bluuebunny/crossref_metadata_2025 huggingface dataset:
Pulvinar Slits: Cellulose-deficient and De-Methyl-Esterified Homogalacturonan-Rich Structures in a Legume Motor Cell
Authors: Takahara Masahiro , Tsugawa Satoru , Sakamoto Shingo , Demura Taku , Nakata Miyuki T
Published: March 2022

The cortical motor cells (CMCs) in a legume pulvinus execute the reversible deformation in leaf movement that is driven by changes in turgor pressure. In contrast to the underlying osmotic regulation proper, the cell wall structure of CMCs that contributes to the movement has yet to be characterized in detail. Here, we report that the cell wall of CMCs has circumferential slits with low levels of cellulose deposition. This structure is unique and distinct from that of any other primary cell walls reported so far; thus, we named them “pulvinar slits.” Notably, we predominantly detected de-methyl-esterified homogalacturonan inside pulvinar slits, with a low deposition of highly methyl-esterified homogalacturonan, as with cellulose. In addition, Fourier-transform infrared spectroscopy analysis indicated that the cell wall composition of pulvini is different from that of other axial organs, such as petioles or stems. Moreover, monosaccharide analysis showed that pulvini are pectin-rich organs like developing stems and that the amount of galacturonic acid in pulvini is greater than in developing stems. Computer modeling suggested that pulvinar slits facilitate anisotropic extension in the direction perpendicular to the slits in the presence of turgor pressure. When tissue slices of CMCs were transferred to different extracellular osmotic conditions, pulvinar slits altered their opening width, indicating their deformability. In this study, we thus characterized a distinctive cell wall structure of CMCs, adding new knowledge to repetitive and reversible organ deformation as well as the structural diversity and function of the plant cell wall.
Robustness of field studies evaluating biodiversity responses to invasive species management in New Zealand
Authors: Allen Robert B. , Forsyth David M. , MacKenzie Darryl I. , Peltzer Duane A.
Published: March 2022

Benefits of invasive species management for terrestrial biodiversity are widely expected and promoted in New Zealand. Evidence for this is presented in policy and scientific reviews of the literature, but the robustness and repeatability of the underpinning evidence-base remains poorly understood. We evaluated the design of field-based studies assessing biodiversity responses to invasive species management in 155 peer-reviewed articles published across 46 journals from 2010 - 2019. Each study was assessed against nine principles of experimental design, covering robustness of sampling and avoidance of bias. These principles are important in New Zealand to detect treatment effects from environmental variability driven by underlying gradients such as soil fertility, climate and disturbance. Fifty two percent of studies defined a sampling universe and 68% of studies specified the treatment. Whereas, 54%, 74%, and 50% of studies did not utilise replication, representatively sample the universe, or quantify invasive species, respectively. Ninety five percent of studies quantified biodiversity responses, although a high proportion of these did not representatively sample replicates. Initial conditions and accounting for effects of experimental implementation were not utilised in 57% and 84% of studies respectively. No studies avoided observer/analyst bias using blinding methods, despite this being widely adopted in other fields. Ordinal logistic regression showed these principles varied in how robustly they were applied among categories of biodiversity responses and invasive species. Our findings suggest that greater attention to experimental design principles is desirable: supported by researchers, funding agencies, reviewers, and journal editors. Greater resources is not necessarily a solution to these design issues. Undertaking fewer studies, that are individually more expensive because they better adhere to experimental design principles, is one alternative. Our intent in this article is to improve the robustness of future field studies for at least some principles. Robust designs have enduring value, reduce uncertainty and increase our understanding of when, where and how often the impacts of invasive species on biodiversity are indeed reversible.
SARS-CoV-2 reinfections during the first three major COVID-19 waves in Bulgaria
Authors: Marinov Georgi K. , Mladenov Mladen , Rangachev Antoni , Alexiev Ivailo
Published: March 2022

Background

The COVID-19 pandemic has had a devastating impact on the world over the past two years (2020-2021). One of the key questions about its future trajectory is the protection from subsequent infections and disease conferred by a previous infection, as the SARS-CoV-2 virus belongs to the coronaviruses, a group of viruses the members of which are known for their ability to reinfect convalescent individuals. Bulgaria, with high rates of previous infections combined with low vaccination rates and an elderly population, presents a somewhat unique context to study this question.

Methods

We use detailed governmental data on registered COVID-19 cases to evaluate the incidence and outcomes of COVID-19 reinfections in Bulgaria in the period between March 2020 and early December 2021.

Results

For the period analyzed, a total of 4,106 cases of individuals infected more than once were observed, including 31 cases of three infections and one of four infections. The number of reinfections increased dramatically during the Delta variant-driven wave of the pandemic towards the end of 2021. We observe a moderate reduction of severe outcomes (hospitalization and death) in reinfections relative to primary infections, and a more substantial reduction of severe outcomes in breakthrough infections in vaccinated individuals.

Conclusions

In the available datasets from Bulgaria, prior infection appears to provide some protection from severe outcomes, but to a lower degree than the reduction in severity of breakthrough infections in the vaccinated compared to primary infections in the unvaccinated.
Seasonality in living kidney donation in the United States from 1995-2019
Authors: Arking Andrew , Kaddu Gabriella , Segev Dorry L. , Garonzik-Wang Jacqueline , Muzaale Abimereki D. , Ammary Fawaz Al
Published: March 2022

For nearly two decades, the annual number of US living kidney donors has been characterized by worrying patterns of decline and no factors have been identified to explain and reverse these patterns. Evidence suggests that there is seasonality in living kidney donation; herein we investigate whether potentially modifiable social, economic, and structural issues might explain this seasonality. Using donor-registry data from the Scientific Registry of Transplant Recipients, we described this seasonality in living kidney donation and used Poisson regression stratified by both donor–recipient biological relationship and estimated household income tertile to quantify these trends. In every decade from 1980-2020, there was a summer-only surge in living kidney donations (13%-25% for biologically related donors and 10%-17% for unrelated donors). This summer-only surge was evident for the months of June, July, and August when compared with January for each given year and statistically significant in some groups (range of incidence rate ratio [IRR] for related donors: 1.05-1.34; IRR for unrelated donors: 1.08-1.19). We observed this summer-only surge across all three income tertiles ($73,544+, $52,635- $73,544, and <$52,635) and regardless of donor-recipient relationship. Seasonal variation in donation is associated with structural factors, which may serve as potential targets for interventions to increase donation.
Clinical improvement of DM1 patients reflected by reversal of disease-induced gene expression in blood
Authors: van Cruchten Remco T.P. , van As Daniël , Glennon Jeffrey C. , van Engelen Baziel G.M. , ’t Hoen Peter A. C. ,   the OPTIMISTIC consortium,   the ReCognitION consortium
Published: March 2022

Myotonic dystrophy type 1 (DM1) is an incurable multisystem disease caused by a CTG-repeat expansion in the DM1 protein kinase (DMPK) gene. The OPTIMISTIC clinical trial demonstrated positive and heterogenous effects of cognitive behavioral therapy (CBT) on the capacity for activity and social participations in DM1 patients. Here, we performed mRNA sequencing of full blood for 27 patients of the OPTIMISTIC cohort before and after the CBT intervention. We identified 608 genes for which their expression was significantly associated with the disease causing CTG-repeat expansion, as well as 1176 genes significantly associated with the average clinical response towards the intervention. Remarkably, all 97 genes significantly associated with both returned to more normal levels in patients who benefited most from CBT. This trend was consistent with the difference observed between DM1 patients and controls in an earlier study of blood mRNA expression levels, singling these genes out as candidate biomarkers for therapy response. Together these results highlight the ability to find disease relevant information in full blood of DM1 patients, opening new avenues to monitor therapy effects.
Do recent family physician graduates practice differently? A longitudinal study of primary care visits and continuity in four Canadian provinces
Authors: Rudoler David , Peterson Sandra , Stock David , Taylor Carole , Wilton Andrew S. , Blackie Doug , Burge Fred , Glazier Richard , Goldsmith Laurie J. , Grudniewicz Agnes , Hedden Lindsay , Jamieson Margaret , Katz Alan , MacKenzie Adrian , Marshall Emily Gard , McCracken Rita , McGrail Kimberlyn , Scott Ian , Wong Sabrina T. , Lavergne M. Ruth
Published: March 2022

Background: Complaints about lack of access to family physicians (FPs) has led to concerns about the role recent physician graduates have had in changes in the supply of primary care services in Canada. This study investigates the impact of career stage, time period, and graduation cohort on family physician practice volume and continuity over two decades.

Methods: Retrospective-cohort study of family physician practice from 1997/98 to 2017/18. Administrative health and physician claims data were collected in British Columbia, Manitoba, Ontario and Nova Scotia. The study included all physicians registered with their respective provincial regulatory colleges with a medical specialty of family practice and/or billed the provincial health insurance system for patient care as family physicians. Median polish analysis of patient contacts and physician-level continuity was completed to isolate years-in-practice, period, and cohort effects.

Results: Median patient contacts per provider fell over time in the four provinces examined. In all four provinces, median contacts increased with years in practice until mid-to-late-career and declined into end-of-career. We found no relationship between graduation cohort and practice volume or FP-level continuity.

Interpretation: Recent cohorts of family physicians practice similarly to predecessors in terms of practice volumes and continuity of care. Since FPs of all career stages show declining patient contacts, system-wide solutions to recent challenges in the accessibility of primary care in Canada are needed.
Rates of regional tau accumulation in ageing and across the Alzheimer's disease continuum: An AIBL 18F-MK6240 PET study
Authors: Krishnadas Natasha , Doré Vincent , Robertson Jo , Ward Larry , Fowler Christopher , Masters Colin L , Bourgeat Pierrick , Fripp Jurgen , Villemagne Victor L , Rowe Christopher C
Published: March 2022

Background
Tau PET imaging enables prospective longitudinal observation of the rate and location of tau accumulation in Alzheimer's disease (AD). 18F-MK6240 is a newer, high affinity tracer for the paired helical filaments of tau in AD. It is widely used in clinical trials, despite sparse longitudinal natural history data. We aimed to evaluate the impact of disease stage, and two reference regions on the magnitude and effect size of regional change.
Methods
One hundred and fifty-eight participants: 83 cognitively unimpaired (CU) Aβ-, 37 CU Aβ+, 19 mild cognitively impaired (MCI) Aβ+ and 19 AD Aβ+ had annual 18F-MK6240 PET for one or two years (mean 1.6 years). Standardized uptake value ratios (SUVR) were generated for three in-house composite ROI: mesial temporal (Me), temporoparietal (Te), and rest of neocortex (R), and a Free-Surfer derived meta-temporal (MT) ROI. Two reference regions were examined: cerebellar cortex (SUVRCb) and eroded subcortical white matter (SUVRWM).
Results
Low rates of accumulation were seen in CU Aβ-, predominantly in the mesial temporal lobe (MTL). In CU Aβ+, increase was greatest in the MTL, particularly the amygdala. In MCI Aβ+, a similar increase was seen in MTL, but also globally in the cortex. In AD Aβ+, greatest increase was in temporoparietal and frontal regions, with a decrease in the MTL. In CU and MCI increases were greater using SUVRWM. In AD, the SUVRCb showed marginally greater increase. Interpolation of the data suggests it takes approximately two decades to accumulate tau to the typical levels found in AD, similar to the rates of accumulation of Aβ plaques.
Conclusions
The rate of tau accumulation varies according to brain region and baseline disease stage, confirming previous reports. The PET measured change is greater, with fewer outliers, using an eroded white matter reference region, except in AD. While the eroded subcortical white matter reference may be preferred for trials in preclinical AD, the cerebellar cortex would be preferred for trials in symptomatic AD.
The Microbiome in Adult Acute Appendicitis
Authors: Lee Mei Sze , Sulit Arielle , Frizelle Frank , Purcell Rachel
Published: March 2022

Acute appendicitis is a common acute surgical emergency; however, the pathogenesis of adult appendicitis remains poorly understood. The microbiome is increasingly thought to play a key role in inflammatory disease of the bowel and similarly, may play a role in appendicitis.

Aim

This study aimed to characterize the microbiome of adult acute appendicitis in a prospective cohort.

Method

We recruited 60 adults with acute appendicitis and 20 healthy controls. Rectal swabs were taken from each patient. After DNA extraction, 16S rRNA amplicon sequencing was carried out for analysis of diversity and taxonomic abundance.

Results

Phylogenetic sequencing of the samples indicated that there is a difference between the microbial composition of those with acute appendicitis and healthy controls, with a statistically significant decrease in alpha diversity in rectal swabs of appendicitis patients compared to healthy controls. At the genus level, we saw an increased abundance of potential pathogens, e.g. Parvimonas and Acinetobacter, and a decrease in commensal taxa such as Faecalibacterium, Blautia and Lachnospiraceae in appendicitis patients compared to healthy controls.

Conclusion

An imbalance in the gut microbiome may contribute to the pathogenesis of adult acute appendicitis.
Comparison of radiation dose and cancer risk between conventional polychromatic imaging and gemstone spectral imaging in computed tomography (CT) examination: a phantom study
Authors: Wu Yuliang , Li Songyang , Fang Tianqi , Deng Wei , Liu Guoqing , Wu Jiabao , Chen Xianjie , Chen Hanwei , Ma Jinting , Huang Bingsheng
Published: March 2022

This study aims to measure the radiation doses and estimate the associated lifetime attributable risks (LARs) of cancer incidence resulted from gemstone spectral imaging (GSI) and conventional polychromatic imaging (CPI) in computed tomography (CT) examinations. Organ doses were measured using an adult phantom and thermoluminescent dosimeters. Four scans, including head CT, thorax CT, abdomen CT and pelvis CT, were performed on the phantom. LARs of cancer incidence were estimated for Chinese population and US population. The effective doses of thorax CT was the highest. With CPI, it was 7.94 mSv for females, and 7.74 mSv for males. With GSI, it was 6.81 mSv for females, and 7.69 mSv for males. With GSI, the corresponding LARs for males and females aged 20-70 years were less than 0.062% and 0.154%, respectively. With CPI, these values were less than 0.074% and less than 0.171%, respectively. The LARs decreased in both US population and Chinese population when exposed age increased, with thorax scan causing the highest risk. GSI does not increased radiation exposure or cancer risk compared to CPI. These findings may allow the application of GSI in patients referred for CT.
Immortal time bias in older vs younger age groups: a simulation study with application to a population-based cohort of patients with colon cancer
Authors: Pilleron Sophie , Maringe Camille , Morris Eva JA , Leyrat Clémence
Published: March 2022

Background

In observational studies, the risk of immortal time bias (ITB) increases with the likelihood of early death, itself increasing with age. We investigated how age impacts the magnitude of ITB when estimating the effect of surgery on one-year overall survival (OS) in patients with stage IV colon cancer aged 50-74 and 75-99 in England.

Method

Using simulations, we compared estimates from a time-fixed exposure model to three methods addressing ITB: time-varying exposure, delayed entry, and landmark methods. We then estimated the effect of surgery on one-year OS using a national population-based cohort of patients derived from the CORECT-R resource.

Results

In simulations, the magnitude of ITB was larger among older patients when their probability of early death increased or treatment was delayed. The bias was corrected using the methods addressing ITB. When applied to CORECT-R data, these methods yielded smaller effects than the time-fixed exposure approach but effects were similar in both age groups.

Conclusion

ITB must be addressed in all longitudinal studies, particularly, when investigating the effect of an exposure on an outcome in different groups of people (e.g., age groups) with different distributions of exposure and outcomes.
Different structural variant prediction tools yield considerably different results in Caenorhabditis elegans
Authors: Lesack Kyle , Mariene Grace M. , Andersen Erik C. , Wasmuth James D.
Published: March 2022

The accurate characterization of structural variation is crucial for our understanding of how large chromosomal alterations affect phenotypic differences and contribute to genome evolution. Whole-genome sequencing is a popular approach for identifying structural variants, but the accuracy of popular tools remains unclear due to the limitations of existing benchmarks. Moreover, the performance of these tools for predicting variants in non-human genomes is less certain, as most tools were developed and benchmarked using data from the human genome.

To address this problem, multiple short- and long-read tools were benchmarked using real and simulated Caenorhabditis elegans whole-genome sequence data. To evaluate the use of long-read data for the validation of short-read predictions, the agreement between predictions from a short-read ensemble learning method and long-read tools were compared. The results obtained from simulated data indicate that the best performing tool is contingent on the type and size of the variant, as well as the sequencing depth of coverage. These results also highlight the need for reference datasets generated from real data that can be used as ‘ground truth’ in benchmarks.
Comparing the potential of microglia preparations generated using different human iPSC-based differentiation methods to model microglia-mediated mechanisms of amyotrophic lateral sclerosis pathophysiology
Authors: Tang Ye Man , Pulimood Nisha S. , Stifani Stefano
Published: March 2022

Microglia are the resident immune cells of nervous system. In healthy conditions, microglia actively patrol neural tissues in a homeostatic state, which can rapidly change to an activated state in response to local injury/disease. Dysregulated microglia activation is a hallmark of disorders and diseases of the nervous system, including motor neuron diseases such as amyotrophic lateral sclerosis (ALS). The elucidation of the roles of microglia in human biology and disease has recently benefitted from the development of human induced pluripotent stem cell (iPSC)-based approaches to generate microglia-like cells. Microglia represent a heterogeneous group of cells with spatial diversity in both health and disease. This situation poses a considerable challenge along the path towards establishing the most pathologically-relevant human iPSC-derived microglia preparations to investigate the complex roles of microglia in ALS and other neurological diseases. The success of these approaches must account for microglia diversity in different regions of the brain and spinal cord. In this study, we compared the transcriptomes of human iPSC-derived microglia generated using different methods to determine whether or not separate strategies can be used to generate microglia with distinct transcriptional signatures in vitro. We show that microglia derived using two different methods display distinct in vitro maturation characteristics. We also reveal that different derivation methods give rise to preparations comprising human microglia with distinct transcriptomic signatures resembling the gene profiles of specific microglia subpopulations in vivo. These findings suggest that a careful, and coordinated, implementation of multiple microglia differentiation methods from human iPSCs can be an effective approach towards the goal of generating multiple microglia subtypes that will offer enhanced model systems to account for microglia heterogeneity in vivo. Spatially-defined human iPSC-derived microglia would represent an enhanced tool to study the multiple levels of involvement of microglia in mechanisms of motor neuron degeneration in ALS, as well as other neurological diseases and disorders.
THE STABILIZATAION OF C-MYC BY THE NOVEL CELL CYCLE REGULATOR, SPY1, DECREASES EFFICACY OF BREAST CANCER TREATMENTS
Authors: Ferraiuolo Rosa-Maria , Fifield Bre-Anne , Hamm Caroline , Porter Lisa Ann
Published: March 2022

Purpose

c-Myc is frequently upregulated in breast cancers, however, targeting c-Myc has proven to be a challenge. Targeting of downstream mediators of c-Myc, such as the ‘cyclin-like’ cell cycle regulator Spy1, may be a viable therapeutic option in a subset of breast cancer subtypes.

Methods

Mouse mammary tumour cells isolated from MMTV-Myc mice and human breast cancer cell lines were used to manipulate Spy1 levels followed by tamoxifen or chemotherapeutic treatment with a variety of endpoints. Patient samples from TNBC patients were obtained and constructed into a TMA and stained for c-Myc and Spy1 protein levels.

Results

Over time, MMTV-Myc cells show a decreased response to tamoxifen treatment with increasing levels of Spy1 in the tamoxifen-resistant cells. shRNA against Spy1 re-establishes tamoxifen sensitivity. Spy1 was found to be highly elevated in human TNBC cell and patient samples, correlating to c-Myc protein levels. c-Myc was found to be stabilized by Spy1 and knocking down Spy1 in TNBC cells shows a significant increase in response to chemotherapy treatments.

Conclusions

Understanding the interplay between protein expression level and response to treatment is a critical factor in developing novel treatment options for breast cancer patients. These data have shown a connection between Spy1 and c-Myc protein levels in more aggressive breast cancer cells and patient samples. Furthermore, targeting c-Myc has proven difficult, these data suggest targeting Spy1 even when c-Myc is elevated can confer an advantage to current chemotherapies.
Opiate responses are controlled by interactions ofOprm1andFgf12loci in rodents: Correspondence to human GWAS findings
Authors: Lemen Paige M. , Zuo Yanning , Hatoum Alexander S. , Dickson Price E. , Mittleman Guy , Agrawal Arpana , Reiner Benjamin C. , Berrettini Wade , Ashbrook David G. , Gunturkun Mustafa Hakan , Mulligan Megan K. , Williams Robert W , Telese Francesca , Chen Hao
Published: March 2022

We mapped high-precision time-series data (15 min bins for 3 hours) generated for ~ 700 adult BXD mice across 105 morphine- and naloxone-related traits using new sequence-derived marker maps and a linear-mixed model. We confirm a previously mapped sex-independent effect of initial locomotor responses to morphine (50 mg/kg ip) that maps precisely toOprm1on chromosome (Chr) 10, with the linkage score reaching −log10P of ~12.4 (with a high B allele) at 75 min and exhausted by 160 min. We detected a new modulator of opiate locomotor activation in both sexes on Chr 16, with a peak linkage that climbs from 105 through to 180 min after injection. This locus includes one compelling candidate—fibroblast growth factor 12 (Fgf12). We also detected a strong, but transient epistatic interaction between these two loci. Single nuclei transcriptomic analyses in rats demonstrates that expression ofOprm1andFgf12mRNA covary in one specific subtype ofDrd1medium spiny neurons. Our Bayesian network analysis identified that a cascade of MAP kinases—Mapk8ip2, Map3k11, andMap3k12—are part of theOprm1–Fgf12network. This is the first demonstration of a time-dependent epistatic interaction modulating drug response in mammals with interesting mechanistic implications. Analysis ofOPRM1andFGF12gene networks in human GWAS data highlights enrichment of signals associated with substance use disorder.
Use of Avian Adeno-Associated Virus for the Delivery of Transgenes to Pekin Duck Cells in vitro and in vivo
Authors: Robinson Carleigh R. , Fraley Gregory S. , Kopek Benjamin G.
Published: March 2022

The Pekin duck was domesticated between 4000 and 10,000 years ago from the Mallard duck and is the predominate meat type duck in the world (Cherry and Morris, 2008). The global production of waterfowl is a rapidly growing industry. Total meat duck production increased from 2.9 million tons in 2000 to nearly 4.4 million tons in 2013, a growth rate of 3.2% per year, and further increased to 7.2 million tons in 2018, and valued at $19B in 2019 (IndexBox, 2019). Pekin ducks (Anas platyrhynchus domesticus) are the fastest growing poultry species growing from hatch to 4.5 kg market weight in as little as 28 days (Blois et al., 2019; Campbell et al., 2015; Cherry and Morris, 2008). Thus, there is a need to study the growth and reproduction of this economically important species. While Pekin duck biology is being explored by many researchers, there are fewer molecular tools available for duck compared to other poultry species and many fewer compared to mammalian systems. For example, one molecular tool commonly used to interrogate other systems are adeno-associated virus (AAV) vectors. AAV vectors are being utilized by many researchers to deliver transgenes to target tissues or cells and for genetic manipulation. Recently, avian adeno-associated virus (A3V) has been used to deliver genes to the cells and neurons of the domestic chicken (Gallus gallus domesticus). Here, we show that A3V can be used to deliver genes to Pekin duck neurons and cells in culture. Together, these results suggest that A3V will be a useful molecular tool in Pekin duck research.

Summary Statement

Here we demonstrate the use of viral vectors to deliver transgenes to Pekin duck cells. These vectors can be used to advance understanding of reproduction in this economically important species.
Retained introns in long RNA-seq reads are not reliably detected in sample-matched short reads
Authors: David Julianne K. , Maden Sean K. , Wood Mary A. , Thompson Reid F. , Nellore Abhinav
Published: March 2022

There is growing interest in retained introns in a variety of disease contexts including cancer and aging. Many software tools have been developed to detect retained introns from short RNA-seq reads, but reliable detection is complicated by overlapping genes and transcripts as well as the presence of unprocessed or partially processed RNAs. We compared introns detected by 5 tools using short RNA-seq reads with introns observed in long RNA-seq reads from the same biological specimens and found: (1) significant disagreement among tools (Fleiss’ κ = 0.231) such that 52.4% of all detected intron retentions were not called by more than one tool; (2) that no tool achieved greater than 20% precision or 35% recall under generous conditions; and (3) that retained intron detectability was adversely affected by greater intron length and overlap with annotated exons.
Integration of multi-omics data reveals interplay between brassinosteroid and TORC signaling in Arabidopsis
Authors: Montes Christian , Wang Ping , Liao Ching-Yi , Nolan Trevor M , Song Gaoyuan , Clark Natalie M , Elmore J. Mitch , Guo Hongqing , Bassham Diane C , Yin Yanhai , Walley Justin W
Published: March 2022

Brassinosteroids (BR) and Target of Rapamycin Complex (TORC) are two major actors coordinating plant growth and stress responses. BRs function through a signaling pathway to extensively regulate gene expression and TORC is known to regulate translation and autophagy. Recent studies revealed that these two pathways crosstalk, but a system-wide view of their interplay is still missing. Thus, we quantified the level of 23,975 transcripts, 11,183 proteins, and 27,887 phosphorylation sites in wild-type Arabidopsis and in mutants with altered levels of either BRASSINOSTEROID INSENSITIVE 2 (BIN2) or REGULATORY ASSOCIATED PROTEIN OF TOR 1B (RAPTOR1B), two key players in BR and TORC signaling, respectively. We found that perturbation of BIN2 or RAPTOR1B levels affects a common set of gene-products involved in growth and stress responses. Furthermore, we used the multi-omic data to reconstruct an integrated signaling network. We screened 41 candidate genes identified from the reconstructed network and found that loss of function mutants of many of these proteins led to an altered BR response and/or modulated autophagy activity. Altogether, these results establish a predictive network that defines different layers of molecular interactions between BR-or TORC-regulated growth and autophagy.
Interstrand Crosslinks in Donor DNA Boost Gene Editing in Human Cells
Authors: Ghasemi Hannah I. , Bacal Julien , Yoon Amanda , Cruz Carmen , Vu Jonathan , Gardner Brooke , Richardson Chris D.
Published: March 2022

Co-introduction of targeted nucleases and DNA/RNA templates encoding new genomic sequence is the basis for rapid, effective, and iterable gene editing workflows for therapeutic, agricultural, and basic science applications. Extensive optimization of reagent delivery and nuclease activity have improved genome editing workflows, but comparatively few efforts have been made to alter the gene editing activity of template molecules. Here, we report template DNA modified with interstrand crosslinks (ICLs) – xHDRTs - increases editing frequencies in Cas9-directed gene editing workflows by up to five-fold. xHDRTs increase gene editing frequencies independent of DNA template topology, amount of sequence added, or cell type. Gene editing using xHDRTs requires the DNA repair kinase, ATR, and partially requires Fanconi Anemia proteins, including FANCA, but is independent of other ICL-repair pathways. Covalent modification of donor DNA thus presents a compelling opportunity to improve nonviral gene editing workflows.
Species-specific chromatin landscape determines how transposable elements shape genome evolution
Authors: Huang Yuheng , Shukla Harsh , Lee Yuh Chwen G.
Published: March 2022

Transposable elements (TEs) are selfish genomic parasites that increase their copy number at the expense of host fitness. The “success,” or genome-wide abundance, of TEs differs widely between species. Deciphering the causes for this large variety in TE abundance has remained a central question in evolutionary genomics. We previously proposed that species-specific TE abundance could be driven by the inadvertent consequences of host-direct epigenetic silencing of TEs—the spreading of repressive epigenetic marks from silenced TEs into adjacent sequences. Here, we compared this TE-mediated “epigenetic effect” in six species in the Drosophila melanogaster subgroup to dissect step-by-step the role of such effect in determining genomic TE abundance.

We found that TE-mediated spreading of repressive marks is prevalent and substantially varies across and even within species. While this TE-mediated effect alters the epigenetic states of adjacent genes, we surprisingly discovered that the transcription of neighboring genes could reciprocally impact this spreading. Importantly, our multi- species analysis provides the power and appropriate phylogenetic resolution to connect species-specific host chromatin regulation, TE-mediated epigenetic effects, the strength of natural selection against TEs, and genomic TE abundance unique to individual species. Our findings point towards the importance of host chromatin landscapes in shaping genome evolution through the epigenetic effects of a selfish genetic parasite.
Evolutionary Principles of Bacterial Signaling Capacity and Complexity
Authors: Mo Ran , Liu Yugeng , Chen Yuanyuan , Mao Yingjin , Gao Beile
Published: March 2022

Microbes rely on signal transduction systems to sense and respond to environmental changes for survival and reproduction. It is generally known that niche adaptation plays an important role in shaping the signaling repertoire. However, the evolution of bacterial signaling capacity lacks systematic studies with a temporal direction. Particularly, it is unclear how complexity evolved from simplicity or vice versa for signaling networks. Here we examine the evolutionary processes of major signal transduction systems in Campylobacterota (formerly Epsilonproteobacteria), a phylum with sufficient evolutionary depth and ecological diversity. Evolution of signaling systems within Campylobacterota shows two opposite trends. During niche expansion, signaling complexity increases with gene expansions through horizontal gene transfer (HGT), gene duplication, fusion and fission, which create opportunities for genetic innovation and pathway integration. In contrast, as the lineages adapt to a specialized niche, complexity decreases with massive gene losses that lead to the decline or disappearance of pathways mediated by multiple transmitters. Overall, signaling capacity and complexity arise and drop together in Campylobacterota, determined by sensory demand, genetic resources and co-evolution within the genomic context. These findings reflects plausible evolutionary principles for other cellular networks and genome evolution of the Bacteria domain.
Visualization of mRNA Expression in Pseudomonas aeruginosa Aggregates Reveals Spatial Patterns of Fermentative and Denitrifying Metabolism
Authors: Livingston Jadzia , Spero Melanie A. , Lonergan Zachery R. , Newman Dianne K.
Published: March 2022

Gaining insight into the behavior of bacteria at the single cell level is important given that heterogeneous microenvironments strongly influence microbial physiology. The hybridization chain reaction (HCR) is a technique that provides in situ molecular signal amplification, enabling simultaneous mapping of multiple target RNAs at small spatial scales. To refine this method for biofilm applications, we designed and validated new probes to visualize expression of key catabolic genes in Pseudomonas aeruginosa aggregates. In addition to using existing probes for the dissimilatory nitrate reductase (narG), we developed probes for a terminal oxidase (ccoN1), nitrite reductase (nirS), nitrous oxide reductase (nosZ), and acetate kinase (ackA). These probes can be used to determine gene expression levels both in liquid culture and in biofilms. Using these probes, we quantified gene expression across oxygen gradients in aggregate populations grown using the agar block biofilm assay (ABBA). We observed distinct patterns of catabolic gene expression, with upregulation occurring in particular ABBA regions both within individual aggregates and over the aggregate population. Aerobic respiration (ccoN1) showed peak expression under oxic conditions, whereas fermentation (ackA) showed peak expression in the anoxic cores of high metabolic activity aggregates near the air-agar interface. Denitrification genes narG, nirS, and nosZ showed peak expression in hypoxic and anoxic regions, although nirS expression was much stronger in anoxic environments compared to other denitrification genes. These results reveal that the microenvironment correlates with catabolic gene expression in aggregates, and demonstrate the utility of HCR in unveiling cellular activities at the microscale in heterogeneous populations.

Importance

To understand bacteria in diverse contexts we must understand the variations in behaviors and metabolisms they express spatiotemporally. Populations of bacteria are known to be heterogeneous, but the ways this variation manifests can be challenging to characterize due to technical limitations. By focusing on energy conservation, we demonstrate that HCR v3.0 can visualize nuances in gene expression, allowing us to understand how metabolism in Pseudomonas aeruginosa biofilms responds to microenvironmental variation at high spatial resolution. We validated probes for four catabolic genes: a constitutively expressed oxidase, acetate kinase, nitrite reductase, and nitrous oxide reductase. We showed that the genes for different modes of metabolism are expressed in overlapping but distinct subpopulations according to oxygen concentrations in a predictable fashion. The spatial transcriptomic technique described here has the potential to be used to map microbial activities across diverse environments.
The Speech Envelope Following Response in Normal and Hearing Impaired Listeners
Authors: Wartenberg Tijmen , Garrett Markus , Verhulst Sarah
Published: March 2022

The aim of this work was to investigate the perceptual relevance of the frequency following response to the syllable /da/ for speech intelligibility in noise based on age and hearing deficits. Recordings of the auditory evoked potential from young normal hearing (NH) and older individuals with both normal hearing and high-frequency (HF) hearing loss were analyzed. EFR metrics obtained in quiet and noise condition were calculated and correlated with speech reception. The envelope following responses were analyzed in terms of amplitude, latency and noise robustness. The response was first simulated to form predictions on the effect of cochlear synaptopathy and outer hair cell loss on the EFR. The experimental findings were in line with the computational predictions in the found observation that the EFR was reduced as a consequence of ageing and HF hearing loss. Both the audiogram and the speech EFR magnitude fell short in the individual prediction of SRT in stationary noise, but they accounted well for group performance. We also obtained within-group EFR latency with a cross covariance matrix. Validation of the method confirmed that speech EFR latency was predictive of click ABR Wave V peak latency. Moreover, statistical analysis not only showed that the robustness of the EFR obtained in the noise condition was dependent on the degree of high-frequency hearing loss in the older NH adults, but also dependent on the EFR magnitude in the NH younger adults. These findings provide evidence towards the important role of the EFR in speech-in-noise perception.
PP2A forestalls mitophagy by dephosphorylating Parkin and ubiquitin
Authors: Ye Shang-Xiang , Liu Xing-Yu , Nie Ze-Feng , Meng Ling-Shen , Dong Xu , Li Fang-Fang , Gong Zhou , Yang Wei , Zhang Wei-Ping , Tang Chun
Published: March 2022

Mitophagy is a selective autophagic process that removes damaged mitochondria. PINK1-Parkin axis is primarily responsible for initiating mitophagy via feedforward mechanism, in which PINK1 phosphorylates ubiquitin and Parkin, and Parkin gains E3 ligase activity at the mitochondrial outer membrane. However, the phosphatase of pParkin is unknown, and the braking mechanism of mitophagy is incomplete. Here we report that protein phosphatase 2A (PP2A) catalyzes the dephosphorylation of Parkin and ubiquitin, with Cα, Aα, and B55α, the catalytic scaffolding and regulatory subunits, respectively. Up- or down-regulation of PP2A protein level in cells by over-expression or transfection of specific siRNA decreases or increases Parkin and ubiquitin phosphorylation levels, respectively. Consequently, PP2A phosphatase activity negatively modulates mitochondrial translocation of Parkin and forestalls mitophagy. Our finding thus places PP2A, an enzyme already known for its involvement in the pathogenesis of neurodegenerative diseases, further intertwined with the PINK1-Parkin signaling pathway.
Non-histone protein methylation inTrypanosoma cruziepimastigotes
Authors: de Almeida Rafael Fogaça , Rodrigues Lucena Aline Castro , Batista Michel , Marchini Fabricio Klerynton , de Godoy Lyris Martins Franco
Published: March 2022

Post-translational methylation of proteins, which occurs in arginines and lysines, modulates several biological processes at different levels of cell signaling. Recently, methylation has been demonstrated in the regulation beyond histones, for example, in the dynamics of protein-protein and protein-nucleic acid interactions. However, the presence and role of non-histone methylation inTrypanosoma cruzi, the etiologic agent of Chagas disease, has not yet been elucidated. Here, we applied mass spectrometry-based-proteomics (LC-MS/MS) to describe the methylproteome ofT. cruziepimastigotes. A total of 1253 methyl sites in 824 methylated proteins were identified, of these, 712 (86.4%) proteins were identified in two or more biological replicates. Our results of functional enrichment analysis, protein-protein interaction, and co-occurrence with other PTMs, show that protein methylation impacts different levels of biological processes, such translation, RNA and DNA binding, amino acid and carbohydrate metabolism. In addition, 171 methylated proteins were previously reported with phosphorylation sites inT. cruzi, including flagellar proteins and RNA binding proteins, indicating that there may be an interplay between these different modifications in non-histone proteins. Our results show that a broad spectrum of functions are affected by methylation indicating its potential to impact important processes in the biology of the parasite and other trypanosomes.

Statement of the significance

Trypanosoma cruziis a protozoan parasite that causes Chagas’ disease in humans and faces different environmental changes throughout its life cycle. Protein methylation is an important post-translational modification by which cells respond and adapt to the environment. We applied mass spectrometry-based proteomics and reported the first proteomic analysis of arginine and lysine methylation inT. cruzi. Our data demonstrate that protein methylation is broad and impacts different cellular processes inT. cruzi. This study represents a significant advance in the investigation of the importance of protein methylation in trypanosomes.
Dynamics of endogenous PARP1 and PARP2 during DNA damage revealed by live-cell single-molecule imaging
Authors: Mahadevan Jyothi , Jha Asmita , Rudolph Johannes , Bowerman Samuel , Narducci Domenic , Hansen Anders S , Luger Karolin
Published: March 2022

Summary/Abstract

PARP1 contributes to genome architecture and DNA damage repair through its dynamic association with chromatin. PARP1 and PARP2 (PARP1/2) recognize damaged DNA and recruit the DNA repair machinery. Using single molecule microscopy in live cells, we monitored the movement of PARP1/2 on undamaged and damaged chromatin. We identify two classes of freely diffusing PARP1/2 and two classes of bound PARP1/2. The majority (> 60%) of PARP1/2 diffuse freely in both undamaged and damaged nuclei and in the presence of inhibitors of PARP1/2 used for cancer therapy (PARPi). Laser induced DNA damage results in a small fraction of slowly diffusing PARP1 and PARP2 to become transiently bound. Treatment of cells with PARPi in the presence of DNA damage causes subtle changes in the dynamics of bound PARP1/2, in contrast to bulk studies that suggest PARP trapping. Our results imply that next-generation PARPi could specifically target the small fraction of DNA-bound PARP1/2.
Predicting the structure of large protein complexes using AlphaFold and Monte Carlo tree search
Authors: Bryant Patrick , Pozzati Gabriele , Zhu Wensi , Shenoy Aditi , Kundrotas Petras , Elofsson Arne
Published: March 2022

AlphaFold can predict the structure of single- and multiple-chain proteins with very high accuracy. However, the accuracy decreases with the number of chains, and the available GPU memory limits the size of protein complexes which can be predicted. Here we show that one can predict the structure of large complexes starting from predictions of subcomponents. We assemble 91 out of 175 complexes with 10-30 chains from predicted subcomponents using Monte Carlo tree search, with a median TM-score of 0.51. There are 30 highly accurate complexes (TM-score ≥0.8, 33% of complete assemblies). We create a scoring function, mpDockQ, that can distinguish if assemblies are complete and predict their accuracy. We find that complexes containing symmetry are accurately assembled, while asymmetrical complexes remain challenging. The method is freely available and accesible as a Colab notebook https://colab.research.google.com/github/patrickbryant1/MoLPC/blob/master/MoLPC.ipynb.
Piezo2 expressing nociceptors mediate mechanical sensitization in experimental osteoarthritis
Authors: Obeidat Alia M. , Wood Matthew J. , Ishihara Shingo , Li Jun , Wang Lai , Ren Dongjun , Bennett David A. , Miller Richard J. , Malfait Anne-Marie , Miller Rachel E.
Published: March 2022

Osteoarthritis is a very common painful joint disease, for which few treatment options exist. New non-opioid targets are needed for addressing osteoarthritis pain, which is mechanical in nature and associated with daily activities such as walking and climbing stairs. Piezo2 has been implicated in development of mechanical pain, but the mechanisms by which this occurs remain poorly understood. We observed that in two different murine models of osteoarthritis (destabilization of the medial meniscus and natural aging), nociceptor-specific Piezo2 conditional knock-out mice developed osteoarthritic joint damage, but were protected from associated mechanical sensitization. Since nerve growth factor (NGF) is known to mediate nociceptor sensitization, and antibodies that neutralize NGF are effective as a treatment for osteoarthritis pain, we explored the effects of intra-articularly injected NGF on the development of mechanical joint pain. Wild-type mice developed knee swelling and mechanical pain in response to intra-articular NGF, while nociceptor-specific Piezo2 conditional knock-out mice were protected from these effects. Single cell RNA sequencing and in situ hybridization of mouse and human lumbar dorsal root ganglia (DRG) revealed that a subset of nociceptors co-express Piezo2 and Ntrk1 (the gene that encodes the NGF receptor TrkA). These results indicate that Piezo2 plays a key role in nociceptor sensitization processes in the osteoarthritic joint, and targeting Piezo2 may represent a novel therapy for osteoarthritis pain control.

One Sentence Summary

Nociceptor sensitization to mechanical stimuli is dependent on Piezo2 in mouse models of osteoarthritis.