic resistance gene and transformants were selected using either arsenic or the antibiotic with similar efficiency. The plasmids and conditions to use the arsB gene as a selectable marker have been optimized. Furthermore, we have generated an integrative vector to delete the whole arsBHC operon that allows easy introduction of regulated genes in this locus. Analysis of this strain have shown that the ΔarsBHC mutant has a higher sensitivity to arsenite than the SARSB strain, even when they are complemented with an arsB copy. These suggest that arsH, arsC or both could have an additional role in arsenite resistance.
Quantifying (non)parallelism of gut microbial community change using multivariate vector analysis
Authors: Härer Andreas , Rennison Diana J. 
Published: September 2022

AbstractParallel evolution of phenotypic traits is regarded as strong evidence for natural selection and has been studied extensively in a variety of taxa. However, we have limited knowledge of whether parallel evolution of host organisms is accompanied by parallel changes of their associated microbial communities (i.e., microbiotas), which are crucial for their hosts’ ecology and evolution. Determining the extent of microbiota parallelism in nature can improve our ability to identify the factors that are associated with (putatively adaptive) shifts in microbial communities. While it has been emphasized that (non)parallel evolution is better considered as a quantitative continuum rather than a binary phenomenon, quantitative approaches have rarely been used to study microbiota parallelism. We advocate using multivariate vector analysis (i.e., phenotypic change vector analysis) to quantify direction and magnitude of microbiota changes and discuss the applicability of this approach for studying parallelism. We exemplify its use by reanalyzing gut microbiota data from multiple fish species that exhibit parallel shifts in trophic ecology. This approach provides an analytical framework for quantitative comparisons across host lineages, thereby providing the potential to advance our capacity to predict microbiota changes. Hence, we emphasize that the development and application of quantitative measures, such as multivariate vector analysis, should be further explored in microbiota research in order to better understand the role of microbiota dynamics during their hosts’ adaptive evolution, particularly in settings of parallel evolution.
Conformational changes linked to ADP release from human cardiac myosin bound to actin-tropomyosin
Authors: Doran M.H. , Rynkiewicz M.J. , Rassici D. , Bodt S.M.L. , Barry M. E. , Bullitt E. , Yengo C.M. , Moore J.R. , Lehman W. 
Published: September 2022

AbstractFollowing binding to the thin filament, β-cardiac myosin couples ATP-hydrolysis to conformational rearrangements in the myosin motor that drive myofilament sliding and cardiac ventricular contraction. However, key features of the cardiac-specific actin-myosin interaction remain uncertain, including the structural effect of ADP release from myosin, which is ratelimiting during force generation. In fact, ADP release slows under experimental load or in the intact heart due to the afterload, thereby adjusting cardiac muscle power output to meet physiological demands. To further elucidate the structural basis of this fundamental process, we used a combination of cryo-EM reconstruction methodologies to determine structures of the human cardiac actin-myosin-tropomyosin filament complex at better than 3.4 Å-resolution in the presence and in the absence of Mg2+·ADP. Focused refinements of the myosin motor head and its essential light chains in these reconstructions reveal that small changes in the active site are coupled to significant rigid body movements of the myosin converter domain and a 16-degree lever arm swing. Our structures provide a mechanistic framework to understand the effect of ADP binding and release on human cardiac β-myosin and offer insights into the force-sensing mechanism displayed by the cardiac myosin motor.Short SummaryCryo-EM was used to elucidate high-resolution structures of actin-tropomyosin filaments decorated with human cardiac myosin in the rigor and ADP-bound states. Differences in the myosin lever arm orientation detected correlate with the overall actomyosin-linked force-sensitivity.
Nhlh1 and Nhlh2, a global transcriptional mechanism regulating commissural axon projection via Robo3 activation
Authors: Masuda Aki , Ajima Rieko , Saga Yumiko , Hirata Tatsumi , Zhu Yan 
Published: September 2022

SUMMARYCommissural neurons are highly heterogeneous in their developmental origins, neurotransmitter type and function, but all share the common feature of projecting axons across the midline. The floor plate-crossing commissural axons in mammals, from the spinal cord to the midbrain, are guided by a conserved molecular mechanism relying primarily on Netrin-1/DCC/Robo3 signaling. Up to date, we know very little about the upstream transcriptional program that specify commissural axon laterality, neither do we know if a common mechanism operates in all commissural neurons. Here, we identified a pair of highly related helix-loop-helix transcription factors, Nhlh1 and Nhlh2, as a global transcriptional mechanism that controls the laterality of all floor plate-crossing commissural axons. Forced expression of Nhlh1/2 induce ectopic Robo3 expression and contralateral axon projections. And mutant mice deficient in both genes show a remarkable reduction in Robo3 expression and a total lack of ventral commissures from the spinal cord to the midbrain. This global mechanism may interact with neuron type specific mechanism to achieve specific generation of commissural circuits.
Taurine as a key intermediate for host-symbiont interaction in the tropical spongeIanthella basta
Authors: Moeller Florian U. , Herbold Craig W. , Schintlmeister Arno , Mooshammer Maria , Motti Cherie , Behnam Faris , Watzka Margarete , Schweder Thomas , Albertsen Mads , Richter Andreas , Webster Nicole S. , Wagner Michael 
Published: September 2022

AbstractMarine sponges are critical components of marine benthic fauna assemblages where their filter-feeding and reef-building capabilities provide bentho-pelagic coupling and crucial habitat. As potentially the oldest representation of a metazoan-microbe symbiosis, they also harbor dense, diverse, and species-specific communities of microbes, which are increasingly recognized for their contributions to dissolved organic matter (DOM) processing. Recent omics-based studies of marine sponge microbiomes have proposed numerous pathways of dissolved metabolite exchange between the host and symbionts within the context of the surrounding environment, but few studies have sought to experimentally interrogate these pathways. By using a combination of metaproteogenomics and laboratory incubations coupled with isotope-based functional assays, we showed that the dominant gammaproteobacterial symbiont ‘CandidatusTaurinisymbion ianthellae’ residing in the marine sponge,Ianthella basta, expresses a pathway for the import and dissimilation of taurine, a ubiquitously occurring sulfonate metabolite in marine sponges. ‘CandidatusTaurinisymbion ianthellae’ incorporates taurine-derived carbon and nitrogen while, at the same time, oxidizing the dissimilated sulfite into sulfate for export. Furthermore, we found that taurine-derived ammonia is exported by the symbiont for immediate oxidation by the dominant ammonia-oxidizing thaumarchaeal symbiont ‘CandidatusNitrosospongia ianthellae’. Metaproteogenomic analyses also indicate that ‘CandidatusTaurinisymbion ianthellae’ likely imports DMSP and possesses both pathways for DMSP demethylation and cleavage, enabling it to use this compound as a carbon and sulfur source for biomass, as well as for energy conservation. These results highlight the important role of biogenic sulfur compounds in the interplay betweenIanthella bastaand its microbial symbionts.
Cd-induced cytosolic proteome changes in the cyanobacterium Anabaena sp. PCC7120 are mediated by LexA as one of the regulatory proteins
Authors: Srivastava Akanksha , Kumar Arvind , Biswas Subhankar , Srivastava Vaibhav , Rajaram Hema , Mishra Yogesh 
Published: September 2022

AbstractLexA, a well-characterized transcriptional repressor of the SOS genes in heterotrophic bacteria, has been shown to regulate diverse genes in cyanobacteria. An earlier study showed that LexA overexpression in a cyanobacterium, Anabaena sp. PCC7120 reduces its tolerance to Cd stress. This was later shown to be due to modulation of photosynthetic redox poising by LexA under Cd stress. However, in light of the global regulatory nature of LexA and the prior prediction of AnLexA-box in a few heavy metal-responsive genes, we speculated that LexA has a broad role in Cd stress tolerance, with regulation over a variety of Cd stress-responsive genes in addition to the regulation on genes related with photosynthetic redox poising. Thus, to further expand the knowledge on the regulatory role of LexA in Cd stress tolerance, a cytosolic proteome profiling of Anabaena constitutively overexpressing LexA upon Cd stress was performed. The proteomic study revealed 25 differentially accumulated proteins (DAPs) in response to the combined effect of LexA overexpression and Cd stress, and the other 11 DAPs exclusively in response to either LexA overexpression or Cd stress. The 36 identified proteins were related with a variety of functions, including photosynthesis, carbon metabolism, antioxidative defence, protein turnover, chaperones, post-transcriptional modifications, and a few unknown and hypothetical proteins. The regulation of LexA on corresponding genes, as well as six previously reported Cd efflux transporters, was further validated by the presence of AnLexA-boxes, transcript, and/or promoter analyses. In a nutshell, this study identifies the regulation of LexA on several genes and proteins of various functional categories in Anabaena that are responsive to Cd stress, hence expanding the regulatory role of LexA under Cd stress.HighlightsLexA overexpression was earlier shown to decrease Cd stress tolerance in Anabaena.We examined the combined effect of LexA overexpression and Cd on Anabaena proteome.Upon LexA overexpression or Cd stress or both, 36 differential proteins were found.In silico, transcript and EMSA proved LexA regulation on them and few transporters.The findings of this study extended the regulatory role of LexA in Cd tolerance.
Saccharomyces cerevisiae DJ-1 paralogs maintain genome integrity through glycation repair of nucleic acids and proteins
Authors: Susarla Gautam , Kataria Priyanka , Kundu Amrita , D’Silva Patrick 
Published: September 2022

AbstractReactive carbonyl species (RCS) such as methylglyoxal and glyoxal are potent glycolytic intermediates that extensively damage cellular biomolecules leading to genetic aberration and protein misfolding. Hence, RCS levels are crucial indicators in the progression of various pathological diseases. Besides the glyoxalase system, emerging studies report highly conserved DJ-1/ThiJ/PfpI superfamily proteins as critical regulators of RCS. DJ-1 superfamily proteins, including the human DJ-1, a genetic determinant of Parkinson’s disease possess diverse physiological functions paramount for combating multiple stressors. Although S. cerevisiae retains four DJ-1 orthologs (namely Hsp31, Hsp32, Hsp33, and Hsp34), their physiological relevance and collective requirement are still obscure due to their close sequence similarity. Here, we report for the first time that the yeast DJ-1 orthologs function as novel enzymes involved in the preferential scavenge of glyoxal and methylglyoxal, toxic metabolites, and genotoxic agents. At the cellular level, their collective loss induces chronic glycation of the proteome, and nucleic acids, resulting in a spectrum of genetic mutations and reduced mRNA translational efficiency. Furthermore, the Hsp31 paralogs efficiently repair severely glycated macromolecules derived from carbonyl modifications. They also participate in genome maintenance as their absence upregulates DNA damage response pathways when exposed to different genotoxins. Interestingly, yeast DJ-1 orthologs provide robust organellar protection by redistributing into mitochondria to alleviate the glycation damage of mitochondrial DNA and proteins. Taken together, our study uncovers the existence of a novel glycation repair pathway in S. cerevisiae and a possible neuroprotective mechanism of how hDJ-1 confers mitochondrial health during carbonyl stress.
The NanoLuc Assay System for Accurate Real-Time Monitoring of Mitochondrial Protein Import within Intact Mammalian Cells
Authors: Needs Hope I , Pereira Gonçalo C. , Henley Jeremy M , Collinson Ian 
Published: September 2022

AbstractOnly a few proteins (13 in humans) are encoded by the mammalian mitochondrial genome. Therefore, the other mitochondrial resident proteins (>1000) must be recruitedviaspecialised import pathways. Protein import is critical for mitochondrial biogenesis and bioenergetic function and health; loss of function has been implicated with a wide range of pathologies. Despite this, our understanding of the kinetic and dynamics of import is somewhat limited, particularly within mammalian cells. Here, we report an adaptation of an assay system, established previously to monitor mitochondrial import into isolated yeast mitochondria, to quantitatively monitor mitochondrial import inside mammalian cells. The reporting is based on a split luciferase, whereby the large fragment is segregated in the mitochondrial matrix and the small complementary fragment is fused to the C-terminus of a recombinant precursor protein destined for import. Following import successively through the TOM complex of the outer membrane and the TIM23 complex of the inner membrane, the complementary fragments combine to form an active luciferase. The resultant luminescent signal provides a sensitive, accurate, free of noise and continuous measure of protein import, enabling mathematical model fitting to identify and understand the steps that make up import. This advance allows detailed mechanistic examination of the transport process in live cells. In addition, the assay will enable characterisation of the protein import when the machinery is challenged; for example, in situations associated with disease. Moreover, the assay is compatible with high throughput for large data set collection and kinetic modelling, as well as for drug screening and characterisation. Our set-up also has the potential to be adapted for the analysis of alternative transport systems and different cell types, and even for multicellular model organisms.
N-linked glycosylation increases horse radish peroxidase rigidity leading to enhanced activity and stability
Authors: Ramakrishnan Krithika , Johnson Rachel L. , Winter Sam D. , Worthy Harley L. , Thomas Chris , Humer Diana , Spadiut Oliver , Hindson Sarah H. , Wells Stephen , Barratt Andrew H. , Menzies Georgina E. , Pudney Christopher R. , Jones D. Dafydd 
Published: September 2022

ABSTRACTGlycosylation is the most prevalent protein post-translational modification, with a quarter of glycosylated proteins having enzymatic properties. Yet the full impact of glycosylation on the protein structure-function relationship, especially in enzymes, is still limited. Here we show glycosylation rigidifies the important commercial enzyme horseradish peroxidase (HRP), which in turn increases its activity and stability. Circular dichroism spectroscopy revealed that glycosylation increased holo-HRP’s thermal stability and promoted significant helical structure in the absence of haem (apo-HRP). Glycosylation also resulted in a 10-fold increase in enzymatic turnover towards o-phenylenediamine dihydrochloride when compared to its non-glycosylated form. Utilising a naturally occurring site-specific probe of active site flexibility (Trp117) in combination with red-edge excitation shift fluorescence spectroscopy, we found that glycosylation significantly rigidified the enzyme. In silico simulations confirmed that glycosylation largely decreased protein backbone flexibility, especially in regions close to the active site and the substrate access channel. Thus, our data shows that glycosylation does not just have a passive effect on HRP stability but can exert long range effects that mediate the ‘native’ enzyme’s activity and stability through changes in inherent dynamics.
Antibiotic persistence does not cause phenotypic heterogeneity in tolerance of Escherichia coli to formaldehyde stress but can preserve it through time
Authors: Jordan Isaiah D. , Ticak Tomislav , Lee Jessica A. , Marx Christopher J. 
Published: September 2022

AbstractThe phenomenon of phenotypic heterogeneity, where an isogenic population expresses varying phenotypes, has been uncovered for an expanding variety of organisms and traits. This heterogeneity in phenotype can be physiologically relevant, such as the ability for antibiotic persistence to allow populations to survive lethal conditions due to rare cells being in a slow or non-growing state. Recently, it was discovered that Methylorubrum extorquens, a facultative methylotroph, possesses a continuous spectrum of phenotypic states conferring tolerance to formaldehyde. Formaldehyde is a toxin produced by M. extorquens during growth on methanol. The phenotypic tolerance allowed rapid growth on levels of formaldehyde which were lethal to the majority of the population. Transcriptomics indicated this may be due to upregulation of proteins that attenuate oxidative stress and protein damage rather than increasing formaldehyde oxidation to prevent accumulation. These data suggested that heterogeneity to formaldehyde stress may be present even in non-methylotrophic organisms that do not routinely produce large quantities of formaldehyde, and thus widely distributed across bacteria. To investigate this, we tested Escherichia coli for heterogeneity to formaldehyde stress during growth on glucose. Like M. extorquens, E. coli populations have a wide, continuous range of formaldehyde tolerance thresholds and this tolerance was reversible. Several other features, however, were different from what was found for M. extorquens. Most E. coli growth occurred after formaldehyde levels had dropped, suggesting that persistence could be the cause. The dynamics of antibiotic persistence and formaldehyde tolerance were both tracked but found to not be correlated. On the other hand, the persister cell state can maintain formaldehyde tolerance. These data suggest that persistence can preserve phenotypic heterogeneities in other traits, further expanding its potential role in helping cells survive environmental stressors.
Emergence of Supercoiling-Mediated Regulatory Networks through the Evolution of Bacterial Chromosome Organization
Authors: Grohens Théotime , Meyer Sam , Beslon Guillaume 
Published: September 2022

AbstractDNA supercoiling – the level of twisting and writhing of the DNA molecule around itself – plays a major role in the regulation of gene expression in bacteria by modulating promoter activity. The level of DNA supercoiling is a dynamic property of the chromosome, which varies both at local and global scales, in response to external factors such as environmental perturbations and to internal factors including gene transcription. As such, local variations in supercoiling could in theory couple the expression levels of neighboring genes by creating feedback loops between their transcription rates. However, the impact of such supercoiling-mediated interactions on the regulation of gene expression remains uncertain. In this work, we study how this coupling between transcription and supercoiling could shape genome organization and help regulate gene transcription. We present a model of genome evolution in which individuals whose gene transcription rates are coupled to supercoiling must adapt to two environments that induce different global supercoiling levels. In this model, we observe the evolution of whole-genome regulatory networks that provide control over gene expression by leveraging the transcription-supercoiling coupling, and show that the structure of these networks is underpinned by the organization of genes along the chromosome at several scales. Local variations in DNA supercoiling could therefore help jointly shape both gene regulation and genome organization during evolution.
Quantum analysis of P3a and P3b from auditory single trial ERPs differentiates borderline personality disorder from schizophrenia
Authors: Melkonian Dmitriy , Korner Anthony , Meares Russell , Harris Anthony 
Published: September 2022

AbstractTraditional approaches to EEG modelling use the methods of classical physics to reconstruct scalp potentials in terms of explicit physical models of cortical neuron ensembles. The principal difficulty is that the multiplicity of cellular processes with an intricate array of deterministic and random factors prevents creation of consistent biophysical parameter sets. An original, empirically-testable solution has been recently achieved in our previous studies by a radical departure from the deterministic equations of classical physics to the probabilistic reasoning of quantum mechanics. This crucial step relocates elementary bioelectric sources of EEG signals from the cellular to the molecular level where positively and negatively ions are considered as elementary sources of electricity. The rationale is that despite dramatic differences in cellular machineries, statistical factors governed by the rules of central limit theorem produce EEG waveforms as a statistical aggregate of the synchronized activity of multiple closely-located microscale sources. Using the formalism of nonhomogeneous birth-and-death processes (BDP) the quantum models of microscale events are deduced and linked to the dynamics of macroscale EEG waveforms. This study expands these methods with new features for comprehensive analysis of event related potentials directly from single trials, i.e. the EEG segments which are closely related in timing to cognitive events. We derive a universal model of the components of single trial ERPs both in frequency and time domains. This, for the first time, enables us to quantify all significant cognitive components in single trial ERPs, providing an alternative to the traditional method of averaging. Given P300 as an important objective marker of psychiatric disorders, a methodology which reliably discloses the component compositions of this potential, may have specific diagnostic importance. In this study, reliable identification of the P3a and P3b components from an auditory oddball paradigm provided a means of differentiating borderline personality disorder from schizophrenia.