bluuebunny/crossref_metadata_2025 huggingface dataset:
Interviewing Women with Hidradenitis Suppurativa—Thematic and Content Analysis
Authors: Fisher Shani , Ziv Michael
Published: June 2022

OBJECTIVE

Hidradenitis suppurativa (HS) is a chronic autoinflammatory skin disease with a higher prevalence in women. The disease results in a low quality of life as well as physical and psychological comorbidities. The authors sought to determine the effects of HS on women’s self-perception and life experiences.

METHODS

Semistructured interviews were conducted with 22 women of varying age and family status. The content was transcribed and subjected to both thematic and content analyses.

RESULTS

Five themes and a number of subthemes were revealed, involving physical, emotional, coping, and functional aspects. Somatic features, especially pain, were the most troubling issues, along with the emotional burden of shame and loss of femininity and intimacy. However, women also revealed strength and expressed optimism.

CONCLUSIONS

These findings reveal the inner world of women coping with HS, addressing multiple dilemmas, problems, and concerns. Healthcare providers should pay special attention to the specific needs of these patients. Additional research is needed to further shed light on the impact of HS on women.
Exceptional response to combination ipilimumab and nivolumab in metastatic uveal melanoma: Insights from genomic analysis
Authors: Yu Irene S. , Wee Kathleen , Williamson Laura , Titmuss Emma , An Jianghong , Naderi-Azad Sheida , Metcalf Corey , Yip Stephen , Horst Basil , Jones Steven J.M. , Paton Katherine , Nelson Brad H. , Marra Marco , Laskin Janessa J. , Savage Kerry J.
Published: June 2022

Uveal melanoma is the most common intraocular malignancy and has a poor prognosis compared to other melanoma subtypes with a median overall survival of 6–10 months. With immune checkpoint inhibitor therapy, either PD-1 inhibitor alone or combination ipilimumab/nivolumab (anti-CTLA-4/anti-PD-1), responses are rare and often not durable. We present a case report of a now 66-year-old woman with diffuse metastatic uveal melanoma previously treated with a combination of ipilimumab/nivolumab, followed by maintenance nivolumab. Almost complete resolution of all sites of metastatic disease was observed except for one liver metastasis which regressed partially on immunotherapy. Notably, the patient had a significantly elevated BMI and developed widespread vitiligo on treatment. Whole-genome and transcriptome analysis was performed on the residual liver biopsy and molecular markers that may have contributed to the exceptional response were investigated. Several alterations were observed in genes involved in T-cell responses. Estimates of tumour infiltrating immune cells indicated a high level of plasma cells compared to other uveal melanoma cases, a finding previously associated with indolent disease. The patient also carried several germline SNPs that may have contributed to her treatment response as well as widespread vitiligo. Whole-genome and transcriptome sequencing have provided insight into potential molecular underpinnings of an exceptional treatment response in a tumour type typically associated with poor prognosis. Immunological findings suggest a role for plasma cells in the tumour microenvironment. Elevated BMI and the development of vitiligo may be clinically relevant factors for predicting response to immune checkpoint inhibitor therapy, warranting further studies in patients with uveal melanoma.
The association of vasomotor symptoms during the menopausal transition and cognition in later life
Authors: Peterson Amalia , Gottesman Reena T. , Miller Eliza C. , Tom Sarah E.
Published: June 2022

Objective

The majority of women experience vasomotor symptoms (VMS) during the menopausal transition. Whether self-reported VMS are associated with cognitive test performance later in life remains unclear. The goal of this study was to determine whether a greater burden of VMS is associated with poor later-life cognition.

Methods

The Wisconsin Longitudinal Study is a prospective study of randomly selected Wisconsin high school graduates of the class of 1957. At ages 65 and 72, a random subset of participants completed six cognitive tests, including similarities, letter and category fluency, immediate and delayed word recall, and digit ordering. Nested regression models were used to examine the association between extent of VMS, assessed at age 54, and baseline cognition at 65, adjusting for early-life socioeconomic status, women’s reproductive health variables, intelligence quotient, and midlife income. This series of models was also used to examine the association between VMS and change in cognition score from age 65 to 72. In sensitivity analyses, models were repeated in a sample using multiple imputation for missing covariates.

Results

Of the 5,326 women enrolled, 874 had data onVMS, covariates, and all cognitive tests. In an unadjusted model, higher VMS were associated with a lower similarities score (b = -0.09 95% CI -0.16 to -0.02) at age 65 but no other cognitive tests. In adjusted models, VMS were not related to cognition at age 65 or change in cognition. Results remained similar with multiple imputation.

Conclusions

Our study does not support a relationship between self-reported VMS and cognition later in life.
Art of prevention: Atopic dermatitis in women and families of color-prevalence, recognition, and prevention
Authors: Wilson Britney N. , Alexis Andrew , Murase Jenny E.
Published: June 2022

Atopic dermatitis (AD) is one of the most common skin conditions encountered by dermatologists. Skin of color (SOC) patients, in particular, are 50% more likely to visit a dermatologist for AD than non-SOC patients. While the misdiagnosis of AD in SOC patients is rare, the misinterpretation of severity or undertreatment of disease experienced by this patient population is a common occurrence. Herein, we present this Art of Prevention piece focused on the epidemiology, presentation, treatment, and management of AD in skin of color patients.
Drug-induced Liver Injury in Pediatrics
Authors: Monge-Urrea Fernanda , Montijo-Barrios Ericka
Published: June 2022

Drug-induced liver injury (DILI) is a rare, underdiagnosed cause of liver disease in children. The incidence of DILI in the pediatric population is unknown but it represents around 10% of all DILI cases. The most common hepatotoxic drugs in children are antibiotics and antiepileptics. DILI is classified as intrinsic or idiosyncratic and it presents mostly with 2 patterns of injury: hepatocellular or cholestatic. Diagnosis can be done with help of the Roussel Uclaf Causality Assessment Method (RUCAM) casualty assessment. The mainstay of treatment is prompt withdrawal of the suspect drug.
Radiologic Diagnosis and Staging of Bladder Cancer: An Update
Authors: Awiwi Muhammad O. , Vikram Raghunandan
Published: June 2022

Although cystoscopic biopsy is the standard of care for initial diagnosis and local staging of bladder cancer, radiologic imaging plays a major role in identifying local invasion, nodal status, distant metastasis, and posttreatment surveillance. Recent development of the Vesical Imaging-Reporting and Data System for interpretation of multiparametric magnetic resonance imaging of the bladder has expanded the role diagnostic imaging in the management of bladder cancer. This article reviews multimodality imaging appearances, staging, and differential diagnosis of bladder cancer.
Modern and prebiotic amino acids support distinct structural profiles in proteins
Authors: Tretyachenko Vyacheslav , Vymětal Jiří , Neuwirthová Tereza , Vondrášek Jiří , Fujishima Kosuke , Hlouchová Klára
Published: June 2022

The earliest proteins had to rely on amino acids available on early Earth before the biosynthetic pathways for more complex amino acids evolved. In extant proteins, a significant fraction of the ‘late’ amino acids (such as Arg, Lys, His, Cys, Trp and Tyr) belong to essential catalytic and structure-stabilizing residues. How (or if) early proteins could sustain an early biosphere has been a major puzzle. Here, we analysed two combinatorial protein libraries representing proxies of the available sequence space at two different evolutionary stages. The first is composed of the entire alphabet of 20 amino acids while the second one consists of only 10 residues (ASDGLIPTEV) representing a consensus view of plausibly available amino acids through prebiotic chemistry. We show that compact conformations resistant to proteolysis are surprisingly similarly abundant in both libraries. In addition, the early alphabet proteins are inherently more soluble and refoldable, independent of the general Hsp70 chaperone activity. By contrast, chaperones significantly increase the otherwise poor solubility of the modern alphabet proteins suggesting their coevolution with the amino acid repertoire. Our work indicates that while both early and modern amino acids are predisposed to supporting protein structure, they do so with different biophysical properties and via different mechanisms.
Real-world treatment patterns and clinical outcomes in Korean patients with AML ineligible for first-line intensive chemotherapy: A subanalysis of the CURRENT study, a non-interventional, retrospective chart review
Authors: Bang Soo-Mee , Kang Ka-Won , Song Ik-Chan , Delgado Alexander , Llamas Cynthia , Duan Yinghui , Jeong Ji-Young , Lee Je-Hwan
Published: June 2022

Background

Although most elderly patients with acute myeloid leukemia are ineligible for intensive chemotherapy, treatment options remain limited. CURRENT (UMIN000037786), a real-world, non-interventional, retrospective chart review, evaluated clinical outcomes, clinicopathologic characteristics, and treatment patterns in these patients. We present results from a subanalysis of Korean patients in this study.

Methods

Patients were aged ≥18 years with primary or secondary acute myeloid leukemia ineligible for intensive chemotherapy who initiated first-line systemic therapy or best supportive care between 2015 and 2018 across four centers in Korea. Primary endpoint was overall survival from diagnosis. Secondary endpoints included progression-free survival, time to treatment failure, and response rates. Data analyses were primarily descriptive, with time-to-event outcomes estimated using the Kaplan-Meier method, and Cox regression used to determine prognostic factors for survival.

Results

Among 194 patients enrolled, 84.0% received systemic therapy and 16.0% received best supportive care. Median age at diagnosis was 74 and 78 years, and Eastern Cooperative Oncology Group performance status 0 or 1 was reported in 73.0% and 48.4% of patients, respectively; poor cytogenetic risk was reported in 30.1% and 16.1% of patients. Median overall survival was 7.83 versus 4.50 months, and median progression-free survival was 6.73 versus 4.50 months in the systemic therapy versus best supportive care groups. Prognostic factors (all P <0.05) affecting overall survival included secondary acute myeloid leukemia (hazard ratio, 1.67 [95% CI: 1.13–2.45]), Eastern Cooperative Oncology Group performance status ≥2 (2.41 [1.51–3.83]), poor cytogenetic risk (2.10 [1.36–3.24]), and Charlson comorbidity index ≥1 (2.26 [1.43–3.58]).

Conclusion

Clinical outcomes are poor in Korean patients with acute myeloid leukemia ineligible for intensive chemotherapy who are prescribed current systemic therapies or best supportive care. There is a substantial unmet need for novel agents (monotherapy or in combination) to improve clinical outcomes in this patient population.
Denoising Approach Affects Diagnostic Differences in Brain Connectivity across the Alzheimer’s Disease Continuum
Authors: Blujus Jenna K. , Oh Hwamee ,   the Alzheimer’s Disease Neuroimaging Initiative
Published: June 2022

Graph theory provides a promising technique to investigate Alzheimer’s disease (AD)-related alterations in brain connectivity. However, discrepancies exist in the reported disruptions that occur to network topology across the AD continuum, which may be attributed to differences in the denoising approach used in fMRI processing to remove the effect of non-neuronal sources from signal. The current study aimed to determine if diagnostic differences in graph metrics were dependent on nuisance regression strategy. Sixty cognitively normal (CN), 60 MCI, and 40 AD matched for age, sex, and motion, were selected from the ADNI database for analysis. Resting state images were preprocessed using AFNI (v21.2.04) and 16 nuisance regression approaches were employed, which included the unique combination of four nuisance regressors (derivatives of the realignment parameters, motion censoring [euclidean norm > 0.3mm], outlier censoring [outlier fraction > .10], bandpass filtering [0.01 - 0.1 Hz]). Graph metrics representing network segregation (clustering coefficient, local efficiency, modularity), network integration (largest connected component, path length, local efficiency), and small-worldness (clustering coefficient/path length) were calculated. The results showed a significant interaction between diagnosis and nuisance approach on path length, such that diagnostic differences were only evident when motion derivatives and censoring of both motion and outlier volumes were applied. Further, regardless of the denoising approach, AD patients exhibited less segregated networks and lower small-worldness than CN and MCI. Finally, independent of diagnosis, denoising strategy significantly affected the magnitude of nearly all metrics (except local efficiency), such that models including bandpass filtering had higher graph metrics than those without. These findings suggest the relative robustness of network segregation and small-worldness properties to denoising strategy. However, caution should be taken when interpreting path length findings across studies, as subtle variations in regression approach may account for discrepancies. Continued efforts should be taken towards harmonizing preprocessing pipelines across studies to aid replication efforts and build consensus towards understanding the mechanisms underlying pathological aging.
BOSS: Beta-mixture Unsupervised Oligodendrocytes Segmentation System
Authors: Bae Eunchan , Gonzenbach Virgilio , Orthmann-Murphy Jennifer L , Shinohara Russell T.
Published: June 2022

To develop reparative therapies for multiple sclerosis (MS), we need to better understand the physiology of loss and replacement of oligodendrocytes, the cells that make myelin and the target of damage in MS.In vivotwo-photon fluorescence microscopy allows direct visualization of oligodendrocytes in transgenic mouse models, and promises a deeper understanding of the longitudinal dynamics of replacing oligodendrocytes after damage. However, the task of tracking oligodendrocytes requires extensive human effort and is especially challenging in three-dimensional images. While several models exist for automatically annotating cells in two-dimensional images, few models exist to annotate cells in three-dimensional images and even fewer are designed for tracking cells in longitudinal imaging. Furthermore, the complexity of processes and myelin formed by individual oligodendrocytes can result in the failure of algorithms that are specifically designed for tracking cell bodies alone. Here, we propose a novel beta-mixture unsupervised oligodendrocyte segmentation system (BOSS) that can segment and track oligodendrocytes in three-dimensional images over time that requires minimal human input. We evaluated the performance of the BOSS model on a set of eight images obtained longitudinally. We showed that the BOSS model can segment and track oligodendrocytes similarly to a blinded human observer. Our method offers many advantages, as it does not require fully curated data, reduces computational time, and most importantly recapitulates cell dynamic patterns observed by manually tracking oligodendrocytes. Although BOSS was developed to apply to our studies on oligodendrocytes, we anticipate that it can be modified to study four-dimensional in vivo data of any brain cell.
Quantitative whole-tissue 3D imaging reveals bacteria in close association with mouse jejunum mucosa
Authors: Poceviciute Roberta , Bogatyrev Said R. , Romano Anna E. , Dilmore Amanda H. , Mondragón-Palomino Octavio , Takko Heli , Ismagilov Rustem F.
Published: June 2022

Background

The small intestine (SI) is the primary site of nutrient absorption, so its large surface area lacks the thick protective mucus that is characteristic of the large intestine. Because the SI epithelium is relatively exposed, any microbes that colonize the thin mucosa of the SI may exert a substantial effect on the host. Thus far, potential bacterial colonization of the SI mucosa has only been documented in disease states, suggesting mucosal colonization is a rare occurrence, likely requiring multiple perturbations.

Results

Here, we tested whether we could induce bacterial association with jejunum mucosa by a combination of malnutrition and oral co-gavage with a specific bacterial cocktail (E. coli and Bacteroides spp.) that has previously induced environmental enteropathy in mouse models. To overcome the current limitations in imaging and allow definite determination of whether bacterial colonization of the SI mucosa is occurring, we optimized our previously developed whole-tissue three-dimensional (3D) imaging tools with third-generation hybridization chain reaction (HCR v3.0) probes. Only in mice that were malnourished and gavaged with the bacterial cocktail did we detect dense bacterial clusters surrounding intestinal villi suggestive of colonization. Healthy mice gavaged with bacteria and malnourished mice not gavaged with bacteria showed no evidence of mucosal colonization. Furthermore, in malnourished mice gavaged with bacteria we detected villus loss, which may represent one possible consequence that bacterial colonization of the SI mucosa has on the host.

Conclusions

Our results suggest that dense bacterial colonization of jejunum mucosa is possible in the presence of multiple perturbations and that villus loss may be one possible consequence to such colonization. Furthermore, our results demonstrate the utility of whole-tissue 3D imaging tools. Although 2D imaging of thin sections may have failed to detect and capture the full spatial complexity of such rare events, whole-tissue 3D imaging tools enabled their detection over large areas of intestinal mucosa and visualization of their spatial complexity in 3D.
Lactose-based surfactants complexable with oligonucleic acids as gene delivery agents
Authors: Wilkowska Michalina , Makrocka-Rydzyk Monika , Michocka Katarzyna , Zieliński Ryszard , Sobczak Krzysztof , Kozak Maciej
Published: June 2022

Advances in molecular medicine related to the treatment of genetic disorders and cancer require finding new pathways for gene transfer. Nonviral delivery methods rely on the application of polymers, liposomes and cationic lipid systems used as vehicle. Among these systems, there is increasing interest in surfactants, which, due to their ability to complex with nucleic acids can deliver into cell DNA or RNA molecules of almost any size, which is unattainable with viral gene delivery systems.The main aim of this study was to determine the effect of the concentration of lactose-based surfactants (zwitterionic derivatives of sulfobetaine with carbohydrate moieties) on the structure of DNA/RNA as well as to explore their abilities of nucleic acid complexation. Structural studies of DNA or RNA in complexes with surfactants of two types at various concentrations were conducted using circular dichroism (CD) spectroscopy, gel electrophoresis (GE) and synchrotron radiation small angle X-ray scattering (SR-SAXS). Our studies showed that the examined surfactants have excellent properties of forming complexes with DNA and RNA. Additionally, to determine the cytotoxicity and transfection abilities of the studied lipoplexes, preliminary tests were performed in HeLa and fibroblast cells. The obtained results suggest that these systems have relatively low toxicity; however, further research is needed in this area.
Deciphering inhibitory mechanism of coronavirus replication through host miRNAs-RNA-dependent RNA polymerase (RdRp) interactome
Authors: Morenikeji Olanrewaju B. , Adegbaju Muyiwa S. , Okoh Olayinka S. , Babalola Asegunloluwa E. , Grytsay Anastasia , Braimah Olubumi A. , Akinyemi Mabel O. , Thomas Bolaji N.
Published: June 2022

Despite what we know so far, Covid-19, caused by SARS-CoV-2 virus, remains a pandemic that still require urgent healthcare intervention. The frequent mutations of the SARS-CoV-2 virus has rendered disease control with vaccines and antiviral drugs quite difficult and challenging, with newer variants surfacing constantly. There is therefore the need for newer, effective and efficacious drugs against coronaviruses. Considering the role of RNA dependent, RNA polymerase (RdRp) as an important enzyme necessary for the virus life cycle and its conservation among coronaviruses, we investigated potential host miRNAs that can be employed as broad-range antiviral drugs averse to coronaviruses, with particular emphasis on BCoV, MERS-CoV, SARS-CoV and SARS-CoV-2. miRNAs are small molecules capable of binding mRNA and regulate expression at transcriptional or translational levels. Our hypothesis is that host miRNAs have the potential of blocking coronavirus replication through miRNA-RdRp mRNA interaction. To investigate this, we downloaded the open reading frame (ORF 1ab) nucleotide sequences and used them to interrogate miRNA databases for miRNAs that can bind them. We employed various bioinformatics tools to predict and identify the most effective host miRNAs. In all, we found 27 miRNAs that target RdRp mRNA of multiple coronaviruses, of which three - hsa-miR-1283, hsa-miR-579-3p, and hsa-miR-664b-3p target BCoV, SARS-CoV and SARS-CoV-2. Additionally, hsa-miR-374a-5p has three bovine miRNAs homologs viz bta-miR-374a, bta-miR-374b, and bta-miR-374c. Inhibiting the expression of RdRp enzyme via non-coding RNA is novel and of great therapeutic importance in the control of coronavirus replication, and could serve as a broad-spectrum antiviral, with hsa-miR-1283, hsa-miR-579-3p, and hsa-miR-664b-3p highly promising.
Auditory neural tracking reflects target enhancement but not distractor suppression in a psychophysically augmented continuous-speech paradigm
Authors: Orf Martin , Wöstmann Malte , Hannemann Ronny , Obleser Jonas
Published: June 2022

Selective attention modulates the neural tracking of speech in auditory cortical regions. It is unclear whether this attention modulation is dominated by enhanced target tracking, or suppression of distraction. To settle this long-standing debate, we here employed an augmented electroencephalography (EEG) speech-tracking paradigm with target, distractor, and neutral streams. Concurrent target speech and distractor (i.e., sometimes relevant) speech were juxtaposed with a third, never task-relevant speech stream serving as neutral baseline. Listeners had to detect short target repeats and committed more false alarms originating from the distractor than the neutral stream. Speech tracking revealed target enhancement but no distractor suppression below the neutral baseline. Speech tracking of the target (not distractor or neutral speech) explained single-trial accuracy in repeat detection. In sum, the enhanced neural representation of target speech is specific to processes of attentional gain for behaviourally relevant target speech rather than neural suppression of distraction.
A FRET biosensor, SMART, monitors necroptosis in renal tubular epithelial cells in a cisplatin-induced kidney injury model
Authors: Murai Shin , Takakura Kanako , Sumiyama Kenta , Moriwaki Kenta , Terai Kenta , Komazawa-Sakon Sachiko , Yamaguchi Yoshifumi , Mikami Tetuo , Araki Kimi , Ohmuraya Masaki , Matsuda Michiyuki , Nakano Hiroyasu
Published: June 2022

Necroptosis is a regulated form of cell death involved in various pathological conditions, including ischemic reperfusion injuries, virus infections, and drug-induced tissue injuries. However, it is not fully understood when and where necroptosis occursin vivo. We previously generated a Forster resonance energy transfer (FRET) biosensor, termed SMART (the sensor for MLKL activation based on FRET), which specifically monitored necroptosis in human and murine cell linesin vitro. Here, we generated transgenic (Tg) mice that expressed the SMART biosensor in various tissues. SMART monitored necroptosis, but not apoptosis or pyroptosis, in primary cells, including peritoneal macrophages and embryonic fibroblasts. Moreover, the FRET signal was elevated in renal tubular cells of cisplatin-treated SMART Tg mice compared to untreated SMART Tg mice. Together, SMART Tg mice may provide a valuable tool for monitoring necroptosis in different types of cellsin vitroandin vivo.
Alterations of auditory sensory gating in mice with noise-induced tinnitus treated with nicotine and cannabis extract
Authors: Ciralli Barbara , Malfatti Thawann , Lima Thiago Z. , Silva Sérgio Ruschi B. , Cederroth Christopher R. , Leao Katarina E.
Published: June 2022

Tinnitus is a phantom sound perception affecting both auditory and limbic structures. The mechanisms of tinnitus remain unclear and it is debatable whether tinnitus alters attention to sound and the ability to inhibit repetitive sounds, a phenomenon also known as auditory gating. Here we investigate if noise exposure interferes with auditory gating and whether natural extracts of cannabis or nicotine could improve auditory pre-attentional processing in noise-exposed mice. We used 22 male C57BL/6J mice divided into noise-exposed (exposed to a 9-11 kHz narrow band noise for 1 hour) and sham (no sound during noise exposure) groups. Hearing thresholds were measured using auditory brainstem responses, and tinnitus-like behavior was assessed using Gap prepulse inhibition of acoustic startle. After noise exposure, mice were implanted with multi-electrodes in the dorsal hippocampus to assess auditory event-related potentials in response to paired clicks. The results showed that mice with tinnitus-like behavior displayed auditory gating of repetitive clicks, but with larger amplitudes and longer latencies of the N40 component of the aERP waveform. The combination of cannabis extract and nicotine improved auditory gating ratio in noise-exposed mice without permanent hearing threshold shifts. Lastly, the longer latency of the N40 component appears due to an increased sensitivity to cannabis extract in noise-exposed mice compared to sham mice. The study suggests that the altered central plasticity in tinnitus is more sensitive to the combined actions on the cholinergic and the endocannabinoid systems. Overall, the findings contribute to a better understanding of pharmacological modulation of auditory sensory gating.
PI3P dependent regulation of cell size and autophagy by phosphatidylinositol 5-phosphate 4-kinase
Authors: Ghosh Avishek , Venugopal Aishwarya , Shinde Dhananjay , Sharma Sanjeev , Krishnan Meera , Mathre Swarna , Krishnan Harini , Raghu Padinjat
Published: June 2022

Phosphatidylinositol 3-phosphate (PI3P) and Phosphatidylinositol 5-phosphate (PI5P) are low abundant phosphoinositides crucial for key cellular events such as endosomal trafficking and autophagy. Phosphatidylinositol 5-phosphate 4-kinase (PIP4K) is an enzyme that regulates PI5P in vivo but can act on both PI5P and PI3P, in vitro. In this study, we report a novel role for PIP4K in regulating PI3P levels in Drosophila tissues. Loss-of-function mutants of the only PIP4K gene in Drosophila (dPIP4K29) show reduced cell size in larval salivary glands. We find that PI3P levels are elevated in dPIP4K29 tissues and that reverting PI3P levels back towards wild type, without changes in PI5P levels, can also rescue the reduced cell size phenotype. dPIP4K29 mutants also show an upregulation in autophagy and the reduced cell size can be reverted by decreasing Atg8a, that is required for autophagosome maturation. Lastly, increasing PI3P levels in wild type salivary glands can phenocopy the reduction in cell size and associated upregulation of autophagy seen in dPIP4K29. Thus, our study reports for the first time, a role for a PIP4K-regulated PI3P pool in the control of autophagy and cell size regulation that may explain the reported role of PIP4K in regulating neurodegeneration and tumour growth.
Antisense oligonucleotide therapy rescues disturbed brain rhythms and sleep in juvenile and adult mouse models of Angelman syndrome
Authors: Lee Dongwon , Chen Wu , Kaku Heet Naresh , Zhuo Xinming , Chao Eugene S. , Soriano Armand , Kuncheria Allen , Flores Stephanie , Kim Joo Hyun , Rigo Frank , Jafar-nejad Paymaan , Beaudet Arthur L. , Caudill Matthew S. , Xue Mingshan
Published: June 2022

UBE3A encodes ubiquitin protein ligase E3A, and in neurons its expression from the paternal allele is repressed by the UBE3A antisense transcript (UBE3A-ATS). This leaves neurons susceptible to loss-of-function of maternal UBE3A. Indeed, Angelman syndrome, a severe neurodevelopmental disorder, is caused by maternal UBE3A deficiency. A promising therapeutic approach to treating Angelman syndrome is to reactivate the intact paternal UBE3A by suppressing UBE3A-ATS. Prior studies show that many neurological phenotypes of maternal Ube3a knockout mice can only be rescued by reinstating Ube3a expression in early development, indicating a restricted therapeutic window for Angelman syndrome. Here we report that reducing Ube3a-ATS by antisense oligonucleotides in juvenile or adult maternal Ube3a knockout mice rescues the abnormal electroencephalogram rhythms and sleep disturbance, two prominent clinical features of Angelman syndrome. Importantly, the degree of phenotypic improvement correlates with the increase of Ube3a protein levels. These results indicate that the therapeutic window of genetic therapies for Angelman syndrome is broader than previously thought, and electroencephalogram power spectrum and sleep architecture should be used to evaluate the clinical efficacy of therapies.
Pumping the brakes on RNA velocity – understanding and interpreting RNA velocity estimates
Authors: Zheng Shijie C. , Stein-O’Brien Genevieve , Boukas Leandros , Goff Loyal A. , Hansen Kasper D.
Published: June 2022

RNA velocity analysis of single cells promises to predict temporal dynamics from gene expression. Indeed, in many systems, it has been observed that RNA velocity produces a vector field that qualitatively reflects known features of the system. Despite this observation, the limitations of RNA velocity estimates are poorly understood. Using real data and simulations, we dissect the impact of different steps in the RNA velocity workflow on the estimated vector field. We find that the process of mapping RNA velocity estimates into a low-dimensional representation, such as those produced by UMAP, has a large impact on the result. The RNA velocity vector field strongly depends on the k-NN graph of the data. This dependence leads to significant estimator errors when the k-NN graph is not a faithful representation of the true data structure, a feature that cannot be known for most real datasets. Finally, we establish that RNA velocity estimates expression speed neither at the gene nor cellular level. We propose that RNA velocity is best considered a smoothed interpolation of the observed k-NN structure, as opposed to an extrapolation of future cellular states, and that the use of RNA velocity as a validation of latent space embedding structures is circular.
Strong pathogen competition in neonatal gut colonisation
Authors: Mäklin Tommi , Thorpe Harry A. , Pöntinen Anna K. , Gladstone Rebecca A. , Shao Yan , Pesonen Maiju , McNally Alan , Johnsen Pål J. , Samuelsen Ørjan , Lawley Trevor D. , Honkela Antti , Corander Jukka
Published: June 2022

Bacterial pathogen species and their strains that colonise the human gut are generally understood to compete against both each other and the commensal species colonising this ecosystem. However, currently we are lacking a population-wide quantification of strain-level colonisation dynamics for many common bacterial pathogens and the relationship of colonisation potential to prevalence in disease is unknown. In addition, it is unclear how ecological factors might be modulating the dynamics. Here, using a combination of latest high-resolution metagenomics and strain-level genomic epidemiology methods leveraging large genomic reference libraries of key pathogens, we performed a characterisation of the competition (co-exclusion and co-presence) and colonisation dynamics for a longitudinal cohort of neonatal gut microbiomes. We found a strong inter- and intra-species competition dynamic in the gut colonisation process, but also a number of synergistic relationships among several species belonging to genusKlebsiella, which includes the prominent human pathogenKlebsiella pneumoniae. Additionally, we find no evidence of preferential colonisation by hospital-adapted pathogen lineages in either vaginal or caesarean section birth groups. Our analysis also enables the first unbiased assessment of the strain-level colonisation potential of extra-intestinal pathogenicEscherichia coli(ExPEC) in comparison with their potential to cause bloodstream infections. We determined that the established common ExPEC clones ST73 and ST95 are overall significantly more pathogenic than the more recent, globally circulating multi-drug resistant clone ST131, where only a single subclone (ST131-C2) exhibited excess pathogenic potential. Our study highlights the importance of systematic surveillance of bacterial gut pathogens, not only from disease but also from carriage state, to better inform therapies and preventive medicine in the future.
Dynein dysfunction prevents maintenance of high concentrations of slow axonal transport cargos at the axon terminal: a computational study
Authors: Kuznetsov Ivan A. , Kuznetsov Andrey V.
Published: June 2022

Here we report computational studies of bidirectional transport in an axon, specifically focusing on predictions when the retrograde motor becomes dysfunctional. We are motivated by reports that mutations in dynein-encoding genes can cause diseases associated with peripheral motor and sensory neurons, such as type 2O Charcot-Marie-Tooth disease. We use two different models to simulate bidirectional transport in an axon: an anterograde-retrograde model, which neglects passive transport by diffusion in the cytosol, and a full slow transport model, which includes passive transport by diffusion in the cytosol. As dynein is a retrograde motor, dysfunction should not directly influence anterograde transport. However, our modeling results unexpectedly predict that slow axonal transport fails to transport cargos against their concentration gradient without dynein. The reason is the lack of a physical mechanism for the reverse information flow from the axon terminal, which is required so that the cargo concentration at the terminal could influence the cargo concentration distribution in the axon. Mathematically speaking, to achieve a prescribed concentration at the terminal, equations governing cargo transport must allow for the imposition of a boundary condition postulating the cargo concentration at the terminal. Perturbation analysis for the case when the retrograde motor velocity becomes close to zero predicts uniform cargo distributions along the axon. The obtained results explain why slow axonal transport must be bidirectional to allow for the maintenance of concentration gradients along the axon length. Our result is limited to small cargo diffusivity, which is a reasonable assumption for many slow axonal transport cargos (such as cytosolic and cytoskeletal proteins, neurofilaments, actin, and microtubules) which are transported as large multiprotein complexes or polymers.
Flexible and efficient handling of nanopore sequencing signal data with slow5tools
Authors: Samarakoon Hiruna , Ferguson James M. , Jenner Sasha P. , Amos Timothy G. , Parameswaran Sri , Gamaarachchi Hasindu , Deveson Ira W.
Published: June 2022

Background

Nanopore sequencing is an emerging technology that is being rapidly adopted in research and clinical genomics. We recently developed SLOW5, a new file format for storage and analysis of raw data from nanopore sequencing experiments. SLOW5 is a community-centric, open source format that offers considerable performance benefits over the existing nanopore data format, known as FAST5. Here we introduce slow5tools, a simple, intuitive toolkit for handling nanopore raw signal data in SLOW5 format.

Results

Slow5tools enables lossless FAST5-to-SLOW5 and SLOW5-to-FAST5 data conversion, and a range of tools for structuring, indexing, viewing and querying SLOW5 files. Slow5tools uses multi-threading, multi-processing and other engineering strategies to achieve fast data conversion and manipulation, including live FAST5-to-SLOW5 conversion during sequencing. We outline a series of examples and benchmarking experiments to illustrate slow5tools usage, and describe the engineering principles underpinning its high performance.

Conclusion

Slow5tools is an essential toolkit for handling nanopore signal data, which was developed to support adoption of SLOW5 by the nanopore community. Slow5tools is written in C/C++ with minimal dependencies and is freely available as an open-source program under an MIT licence: https://github.com/hasindu2008/slow5tools.
One model fits all: combining inference and simulation of gene regulatory networks
Authors: Ventre Elias , Herbach Ulysse , Espinasse Thibault , Benoit Gérard , Gandrillon Olivier
Published: June 2022

The rise of single-cell data highlights the need for a nondeterministic view of gene expression, while offering new opportunities regarding gene regulatory network inference. We recently introduced two strategies that specifically exploit time-course data, where single-cell profiling is performed after a stimulus: HARISSA, a mechanistic network model with a highly efficient simulation procedure, and CARDAMOM, a scalable inference method seen as model calibration. Here, we combine the two approaches and show that the same model driven by transcriptional bursting can be used simultaneously as an inference tool, to reconstruct biologically relevant networks, and as a simulation tool, to generate realistic transcriptional profiles emerging from gene interactions. We verify that CARDAMOM quantitatively reconstructs causal links when the data is simulated from HARISSA, and demonstrate its performance on experimental data collected onin vitrodifferentiating mouse embryonic stem cells. Overall, this integrated strategy largely overcomes the limitations of disconnected inference and simulation.

Author summary

Gene regulatory network (GRN) inference is an old problem, to which single-cell data has recently offered new challenges and breakthrough potential. Many GRN inference methods based on single-cell transcriptomic data have been developed over the last few years, while GRN simulation tools have also been proposed for generating synthetic datasets with realistic features. However, except for benchmarking purposes, these two fields remain largely disconnected. In this work, building on a combination of two methods we recently described, we show that a particular GRN model can be used simultaneously as an inference tool, to reconstruct a biologically relevant network from time-course single-cell gene expression data, and as a simulation tool, to generate realistic transcriptional profiles in a non-trivial way through gene interactions. This integrated strategy demonstrates the benefits of using the same executable model for both simulation and inference.
Structural basis of LRPPRC-SLIRP-dependent translation by the mitoribosome
Authors: Singh Vivek , Conor Moran J. , Itoh Yuzuru , Soto Iliana C. , Fontanesi Flavia , Couvillion Mary , Huynen Martijn A. , Churchman Stirling , Barrientos Antoni , Amunts Alexey
Published: June 2022

In mammalian mitochondria, mRNAs are co-transcriptionally stabilized by the protein factor LRPPRC. Here, we characterize LRPPRC as an mRNA delivery factor and report its cryo-EM structure in complex with SLIRP, mRNA and the mitoribosome. The structure shows that LRPPRC associates with the mitoribosomal proteins mS39 and the N-terminus of mS31 through recognition of the LRPPRC helical repeats. Together, the proteins form a corridor for hand-off the mRNA. The mRNA is directly bound to SLIRP, which also has a stabilizing function for LRPPRC. To delineate the effect of LRPPRC on individual mitochondrial transcripts, we used an RNAseq approach, metabolic labeling and mitoribosome profiling that showed a major influence on ND1, ND2, ATP6, COX1, COX2, and COX3 mRNA translation efficiency. Our data suggest that LRPPRC-SLIRP acts in recruitment of mitochondrial mRNAs to modulate their translation. Collectively, the data define LRPPRC-SLIRP as a regulator of the mitochondrial gene expression system.
Molecular interactions underlying the phase separation of HP1α: Role of phosphorylation, ligand and nucleic acid binding
Authors: Her Cheenou , Phan Tien M. , Jovic Nina , Kapoor Utkarsh , Ackermann Bryce E. , Rizuan Azamat , Kim Young , Mittal Jeetain , Debelouchina Galia T.
Published: June 2022

Heterochromatin protein 1α (HP1α) is a crucial component for the proper maintenance of chromatin structure and function. It has been proposed that HP1α functions through liquid-liquid phase separation (LLPS), which allows it to sequester and compact chromatin into transcriptionally repressed heterochromatin regions. In vitro, HP1α can form phase separated liquid droplets upon phosphorylation of its N-terminus extension (NTE) and/or through interactions with DNA and chromatin. While it is known that LLPS requires homodimerization of HP1α and that it involves interactions between the positively charged hinge region of HP1α and the negatively charged phosphorylated NTE or nucleic acid, the precise molecular details of this process and its regulation are still unclear. Here, we combine computational modeling and experimental approaches to elucidate the phase separation properties of HP1α under phosphorylation-driven and DNA-driven LLPS conditions. We also tune these properties using peptides from four HP1α binding partners (Sgo1, CAF-1, LBR, and H3). In phosphorylation-driven LLPS, HP1α can exchange intradimer hinge-NTE interactions with interdimer contacts, which also leads to a structural change from a compacted to an extended HP1α dimer conformation. This process can be enhanced by the presence of positively charged peptide ligands such as Sgo1 and H3 and disrupted by the addition of negatively charged or neutral peptides such as LBR and CAF-1. In DNA-driven LLPS, both positively and negatively charged peptide ligands can perturb phase separation. Our findings demonstrate the importance of electrostatic interactions in the LLPS of HP1α where binding partners can modulate the overall charge of the droplets and screen or enhance hinge region interactions through specific and non-specific effects. Our study illuminates the complex molecular framework that can fine tune the properties of HP1α and that can contribute to heterochromatin regulation and function.
Components of TOR and MAP kinase signaling control chemotropism and pathogenicity in the fungal pathogen Verticillium dahliae
Authors: Vangalis Vasileios , Markakis Emmanouil A. , Knop Michael , Pietro Antonio Di , Typas Milton A. , Papaioannou Ioannis A.
Published: June 2022

Filamentous fungi can sense useful resources and hazards in their environment and direct growth of their hyphae accordingly. Chemotropism ensures access to nutrients, contact with other individuals (e.g., for mating), and interaction with hosts in the case of pathogens. Previous studies have revealed a complex chemotropic sensing landscape during host-pathogen interactions, but the underlying molecular machinery remains poorly characterized. Here we studied mechanisms controlling directed hyphal growth of the important plant-pathogenic fungus Verticillium dahliae towards different chemoattractants. We found that the homologs of the Rag GTPase Gtr1 and the GTPase-activating protein Tsc2, an activator and a repressor of the TOR kinase respectively, play important roles in hyphal chemotropism towards nutrients, plant-derived signals, and heterologous α-pheromone of Fusarium oxysporum. Furthermore, important roles of these regulators were identified in fungal development and pathogenicity. We also found that the mitogen-activated protein kinase (MAPK) Fus3 is required for chemotropism towards nutrients, while the G protein-coupled receptor (GPCR) Ste2 and the MAPK Slt2 control chemosensing of plant-derived signals and α-pheromone. Our study establishes V. dahliae as a suitable model for the analysis of fungal chemotropism and discovers new components of chemotropic signaling, during growth and host-pathogen interactions of V. dahliae.
Precise Transcript Targeting by CRISPR-Csm Complexes
Authors: Colognori David , Trinidad Marena , Doudna Jennifer A.
Published: June 2022

Robust and precise transcript targeting in mammalian cells remains a difficult challenge using existing approaches due to inefficiency, imprecision, and subcellular compartmentalization. Here, we show that the CRISPR-Csm complex, a multi-protein effector from type III CRISPR immune systems in prokaryotes, provides surgical RNA ablation of both nuclear and cytoplasmic transcripts. As part of the most widely occurring CRISPR adaptive immunity pathway, CRISPR-Csm uses a programmable RNA-guided mechanism to find and degrade target RNA molecules without inducing indiscriminate trans-cleavage of cellular RNAs, giving it an important advantage over the CRISPR-Cas13-family enzymes. Using single-vector delivery of the S. thermophilus Csm complex, we observe high-efficiency RNA knockdown (90-99%) and minimal off-target effects in human cells, outperforming existing technologies including shRNA- and Cas13-mediated knockdown. We also find that catalytically inactivated Csm achieves specific and durable RNA binding, a property we harness for live-cell RNA imaging. These results establish the feasibility and efficacy of multi-protein CRISPR-Cas effector complexes as RNA-targeting tools in eukaryotes.
ct2vl: Converting Ct Values to Viral Loads for SARS-CoV-2 RT-qPCR Test Results
Authors: Hill Elliot D. , Yilmaz Fazilet , Callahan Cody , Cheng Annie , Braun Jasper , Arnaout Ramy
Published: June 2022

RT-qPCR is the de facto reference method for detecting the presence of SARS-CoV-2 genomic material in infected individuals (1). Although RT-qPCR is inherently quantitative and despite SARS-CoV-2 viral loads varying by 10 orders of magnitude and therefore being potentially highly clinically informative, in practice SARS-CoV-2 RT-qPCR results are usually reported qualitatively as simply positive or negative. This is both because of the mathematical complexity of converting from Ct values to viral loads and because the same Ct value can correspond to orders-of-magnitude differences in viral load depending on the testing platform (2, 3, 4). To address this problem, here we present ct2vl, a Python package designed to help individual clinical laboratories, investigators, and test developers convert from Ct values to viral loads on their own platforms, using only the data generated during validation of those platforms. It allows any user to convert Ct values to viral loads and is readily applicable to other RT-qPCR tests. ct2vl is open source, has 100% code coverage, and is freely available via the Python Package Index (PyPI).

IMPORTANCE

Up to now, COVID-19 test results have been reported as positive vs. negative, even though “positive” can mean anywhere from 1 copy of SARS-CoV-2 virus per milliliter of transport media to over 1 billion copies/mL, with attendant clinical consequences. Democratizing access to this quantitative data is the first step toward its eventual incorporation into test development, the research literature, and clinical care.