bluuebunny/crossref_metadata_2025_split huggingface dataset:
Extensive genomic divergence among 61 strains ofChlamydia psittaci
Authors: Sachse Konrad , Hölzer Martin , Vorimore Fabien , Barf Lisa-Marie , Lamkiewicz Kevin , Sachse Carsten , Laroucau Karine , Marz Manja 
Published: November 2022

AbstractChlamydia (C.) psittaci, the causative agent of avian chlamydiosis and human psittacosis, is a genetically heterogeneous species. Its broad host range includes parrots and many other birds, but occasionally also humans (via zoonotic transmission), ruminants, horses, swine and rodents. To assess whether there are genetic markers associated with host tropism we comparatively analyzed whole-genome sequences of 61C. psittacistrains. Initially, poorly assembled genomes in public databases were subjected to clean-up, reassembly and polishing. Multiple sequence alignment of the genome sequences revealed four major clades within this species. Clade 1 represents the most recent lineage comprising 40/61 strains and contains 9/10 of the psittacine strains, including type strain 6BC, and 10/13 of human isolates. Clades 2-4 carry strains from different non-psittacine hosts. We found that clade membership correlates with classification schemes based on SNP types,ompAgenotypes, multilocus sequence types as well as plasticity zone (PZ) structure and host preference. Genome analysis also revealed that i) sequence variation in the major outer membrane porin OmpA can result in 3D structural changes of immunogenic domains, ii) past host change of Clade 3 and 4 strains could be associated with loss of MAC/perforin in the PZ, rather than the large cytotoxin, iii) the distinct phylogeny of atypical strains (Clades 3 and 4) is also reflected in their repertoire of inclusion proteins (Inc family) and polymorphic membrane proteins (Pmps). Altogether, our study identified a number of genomic features that can be correlated with the phylogeny and host preference ofC. psittacistrains.
Inhibition of pyruvate dehydrogenase affects the brain protein acylation stronger than PDHA phosphorylation at Ser293
Authors: Aleshin Vasily A. , Sibiryakina Daria A. , Kazantsev Alexey V. , Graf Anastasia V. , Bunik Victoria I. 
Published: November 2022

AbstractAdaptation of an organism to metabolic challenges requires mechanisms coupling metabolism to gene expression. Acylations of metabolic and histone proteins acquire significant attention in this regard. We hypothesize that adaptive response to inhibition of a key metabolic process, catalyzed by the acetyl-CoA-generating pyruvate dehydrogenase (PDH) complex, may be mediated by changed protein acylations. The hypothesis is tested by intranasal administration to animals of PDH-specific inhibitors acetylmethylphosphinate (AcMeP) or methyl ester of acetylphosphonate (AcPMe), followed by assessment of physiological parameters, brain protein acylation system and expression/phosphorylation of PDHA subunit. At a fixed dose, AcMeP, but not AcPMe, decreases acetylation and increases succinylation of the brain proteins of apparent molecular mass of 15-20 kDa. Regarding the 30-50 kDa proteins, a strong inhibitor AcMeP affects acetylation only, while a less efficient AcPMe mostly increases succinylation. No increase in the succinylation of the 30-50 kDa proteins by AcMeP coincides with its induction of desuccinylase SIRT5, not observed in the AcPMe-treated animals. The brain PDHA expression or phosphorylation, the animal behavior or ECG do not significantly differ between the studied animal groups. The data indicate that a short-term inhibition of the brain PDH affects acetylation and/or succinylation of the brain proteins, dependent on the inhibitor potency, protein molecular mass and acylation type. Homeostatic nature of these changes is implied by stability of physiological parameters after the PDH inhibition.
Precise identification of cell states altered in disease with healthy single-cell references
Authors: Dann Emma , Teichmann Sarah A. , Marioni John C. 
Published: November 2022

AbstractSingle cell genomics is a powerful tool to distinguish altered cell states in disease tissue samples, through joint analysis with healthy reference datasets. Collections of data from healthy individuals are being integrated in cell atlases that provide a comprehensive view of cellular phenotypes in a tissue. However, it remains unclear whether atlas datasets are suitable references for disease-state identification, or whether matched control samples should be employed, to minimise false discoveries driven by biological and technical confounders. Here we quantitatively compare the use of atlas and control datasets as references for identification of disease-associated cell states, on simulations and real disease scRNA-seq datasets. We find that reliance on a single type of reference dataset introduces false positives. Conversely, using an atlas dataset as reference for latent space learning followed by differential analysis against a matched control dataset leads to precise identification of disease-associated cell states. We show that, when an atlas dataset is available, it is possible to reduce the number of control samples without increasing the rate of false discoveries. Using a cell atlas of blood cells from 12 studies to contextualise data from a case-control COVID-19 cohort, we sensitively detect cell states associated with infection, and distinguish heterogeneous pathological cell states associated with distinct clinical severities. Our analysis provides guiding principles for design of disease cohort studies and efficient use of cell atlases within the Human Cell Atlas.
The effect of wheat genotype on its microbiome is more evident in roots than rhizosphere and is strongly influenced by time
Authors: Quiza Liliana , Tremblay Julien , Pagé Antoine P. , Greer Charles W. , Pozniak Curtis J. , Li Rong , Haug Brenda , Hemmingsen Sean M. , St-Arnaud Marc , Yergeau Etienne 
Published: November 2022

AbstractCrop breeding has traditionally ignored the plant-associated microbial communities. Consideration of the interactions between plant genotype and associated microbiota is of value since different genotypes of the same crop often harbor distinct microbial communities which can influence the plant phenotype. However, recent studies have reported contrasting results, which led us to hypothesize that the effect of genotype is constrained by time (growth stage, year) and space (plant compartment). To test this hypothesis, we sampled bulk soil, rhizosphere soil and roots of 10 wheat genotypes, twice per year, for 4 years. DNA was extracted and regions of the bacterial 16S rRNA and CPN60 genes and the fungal ITS region were amplified and sequenced. The effect of genotype was highly contingent on the time of sampling and on the plant compartment sampled. Only for a few sampling dates, were the microbial communities significantly different across genotypes. The effect of genotype was most often significant for root microbial communities. The three marker genes used provided a highly coherent picture of the effect of genotype. Taken together, our results confirm that microbial communities in the plant environment strongly vary temporally and spatially and that this can mask the effect of genotype.
DNA-enhanced CuAAC ligand enables live-cell detection of intracellular biomolecules
Authors: Nian Keqing , Liu Yifang , Qiu Yuchen , Zhang Zhuoyu , Brigandi Laura , Wanunu Meni , Rouhanifard Sara H. 
Published: November 2022

AbstractOf the various conjugation strategies for cellular biomolecules, Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC) is the preferred click chemistry approach due to its fast reaction rate and the commercial availability of a wide range of conjugates. While extracellular labeling of biomolecules using CuAAC has been widely adopted, intracellular labeling in live cells has been challenging as the high copper concentrations required for CuAAC reaction is toxic to biological systems. As a critical first step towards CuAAC-mediated intracellular labeling, an ultrasensitive CuAAC ligand is needed to reduce cytosolic copper concentrations while maintaining fast reaction kinetics. Here, we developed BTT-DNA, a new DNA oligomer-conjugated CuAAC ligand for click reaction biomolecular labeling. The DNA oligo attachment serves several purposes, including: 1. Increased localization of copper atoms near the ligand, which enables ligation of azide tags with much lower copper concentrations than commercially available CuAAC ligands and without the addition of exogenous copper salt; 2. Allows nucleic acid template-driven proximity ligation by choosing the attached DNA sequence, 3. Enables the liposome encapsulation and delivery of the ligand into live cells, and 4. Facilitates intracellular labeling of nascent phospholipids in live cells. We demonstrate that BTT-DNA mediated labeling has little to no effect on the overall cell health.
Intranasal self-amplifying RNA SARS-CoV-2 vaccine produces protective respiratory and systemic immunity and prevents viral transmission
Authors: Jennewein Madeleine F. , Schultz Michael D. , Beaver Samuel , Battisti Peter , Bakken Julie , Hanson Derek , Akther Jobaida , Zhou Fen , Mohamath Raodoh , Singh Jasneet , Cross Noah , Kasal Darshan N. , Ykema Matthew , Reed Sierra , Kalange Davies , Cheatwood Isabella R. , Tipper Jennifer L. , Foote Jeremy B. , King R. Glenn , Silva-Sanchez Aaron , Harrod Kevin S. , Botta Davide , Gerhardt Alana , Casper Corey , Randall Troy D. , Lund Frances E. , Voigt Emily A. 
Published: November 2022

AbstractWhile mRNA vaccines have been effective in combating SARS-CoV-2, waning of vaccine-induced antibody responses and lack of vaccine-induced respiratory tract immunity contribute to ongoing infection and transmission. In this work, we compare and contrast intranasal (i.n.) and intramuscular (i.m.) administration of a SARS-CoV-2 self-amplifying RNA (saRNA) vaccine delivered by a nanostructured lipid carrier (NLC). Both i.m. and i.n. vaccines induce potent systemic serum neutralizing antibodies, bone marrow-resident IgG-secreting cells, and splenic T cell responses. The i.n. vaccine additionally induces robust respiratory mucosal immune responses, including SARS-CoV-2-reactive lung-resident memory and lung-homing T cell populations. As a booster following previous i.m. vaccination, the i.n. vaccine also elicits the development of mucosal virus-specific T cells. Both the i.m. and i.n. administered vaccines durably protect hamsters from infection-associated morbidity upon viral challenge, significantly reducing viral loads and preventing challenged hamsters from transmitting virus to naïve cagemates. This saRNA-NLC vaccine’s potent systemic immunogenicity, and additional mucosal immunogenicity when delivered i.n., may be key for combating SARS-CoV-2 and other respiratory pathogens.
Parallel phospholipid transfer by Vps13 and Atg2 determines autophagosome biogenesis dynamics
Authors: Dabrowski Rahel , Tulli Susanna , Graef Martin 
Published: November 2022

AbstractDuring autophagy, rapid membrane assembly expands small phagophores into large double-membrane autophagosomes. Theoretical modelling predicts the majority of autophagosomal phospholipids is derived from highly efficient non-vesicular phospholipid transfer (PLT) across phagophore-ER contacts (PERCS). Currently, the phagophore-ER tether Atg2 is the only PLT protein known to drive phagophore expansionin vivo. Here, our quantitative live-cell-imaging analysis reveals poor correlation between duration and size of forming autophagosomes and number of Atg2 molecules at PERCS of starving yeast cells. Strikingly, we find Atg2-mediated PLT is non-rate-limiting for autophagosome biogenesis, because membrane tether and PLT protein Vps13 localizes to the rim and promotes expansion of phagophores in parallel with Atg2. In the absence of Vps13, the number of Atg2 molecules at PERCS determines duration and size of forming autophagosomes with an apparentin vivotransfer rate of ~200 phospholipids per Atg2 molecule and second. We propose conserved PLT proteins cooperate in channeling phospholipids across organelle contact sites for non-rate-limiting membrane assembly during autophagosome biogenesis.
The N-terminal domain of the human mitochondrial helicase Twinkle has DNA binding activity crucial for supporting processive DNA synthesis by Polymerase γ
Authors: Johnson Laura C. , Singh Anupam , Patel Smita S. 
Published: November 2022

ABSTRACTTwinkle is the ring-shaped replicative helicase within the human mitochondria with high homology to bacteriophage T7 gp4 helicase-primase. Unlike many orthologs of Twinkle, the N-terminal domain (NTD) of human Twinkle has lost its primase activity through evolutionarily acquired mutations. The NTD has demonstrated no observed activity thus far, hence its role has remained unclear. In this study, we have biochemically characterized the isolated NTD and C-terminal domain with linker (CTD) to decipher their contributions to the activities of the full-length (FL) Twinkle. This novel CTD construct hydrolyzes ATP, has weak DNA unwinding activity, and assists Polγ-catalyzed strand-displacement synthesis on short replication forks. However, CTD fails to promote multi-kilobase length product formation by Polγ in rolling-circle DNA synthesis. Thus, CTD retains all the motor functions but struggles to implement them for processive translocation. We show that NTD has DNA binding activity, and its presence stabilizes Twinkle oligomerization. The CTD oligomerizes on its own, but loss of NTD results in heterogeneously-sized oligomeric species. The CTD also exhibits weaker and salt-sensitive DNA binding compared to FL Twinkle. Based on these results, we propose that NTD directly contributes to DNA binding and holds the DNA in place behind the central channel of the CTD like a ‘doorstop’, preventing helicase slippages and sustaining processive unwinding. Consistent with this model, mtSSB compensate for the NTD loss and partially restore kilobase length DNA synthesis by CTD and Polγ. The implications of our studies are foundational for understanding the mechanisms of disease-causing Twinkle mutants that lie in the NTD.
Clamp loader processing is important during DNA replication stress
Authors: Tashjian Tommy F. , Chien Peter 
Published: November 2022

ABSTRACTThe DNA clamp loader is critical to the processivity of the DNA polymerase and coordinating synthesis on the leading and lagging strands. In bacteria the major subunit of the clamp loader, DnaX, has two forms: the essential full-length τ and shorter γ. These are conserved across bacterial species and three distinct mechanisms have been found to create them: ribosomal frameshift, transcriptional slippage, and, inCaulobacter crescentus, proteolysis. This conservation suggests that DnaX processing is evolutionarily important, but its role remains unknown.Here we find a bias against switching from expression of a wild typednaXto a nonprocessableτ-onlyallele inCaulobacter. Despite this bias, cells are able to adapt to theτ-onlyallele with little effect on growth or morphology and only minor defects during DNA damage. Motivated by transposon sequencing, we find that loss of the genesidAin theτ-onlystrain slows growth and increases filamentation. Even in the absence of exogenous DNA damage treatment, theΔsidA τ-onlydouble mutant shows induction of and dependance onrecA, likely due to a defect in resolution of DNA replication fork stalling. We find that some of the phenotypes of theΔsidA τ-onlycan be complemented by expression of γ but that an overabundance of τ-onlydnaXis also detrimental. The data presented here suggest that DnaX processing is important during resolution of replication fork stalling events during DNA replication stress.IMPORTANCEThough the presence of DnaX τ and γ forms is conserved across bacteria, different species have developed different mechanisms to make these forms. This conservation and independent evolution of mechanisms suggest that having two forms of DnaX is important. Despite having been discovered more than 30 years ago, the purpose of expressing both τ and γ is still unclear. Here, we present evidence that expressing two forms of DnaX and controlling the abundance and/or ratio of the forms is important during the resolution of replication fork stalling.
Integration of exogenous and endogenous co-stimulatory signals by CAR-Tregs
Authors: Rosado-Sánchez Isaac , Haque Manjurul , Salim Kevin , Speck Madeleine , Fung Vivian , Boardman Dominic , Mojibian Majid , Raimondi Giorgio , Levings Megan K 
Published: November 2022

ABSTRACTRegulatory T cells (Tregs) expressing chimeric antigen receptors (CAR) are a promising tool to promote transplant tolerance. The relationship between CAR structure and Treg function was studied in xenogeneic, immunodeficient mice, revealing advantages of CD28-encoding CARs. However, these models could underrepresent interactions between CAR-Tregs, antigen-presenting cells (APCs) and donor-specific antibodies. We generated mouse Tregs expressing HLA-A2-specific CARs with different costimulatory domains and compared their function in vitro and in vivo. In vitro assays revealed the CD28-encoding CAR had superior antigen-specific suppression, proliferation and cytokine production. In contrast, in vivo protection from skin allograft rejection and alloantibody production was similar between Tregs expressing CARs encoding CD28, ICOS or PD1, but not GITR, 41BB or OX40, co-stimulatory domains. To reconcile in vitro and in vivo data, we analyzed effects of a CAR encoding CD3ζ but no co-stimulatory domain. These data revealed that exogenous co-stimulation via APCs can compensate for the lack of a CAR-encoded CD28 domain. Thus, Tregs expressing a CAR with or without CD28 are functionally equivalent in vivo. This study reveals a new dimension of CAR-Treg biology and has important implications for the design of CARs for clinical use in Tregs.
Predicting pathological complete response to neoadjuvant chemotherapy in breast cancer from routine diagnostic histopathology biopsies
Authors: Aswolinskiy Witali , Munari Enrico , Horlings Hugo M. , Mulder Lennart , Bogina Giuseppe , Sanders Joyce , Liu Yat-Hee , van den Belt-Dusebout Alexandra W. , Tessier Leslie , Balkenhol Maschenka , Hoven Jeffrey , Wesseling Jelle , van der Laak Jeroen , Lips Esther H. , Ciompi Francesco 
Published: November 2022

AbstractPurposeInvasive breast cancer patients are increasingly being treated with neoadjuvant chemotherapy, however, only a fraction of the patients respond to it completely. To prevent over-treating patients with a toxic drug, there is an urgent need for biomarkers capable of predicting treatment response before administering the therapy. In this retrospective study, we developed interpretable, deep-learning based biomarkers to predict the pathological complete response (pCR, i.e. the absence of tumor cells in the surgical resection specimens) to neoadjuvant chemotherapy from digital pathology H&E images of pre-treatment breast biopsies.Experimental DesignOur approach consists of two steps: In the first step, using deep learning, mitoses are detected and the tissue segmented into several morphology compartments including tumor, lymphocytes and stroma. In the second step, computational biomarkers are derived from the segmentation and detection output to encode slide-level relationships between the morphological structures with focus on tumor infiltrating lymphocytes (TILs). We developed and evaluated our method on slides from N=721 patients from three European medical centers with triple-negative and Luminal B breast cancers.ResultsThe investigated biomarkers yield statistically significant prediction performance for pCR with areas under the receiver operating characteristic curve between 0.66 and 0.88 depending on the cancer subtype and center.ConclusionThe proposed computational biomarkers predict pathological complete response, but will require more evaluation and finetuning for clinical application. The results further corroborate the potential role of deep learning to automate TILs quantification, and their predictive value in breast cancer neoadjuvant treatment planning.
Accuracy of left ventricular mechanical dyssynchrony indices for mechanical characteristics of left bundle branch block using cardiovascular magnetic resonance feature tracking
Authors: Loewenstein Daniel E. , Wieslander Björn , Heiberg Einar , Axelsson Jimmy , Klem Igor , Nijveldt Robin , Schelbert Erik B. , Sörensson Peder , Sigfridsson Andreas , Strauss David G. , Kim Raymond J. , Atwater Brett D. , Ugander Martin 
Published: November 2022

AbstractAimsMore than 90% of patients with left bundle branch block (LBBB) and reduced left ventricular (LV) ejection fraction have LV dyssynchrony and a high probability of response to cardiac resynchronization therapy (CRT). A subgroup of patients with non-specific intraventricular conduction delay (IVCD) have a LBBB-like LV activation pattern when studied using invasive mapping and advanced echocardiographic techniques. These patients also frequently benefit from CRT but these patients have proven difficult to identify using ECG criteria. Cardiovascular magnetic resonance (CMR) imaging indices of dyssynchrony may identify patients with IVCD who may benefit from CRT but their relative accuracies for identification of LV dyssynchrony remains unknown. We compared the LV dyssynchrony classification accuracy of two commonly available CMR indices in a study population of patients with severely reduced LV ejection fraction and no scar, and either LBBB or normal conduction (normal QRS duration and axis, controls).Methods and resultsIn LBBB (n=44) and controls (n=36), using CMR feature tracking circumferential strain, dyssynchrony was quantified as the circumferential uniformity ratio estimate (CURE) and the systolic stretch index (SSI). Deidentified CMR image-data were made publicly available. Both CURE and SSI quantified more severe dyssynchrony in LBBB compared to controls (p<0.001 for both). SSI more frequently discriminated LBBB and normal conduction LV activation patterns than CURE (area under the receiver operating characteristics curve [95% confidence interval] 0.98 [0.95-1.00] for SSI vs 0.77 [0.66-0.86] for CURE, p<0.001).ConclusionsSSI is superior to CURE for discriminating synchronous and dyssynchronous LV activation and should be further studied in the setting of non-LBBB conduction abnormalities.