bluuebunny/crossref_metadata_2025 huggingface dataset:
Drosophila SUMM4 complex couples insulator function and DNA replication timing control
Authors: Andreyeva Evgeniya N. , Emelyanov Alexander V. , Nevil Markus , Sun Lu , Vershilova Elena , Hill Christina A. , Keogh Michael-C. , Duronio Robert J. , Skoultchi Arthur I. , Fyodorov Dmitry V.
Published: October 2021

Abstract

Asynchronous replication of chromosome domains during S phase is essential for eukaryotic genome function, but the mechanisms establishing which domains replicate early versus late in different cell types remain incompletely understood. Drosophila SNF2-related factor SUUR imparts under- replication of late-replicating intercalary heterochromatin in polytene chromosomes. SUUR negatively regulates DNA replication fork progression; however, its mechanism of action remains obscure. Here we developed a novel method termed MS-Enabled Rapid protein Complex Identification (MERCI) to isolate a stable stoichiometric native complex SUMM4 that comprises SUUR and a chromatin boundary protein Mod(Mdg4)-67.2. Mod(Mdg4) stimulates SUUR ATPase activity and is required for a normal spatiotemporal distribution of SUUR in vivo. SUUR and Mod(Mdg4)-67.2 together mediate the activities of gypsy insulator that prevent certain enhancer-promoter interactions and establish euchromatin-heterochromatin barriers in the genome. Furthermore, SuUR or mod(mdg4) mutations reverse under-replication of intercalary heterochromatin. Thus, SUMM4 can impart late replication of intercalary heterochromatin by attenuating the progression of replication forks through euchromatin/heterochromatin boundaries. Our findings reveal that DNA replication can be delayed by a chromatin barrier and uncover a critical role for architectural proteins in replication control. They suggest a mechanism for replication timing that does not depend on an asynchronous firing of replication origins.
RABENOSYN separation-of-function mutations uncouple endosomal recycling from lysosomal degradation causing a distinct Mendelian Disorder
Authors: Paul Franziska , Ng Calista , Nafissi Shahriar , Nilipoor Yalda , Tavasoli Ali Reza , Mohamad Sahari Umar Bin , Bonnard Carine , Wong Pui-Mun , Nabavizadeh Nasrinsadat , Estiar Mehrdad A. , Majoie Charles B. , Lee Hane , Nelson Stanley F. , Gan-Or Ziv , Rouleau Guy A. , Van Veldhoven Paul P. , Massie Rami , Hennekam Raoul C. , Kariminejad Ariana , Reversade Bruno
Published: October 2021

Rabenosyn (RBSN) is a conserved endosomal protein necessary for regulating internalized cargo. Here, we present genetic, cellular and biochemical evidence that two distinct RBSN missense variants are responsible for a novel Mendelian disorder consisting of progressive muscle weakness, facial dysmorphisms, ophthalmoplegia and intellectual disability. Using exome sequencing, we identified recessively-acting germline alleles p.Arg180Gly and p.Gly183Arg which are both situated in the FYVE domain of RBSN. We find that these variants abrogate binding to its cognate substrate PI3P and thus prevent its translocation to early endosomes. Although the endosomal recycling pathway was unaltered, mutant p.Gly183Arg patient fibroblasts exhibit accumulation of cargo tagged for lysosomal degradation. Our results suggest that these variants are separation-of-function alleles, which cause a delay in endosomal maturation without affecting cargo recycling. We conclude that distinct germline mutations in RBSN cause non-overlapping phenotypes with specific and discrete endolysosomal cellular defects.
Humoral immune responses to COVID-19 vaccination in people living with HIV receiving suppressive antiretroviral therapy
Authors: Brumme Zabrina L. , Mwimanzi Francis , Lapointe Hope R. , Cheung Peter , Sang Yurou , Duncan Maggie C. , Yaseen Fatima , Agafitei Olga , Ennis Siobhan , Ng Kurtis , Basra Simran , Lim Li Yi , Kalikawe Rebecca , Speckmaier Sarah , Moran-Garcia Nadia , Young Landon , Ali Hesham , Ganase Bruce , Umviligihozo Gisele , Omondi F. Harrison , Atkinson Kieran , Sudderuddin Hanwei , Toy Junine , Sereda Paul , Burns Laura , Costiniuk Cecilia T. , Cooper Curtis , Anis Aslam H. , Leung Victor , Holmes Daniel , DeMarco Mari L. , Simons Janet , Hedgcock Malcolm , Romney Marc G. , Barrios Rolando , Guillemi Silvia , Brumme Chanson J. , Pantophlet Ralph , Montaner Julio S.G. , Niikura Masahiro , Harris Marianne , Hull Mark , Brockman Mark A.
Published: October 2021

Humoral responses to COVID-19 vaccines in people living with HIV (PLWH) remain incompletely understood. We measured circulating antibodies against the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein, ACE2 displacement and live viral neutralization activities one month following the first and second COVID-19 vaccine doses in 100 adult PLWH and 152 controls. All PLWH were receiving suppressive antiretroviral therapy, with median CD4+ T-cell counts of 710 (IQR 525-935) cells/mm3. Nadir CD4+ T-cell counts ranged as low as <10 (median 280; IQR 120-490) cells/mm3. After adjustment for sociodemographic, health and vaccine-related variables, HIV infection was significantly associated with 0.2 log10 lower anti-RBD antibody concentrations (p=0.03) and ∼11% lower ACE2 displacement activity (p=0.02), but not lower viral neutralization (p=0.1) after one vaccine dose. Following two doses however, HIV was no longer significantly associated with the magnitude of any response measured. Rather, older age, a higher burden of chronic health conditions, and having received two ChAdOx1 doses (versus a heterologous or dual mRNA vaccine regimen) were independently associated with lower responses. After two vaccine doses, no significant correlation was observed between the most recent or nadir CD4+ T-cell counts and vaccine responses in PLWH. These results suggest that PLWH with well-controlled viral loads on antiretroviral therapy and CD4+ T-cell counts in a healthy range will generally not require a third COVID-19 vaccine dose as part of their initial immunization series, though other factors such as older age, co-morbidities, vaccine regimen type, and durability of vaccine responses will influence when this group may benefit from additional doses. Further studies of PLWH who are not receiving antiretroviral treatment and/or who have low CD4+ T-cell counts are needed.
A mechanistic mathematical model of initiation and malignant transformation in sporadic vestibular schwannoma
Authors: Paterson Chay , Bozic Ivana , Smith Miriam J. , Hoad Xanthe , Evans D. Gareth R.
Published: October 2021

A vestibular schwannoma (VS) is a relatively rare, benign tumour of the eighth cranial nerve, often involving alterations to the gene NF2. Previous mathematical models of schwannoma incidence have not attempted to account for alterations in specific genes, and could not distinguish between point mutations and loss of heterozygosity (LOH). Here, we present a mechanistic approach to modelling initiation and malignant transformation in Schwannoma. Each parameter is associated with a specific gene or mechanism operative in Schwann cells, and can be determined by combining incidence data with empirical frequencies of pathogenic variants and LOH. This results in new estimates for the base pair mutation rate u = 4.48 × 10−10 and the rate of LOH = 2.03 × 10−6/yr in Schwann cells. In addition to new parameter estimates, we extend the approach to estimate the risk of both spontaneous and radiation-induced malignant transformation. We conclude that radiotherapy is likely to have a negligible excess risk of malignancy for sporadic VS, with a possible exception of rapidly growing tumours.
New insights into the human 26S proteasome function and regulation
Authors: Kišonaitė Miglė , Afanasyev Pavel , Tafilaku Jonida , Rêgo Ana Toste , da Fonseca Paula C. A.
Published: October 2021

The 26S proteasome is a protease complex essential for proteostasis and strict regulation of diverse critical physiological processes, the mechanisms of which are still not fully described. The human 26S proteasome purification was optimized without exogenous nucleotides, to preserve the endogenous nucleotide occupancy and conformation of its AAA-ATPase subunits. This unveiled important effects on the proteasome function and structure resulting from exposure to Ca2+ or Mg2+, with important physiological implications. This sample, with an added model degron designed to mimic the minimum canonical ubiquitin signal for proteasomal recognition, was analysed by high-resolution cryo-EM. Two proteasome conformations were resolved, with only one capable of degron binding. The structural data show that this occurs without major conformation rearrangements and allows to infer into the allosteric communication between ubiquitin degron binding and the peptidase activities. These results revise existing concepts on the 26S proteasome function and regulation, opening important opportunities for further research.
Mesophyll conductance response to short-term changes in CO2 is related to leaf anatomy and biochemistry in diverse C4 grasses
Authors: Pathare Varsha S. , DiMario Robert J. , Koteyeva Nouria , Cousins Asaph B.
Published: October 2021

Mesophyll CO2 conductance (gm) in C3 species responds to short-term (minutes) changes in environment potentially due to changes in some leaf anatomical and biochemical properties and due to measurement artifacts. Compared to C3 species, there is less information about gm responses to short-term changes in environment conditions like pCO2 across diverse C4 species and the potential determinants of these responses.

Using 16 diverse C4 grasses we investigated the response of gm to short-term changes in CO2 and how this response related to the leaf anatomical and biochemical traits.

For all the measured C4-grasses gm increased as CO2 decreased; however, the percent change in gm varied (+13% to +250%) and significantly related to percent changes in leaf transpiration efficiency (TEi). The percent increase in gm was highest in grasses with thinner mesophyll cell walls and greater leaf nitrogen, activities of phosphoenolpyruvate carboxylase (PEPC), Rubisco and carbonic anhydrase, and a higher affinity of PEPC for bicarbonate.

Our study demonstrates that CO2 response of gm varies greatly across diverse C4 grasses and identifies the key leaf anatomical and biochemical traits related to this variation. These findings have implications for improving C4 photosynthetic models, and in attempts to improve TEi through manipulation of gm.
Neural basis for adaptive motor behavior during car driving
Authors: Ohata Ryu , Ogawa Kenji , Imamizu Hiroshi
Published: October 2021

Car driving is supported by motor skills trained through continuous daily practice. One of the skills unique to expert drivers is the ability to detect abrupt changes in the driving environment and then quickly adapt their operation mode to the changes. Previous functional neuroimaging studies on motor control investigated the mechanisms underlying behaviors adaptive to changes in control properties of simple experimental devices such as a computer mouse or a joystick. The switching of multiple internal models mainly engages adaptive behaviors and underlies the interplay between the cerebellum and frontoparietal network (FPN) regions as the neural process. However, it remains unclear whether the neural mechanisms identified through an experimental paradigm using such simple devices also underlie practical driving behaviors. In the current study, we measure functional magnetic resonance imaging (fMRI) activities while participants control a realistic driving simulator inside the MRI scanner. Here, the accelerator sensitivity of a virtual car is abruptly changed, requiring participants to respond to this change as quickly as possible. We first compare brain activities before and after the sensitivity change. As a result, sensorimotor areas, including the left cerebellum, increase their activities after the sensitivity change. Moreover, after the change, activity significantly increases in the inferior parietal lobe and dorsolateral prefrontal cortex, parts of the FPN regions. By contrast, the posterior cingulate cortex, a part of the default mode network, deactivates after the sensitivity change. Our results suggest that the neural bases found in previous experiments using the simpler devices can serve as the foundation of adaptive car driving. At the same time, this study also highlights the unique contribution of non-motor-related regions to addressing the high cognitive demands of driving.
Two-color super-resolution localization microscopy via joint encoding of emitter location and color
Authors: Wang Yujie , Kuang Weibing , Shang Mingtao , Huang Zhen-Li
Published: October 2021

Multi-color super-resolution localization microscopy (SRLM) provides great opportunities for studying the structural and functional details of biological samples. However, current multi-color SRLM methods either suffer from medium to high crosstalk, or require a dedicated optical system and a complicated image analysis procedure. To address these problems, here we propose a completely different method to realize multi-color SRLM. This method is built upon a customized RGBW camera with a repeated pattern of filtered (Red, Green, Blue and Near-infrared) and unfiltered (White) pixels. With a new insight that RGBW camera is advantageous for color recognition instead of color reproduction, we developed a joint encoding scheme of emitter location and color. By combing this RGBW camera with the joint encoding scheme and a simple optical set-up, we demonstrated two-color SRLM with ∼20 nm resolution and < 2% crosstalk (which is comparable to the best reported values). This study significantly reduces the complexity of two-color SRLM (and potentially multi-color SRLM), and thus offers good opportunities for general biomedical research laboratories to use multi-color SRLM, which is currently mastered only by well-trained researchers.
Antibody-mediated depletion of human CLEC-2 in a novel humanised mouse model
Authors: Brown Helena C , Beck Sarah , Navarro Stefano , Di Ying , Soriano Jerez Eva M , Kaczmarzyk Jana , Thomas Steven G. , Mirakaj Valbona , Watson Steve P , Nieswandt Bernhard , Stegner David
Published: October 2021

Platelet C-type lectin-like receptor 2 (CLEC-2) has been proposed as a potential anti-thrombotic target as genetic or antibody-mediated receptor deficiency prevents occlusive thrombus formation in mice. This occurs through interaction with an unknown ligand as the endogenous ligand podoplanin is not present in the vasculature. However, the CLEC-2-podoplanin interaction does have an important role in tumour metastasis. There are currently no methods to test potential human therapeutics targeting CLEC-2, such as antibodies,in vivo. We have therefore generated and characterised a humanised CLEC-2 mouse (hCLEC-2KI) and developed a novel monoclonal anti-human CLEC-2 antibody, HEL1, forin vivotesting. hCLEC-2KImice were phenotypically normal and had comparable platelet glycoprotein receptor expression, activation and aggregation to wildtype platelets. hCLEC-2KImice had both comparable bleeding and vessel occlusion times to WT mice. Challenging hCLEC-2KImice with HEL1 or a second monoclonal anti-hCLEC-2 antibody, AYP1, resulted in transient thrombocytopenia as well as CLEC-2 depletion for more than 2 weeks but had no effect on haemostasis. This illustrates the power of the humanised CLEC-2 mouse model in evaluating novel therapeuticsin vivo, including antibodies that target CLEC-2, as well as the limited effect on haemostasis when targeting CLEC-2.
Natural influent bioaugmentation of activated sludge water resource recovery systems: implications for low temperature nitrification and heterotrophic population structures
Authors: Jauffur Shameem , Bakhshi Zeinab , Frigon Dominic
Published: October 2021

This work aimed at demonstrating the natural bioaugmentation of biological activated sludge systems with nitrifying biomass from influent wastewater in lab-scale reactors. Three sequencing batch reactors (SBR) were fed with sterile synthetic wastewater. While nitrification was complete at a temperature of 8 °C and a SRT of 20 days, it failed when the temperature was lowered to 5 °C, and the SRT decreased to 7 days. In the test period, the sterile synthetic wastewater fed to the Test Reactor was supplemented by influent solids harvested at a full-scale treatment facility at a total suspended solids concentration of 100 mg/L, which corresponded to approximately 5 mg-COD/L of nitrifying biomass. Upon this addition, nitrification was restores. Subsequent halting the supply of influent solids to the Test Reactor led a rapid failure of nitrification and washout of nitrifiers from the SBR. Reproducibility was demonstrated by switching the feed composition between the Test and Negative control reactors. PCR-based amplicon sequencing analyses targeting the amoA, and nxrB genes of the Nitrospira genus have shown that the influent wastewater governed the structure and composition of the activated sludge nitrifying populations. The most abundant ammonia-oxidizing bacteria (AOB) and Nitrospira-related nitrite-oxidizing bacteria (NOB) in the influent seeds occurred as the most dominant ones in the activated sludge. This pattern was observed even when the influent seeds varied over time. The heterotrophic populations were less affected by the influent seeds with the activated sludge and raw sewage showing distinct microbial populations based on principal coordinate analysis (PCoA). However, the immigrant populations appeared to modulate the structure of the activated sludge heterotrophic communities to some extent. These findings provide concrete evidence of the presence of active nitrifiers in raw wastewater capable of supporting nitrification in an otherwise non-conducive environment. This may have important implications on process design, operation and optimization of wastewater treatment systems.

Highlights

Lab-scale reactors fed sterile synthetic wastewater at low temperatures and SRTs.

Nitrification failed when conditions were adjusted to 5 °C and a SRT of 7 days.

Nitrification restored by addition of real wastewater influent solids.

Nitrifiers in solids from sewers naturally bioaugment activated sludge systems.

Activated sludge models should consider the immigration of nitrifiers with influent.

Graphical abstract
A sensitized genetic screen to identify regulators ofC. elegansgermline stem cells
Authors: Robinson-Thiewes Sarah , Kershner Aaron M. , Shin Heaji , Haupt Kimberly A. , Kroll-Connor Peggy , Kimble Judith
Published: October 2021

Germline stem cells (GSCs) inCaenorhabditis elegansare maintained by GLP-1/Notch signaling from the niche and by a downstream RNA regulatory network. Loss of the GLP-1 receptor causes GSCs to precociously undergo meiotic differentiation, the “Glp” phenotype, due to a failure to self-renew.lst-1andsygl-1are functionally redundant direct targets of GLP-1 signaling whose gene products work with PUF RNA binding proteins to promote GSC self-renewal. Whereas single loss-of-function mutants are fertile,lst-1 sygl-1double mutants are sterile and Glp. We set out to identify genes that function redundantly with eitherlst-1orsygl-1to maintain GSCs. To this end, we conducted forward genetic screens for Glp mutants in genetic backgrounds lacking functional copies of eitherlst-1orsygl-1. The screens generated nineglp-1alleles, twolst-1alleles, and one allele ofpole-1, which encodes the catalytic subunit of DNA polymerase ε. Threeglp-1alleles reside in Ankyrin (ANK) repeats not previously mutated.pole-1single mutants have a low penetrance Glp that is enhanced by loss of eitherlst-1orsygl-1. Thus, the screen uncovered one locus that interacts genetically with bothlst-1andsygl-1and generated useful mutations for further studies of GSC regulation.
Real-time imaging of optic nerve head collagen microstructure and biomechanics using instant polarized light microscopy
Authors: Lee Po-Yi , Yang Bin , Hua Yi , Waxman Susannah , Zhu Ziyi , Ji Fengting , Sigal Ian A
Published: October 2021

Current tools lack the temporal or spatial resolution necessary to image many important aspects of the architecture and dynamics of the optic nerve head (ONH). We evaluated the potential of instant polarized light microscopy (IPOL) to overcome these limitations by leveraging the ability to capture collagen fiber orientation and density in a single image. Coronal sections through the ONH of fresh normal sheep eyes were imaged using IPOL while they were stretched using custom uniaxial or biaxial micro-stretch devices. IPOL allows identifying ONH collagen architectural details, such as fiber interweaving and crimp, and has high temporal resolution, limited only by the frame rate of the camera. Local collagen fiber orientations and deformations were quantified using color analysis and image tracking techniques. We quantified stretch-induced collagen uncrimping of lamina cribrosa (LC) and peripapillary sclera (PPS), and changes in LC pore size (area) and shape (convexity and aspect ratio). The simultaneous high spatial and temporal resolutions of IPOL revealed complex ONH biomechanics: i) stretch-induced local deformation of the PPS was nonlinear and nonaffine. ii) under load the crimped collagen fibers in the PPS and LC straightened, without torsion and with only small rotations. iii) stretch-induced LC pore deformation was anisotropic and heterogeneous among pores. Overall, with stretch the pores were became larger, more convex, and more circular. We have demonstrated that IPOL reveals details of collagen morphology and mechanics under dynamic loading previously out of reach. IPOL can detect stretch-induced collagen uncrimping and other elements of the tissue nonlinear mechanical behavior. IPOL showed changes in pore morphology and collagen architecture that will help improve understanding of how LC tissue responds to load.

Highlights

We demonstrate that instant polarized light microscopy allows visualization and quantification of changes in optic nerve head collagen morphology and architecture under dynamic loading

We show crimped collagen fibers in the peripapillary sclera and lamina cribrosa straightening under load, without torsion and with only small rotations.

We show that stretch-induced local deformation of the peripapillary sclera was nonlinear and nonaffine.

We show that stretch-induced lamina cribrosa pore deformation was anisotropic and heterogeneous among pores.

Our results show this novel imaging technique could help understand the role of collagen microstructure in eye physiology, aging, and in biomechanics-related diseases, such as glaucoma and myopia.
Adverse events of special interest for COVID-19 vaccines - background incidences vary by sex, age and time period and are affected by the pandemic
Authors: Nyberg Fredrik , Lindh Magnus , Vanfleteren Lowie E.G.W. , Hammar Niklas , Wettermark Björn , Sundström Johan , Santosa Ailiana , Kirui Brian K. , Gisslén Magnus
Published: October 2021

Background

With large-scale COVID-19 vaccination implemented world-wide, safety signals needing rapid evaluation will emerge. We report population-based, age- and-sex-specific background incidence rates of conditions representing potential vaccine adverse events of special interest (AESI) for the Swedish general population using register data.

Methods

We studied an age/sex-stratified random 10% sample of the Swedish population on 1 Jan 2020, followed for AESI outcomes during 1 year, as the COVID-19 pandemic emerged and developed, before the start of vaccinations. We selected and defined the following outcomes based on information from regulatory authorities, large-scale adverse events initiatives and previous studies: aseptic meningitis, febrile seizure, Kawasaki syndrome, MISC, post-infectious arthritis, arthritis, myocarditis, ARDS, myocardial infarction, stroke, ischemic stroke, hemorrhagic stroke, venous thromboembolism, pulmonary embolism, kidney failure, liver failure, erythema multiforme, disseminated intravascular coagulation, autoimmune thyroiditis, and appendicitis. We calculated incidence rates stratified by age, sex and time period (quarters of 2020), and classified them using Council of International Organizations of Medical Sciences (CIOMS) categories: very common, common, uncommon, rare, or very rare.

Results

We included 972,723 study subjects, representing the Swedish national population on 1 Jan 2020. We found that AESI incidence rates vary greatly by age and in some cases sex. Several common AESIs showed expected increase with age, while some (e.g. appendicitis, aseptic meningitis, autoimmune thyroiditis, Kawasaki syndrome and MISC) were more common in young people, and others exhibited a flatter age pattern (e.g. myocarditis, DIC and erythema multiforme). Consequently, the CIOMS classification for AESIs varied widely according to age. Considerable variability was suggested for some AESI rates across the 4 quarters of 2020, potentially related to pandemic waves, seasonal variation, healthcare system overload or other healthcare delivery effects.

Conclusion

Age, sex, and timing of rates are important to consider when background AESI rates are compared to corresponding rates observed with COVID-19 vaccines.
Influence of sex, season and environmental air quality on experimental human pneumococcal carriage acquisition
Authors: Cheliotis Katerina S. , Jewell Christopher P. , Solórzano Carla , Urban Britta , Collins Andrea M. , Mitsi Elena , Pojar Sherin , Nikolaou Elissavet , German Esther L. , Reiné Jesús , Gordon Stephen B. , Jochems Simon P. , Rylance Jamie , Ferreira Daniela M.
Published: October 2021

Streptococcus pneumoniae (pneumococcus) is the most common identified bacterial cause of pneumonia, and the leading infectious cause of death in children under five years of age worldwide. Pneumococcal disease follows a seasonal pattern with increased incidence during winter. Pneumonia burden is also associated with poor air quality. Nasopharyngeal carriage of the bacterium is a pre-requisite of invasive disease.

We aimed to determine if susceptibility to nasopharyngeal pneumococcal carriage varied by season, and which environmental factors might explain such variation. We also evaluated the influence of sex on susceptibility of carriage. We collated data from five studies in which human volunteers underwent intranasal pneumococcal challenge. Generalised linear mixed effects models were used to identify factors associated with altered risk of carriage acquisition, specifically climate and air-quality data.

During 2011-2017, 374 healthy adults were challenged with type 6B pneumococcus. Odds of carriage were significantly lower in males (OR, 0.61; 95% CI, 0.40-0.92; p = 0.02), and higher with cooler temperatures (OR, 0.79; 95% CI, 0.63-0.99; p = 0.04). Likelihood of carriage also associated with lower concentrations of local fine particulate matter concentrations (PM2.5) and increased local rainfall.

In contrast to epidemiologic series, experimental challenge allowed us to test propensity to acquisition during controlled exposures; immunologic explanations for sex and climatic differences should be sought.
Patch2Self denoising of diffusion MRI in the cervical spinal cord improves intra-cord contrast, signal modelling, repeatability, and feature conspicuity
Authors: Schilling Kurt G. , Fadnavis Shreyas , Batson Joshua , Visagie Mereze , Combes Anna J.E. , McKnight Colin D. , Bagnato Francesca , Garyfallidis Eleftherios , Landman Bennett A. , Smith Seth A. , O’Grady Kristin P.
Published: October 2021

Quantitative diffusion MRI (dMRI) is a promising technique for evaluating the spinal cord in health and disease. However, low signal-to-noise ratio (SNR) can impede interpretation and quantification of these images. The purpose of this study is to evaluate a denoising approach, Patch2Self, to improve the quality, reliability, and accuracy of quantitative diffusion MRI of the spinal cord. Patch2Self is a self-supervised learning-based denoising method that leverages statistical independence of noise to suppress signal components strictly originating from random fluctuations. We conduct three experiments to validate the denoising performance of Patch2Self on clinical-quality, single-shell dMRI acquisitions with a small number of gradient directions: 1) inter-session scanrescan in healthy volunteers to evaluate enhancements in image contrast and model fitting; 2) repeated intra-session scans in a healthy volunteer to compare signal averaging to Patch2Self; and 3) assessment of spinal cord lesion conspicuity in a multiple sclerosis group. We find that Patch2Self improves intra-cord contrast, signal modeling, SNR, and lesion conspicuity within the spinal cord. This denoising approach holds promise for facilitating reliable diffusion measurements in the spinal cord to investigate biological and pathological processes.
Proteasome-associated ubiquitin ligase relays target plant hormone-specific transcriptional activators
Authors: Wang Zhishuo , Orosa-Puente Beatriz , Nomoto Mika , Grey Heather , Potuschak Thomas , Matsuura Takakazu , Mori Izumi C. , Tada Yasuomi , Genschik Pascal , Spoel Steven H.
Published: October 2021

The ubiquitin-proteasome system is vital to hormone-mediated developmental and stress responses in plants. Ubiquitin ligases target hormone-specific transcriptional activators (TAs) for degradation, but how TAs are processed by proteasomes remains unknown. We report that in Arabidopsis the salicylic acid-and ethylene-responsive TAs, NPR1 and EIN3, are relayed from pathway-specific ubiquitin ligases to proteasome-associated HECT-type UPL3/4 ligases. Activity and stability of NPR1 was regulated by sequential action of three ubiquitin ligases, including UPL3/4, while proteasome processing of EIN3 required physical handover between ethylene-responsive SCFEBF2 and UPL3/4 ligases. Consequently, UPL3/4 controlled extensive hormone-induced developmental and stress-responsive transcriptional programmes. Thus, our findings identify unknown ubiquitin ligase relays that terminate with proteasome-associated HECT-type ligases, which may be a universal mechanism for processive degradation of proteasome-targeted TAs and other substrates.

One-Sentence Summary

Transcriptional activators are targeted by proteasomal ubiquitin ligase relays that control their activity and stability.
Inter-subunit Crosstalk via PDZ Synergistically Governs Allosteric Activation of Proapoptotic HtrA2
Authors: Parui Aasna L. , Mishra Vandana , Dutta Shubhankar , Bhaumik Prasenjit , Bose Kakoli
Published: October 2021

Mitochondrial serine protease – High temperature requirement A2 (HtrA2), is associated with various diseases including neurodegenerative disorders and cancer. Despite availability of structural details, the reports on HtrA2’s mechanistic regulation that varies with the type of activation signals still remain non-concordant. To expound the role of regulatory PDZ domains in promoting synergistic coordination between HtrA2 subunits, we generated heterotrimeric HtrA2 variants comprising different numbers of PDZs and/or active-site mutations. Sequential deletion of PDZs from the trimeric ensemble significantly affected its residual activity in a way that proffered a hypothesis advocating intermolecular allosteric crosstalk via PDZ domains in trimeric HtrA2. Furthermore, structural and computational snapshots affirmed the role of PDZs in secondary structural element formation and coordinated reorganization of the N-terminal region and regulatory loops. Therefore, apart from providing cues for devising structure-guided therapeutic strategies, this study establishes a physiologically relevant working model of complex allosteric regulation through a multifaceted trans-mediated cooperatively-shared energy landscape.
Stochastic variational variable selection for high-dimensional microbiome data
Authors: Dang Tung , Kumaishi Kie , Usui Erika , Kobori Shungo , Sato Takumi , Toda Yusuke , Yamasaki Yuji , Tsujimoto Hisashi , Ichihashi Yasunori , Iwata Hiroyoshi
Published: October 2021

Background

The rapid and accurate identification of a minimal-size core set of representative microbial species plays an important role in the clustering of microbial community data and interpretation of clustering results. However, the huge dimensionality of microbial metagenomics datasets is a major challenge for the existing methods such as Dirichlet multinomial mixture (DMM) models. In the framework of the existing methods, the computational burden of identifying a small number of representative species from a large number of observed species remains a challenge.

Results

We proposed a novel framework to improve the performance of the widely used DMM approach by combining three ideas: (i) we extended the finite DMM model to an infinite case by considering Dirichlet process mixtures and estimating the number of clusters as a random variables; (ii) we proposed an indicator variable to identify representative operational taxonomic units that substantially contribute to the differentiation among clusters; and (iii) to address the computational burden of high-dimensional microbiome data, we proposed a stochastic variational inference, which approximates the posterior distribution using a controllable distribution called variational distribution, and stochastic optimization algorithms for fast computation. Using the proposed method, stochastic variational variable selection (SVVS), we analyzed the root microbiome data collected in our soybean field experiment, the human gut microbiome data from three published datasets of large-scale case-control studies and the healthy human microbiome data from the Human Microbiome Project.

Conclusions

SVVS demonstrated a better performance and significantly faster computation than those of the existing methods in all cases of testing datasets. In particular, SVVS is the only method that can analyze massive high-dimensional microbial data with more than 50,000 microbial species and 1,000 samples. Furthermore, recent microbiome studies have suggested that selection of the microbial species used as a core set is important.
Evidence for the Emergence of β-Trefoils by ‘Peptide Budding’ from an IgG-like β-Sandwich
Authors: Longo Liam M. , Kolodny Rachel , McGlynn Shawn E.
Published: October 2021

As sequence and structure comparison algorithms gain sensitivity, the intrinsic interconnectedness of the protein universe has become increasingly apparent. Despite this general trend, β-trefoils have emerged as an uncommon counterexample: They are an isolated protein lineage for which few, if any, sequence or structure associations to other lineages have been identified. If β-trefoils are, in fact, remote islands in sequence-structure space, it implies that the oligomerizing peptide that founded the β-trefoil lineage itself arose de novo. To better understand β-trefoil evolution, and to probe the limits of fragment sharing across the protein universe, we identified both ‘β-trefoil bridging themes’ (evolutionarily-related sequence segments) and ‘β-trefoil-like motifs’ (structure motifs with a hallmark feature of the β-trefoil architecture) in multiple, ostensibly unrelated, protein lineages. The success of the present approach stems, in part, from considering β-trefoil sequence segments or structure motifs rather than the β-trefoil architecture as a whole, as has been done previously. The newly uncovered inter-lineage connections presented here suggest a novel hypothesis about the origins of the β-trefoil fold itself – namely, that it is a derived fold formed by ‘budding’ from an Immunoglobulin-like β-sandwich protein. These results demonstrate how the emergence of a folded domain from a peptide need not be a signature of antiquity and underpin an emerging truth: few protein lineages escape nature’s sewing table.
Integrative analysis of scRNAs-seq and scATAC-seq revealed transit-amplifying thymic epithelial cells expressing autoimmune regulator
Authors: Miyao Takahisa , Miyauchi Maki , Kelly S. Thomas , Terooatea Tommy W. , Ishikawa Tatsuya , Oh Eugene , Hirai Sotaro , Horie Kenta , Takakura Yuki , Ohki Houko , Hayama Mio , Maruyama Yuya , Seki Takao , Yabukami Haruka , Yoshida Masaki , Inoue Azusa , Sakaue-Sawano Asako , Miyawaki Atsushi , Muratani Masafumi , Minoda Aki , Akiyama Nobuko , Akiyama Taishin
Published: October 2021

Medullary thymic epithelial cells (mTECs) are critical for self-tolerance induction in T cells via promiscuous expression of tissue-specific antigens (TSAs), which are controlled by transcriptional regulator AIRE. Whereas AIRE-expressing (Aire+) mTECs undergo constant turnover in the adult thymus, mechanisms underlying differentiation of postnatal mTECs remain to be discovered. Integrative analysis of single-cell assays for transposase accessible chromatin (scATAC-seq) and single-cell RNA sequencing (scRNA-seq) suggested the presence of proliferating mTECs with a specific chromatin structure, which express high levels of Aire and co-stimulatory molecules CD80 (Aire+CD80hi). Proliferating Aire+CD80hi mTECs detected by using Fucci technology express a minimal level of Aire-dependent TSAs and are converted into quiescent Aire+CD80hi mTECs expressing high levels of TSAs after a transit amplification. These data provide evidence for the existence of transit amplifying Aire+mTEC precursors during Aire+mTEC differentiation process of the postnatal thymus.
All-trans retinoic acid induces GADD34 gene expression via transcriptional regulation by Six1-TLE3 and post-transcriptional regulation by p38-TTP in skeletal muscle
Authors: Adachi Yuichiro , Masuda Masashi , Sakakibara Iori , Uchida Takayuki , Niida Yuki , Mori Yuki , Kamei Yuki , Okumura Yosuke , Ohminami Hirokazu , Ohnishi Kohta , Yamanaka-Okumura Hisami , Nikawa Takeshi , Taketani Yutaka
Published: October 2021

All-trans retinoic acid (ATRA) increases the sensitivity to unfolded protein response (UPR) in differentiating leukemic blasts. The downstream transcriptional factors of PERK, a major arm of UPR, regulates muscle differentiation. However, the role of growth arrest and DNA damage-inducible protein 34 (GADD34), one of the downstream factors of PERK, and the effects of ATRA on GADD34 expression in muscle remain unclear. In this study, we identified ATRA increased the GADD34 expression independent of the PERK signal in the gastrocnemius muscle of mice. ATRA up-regulated GADD34 expression through the transcriptional activation of it via inhibiting the interaction of homeobox Six1 and transcription co-repressor TLE3 with the MEF3-binding site on the GADD34 gene promoter in myoblasts. ATRA also inhibited the interaction of TTP, which induces mRNA degradation, with AU-rich element on GADD34 mRNA via p38 MAPK, resulting in the instability of GADD34 mRNA. Overexpressed GADD34 in myoblasts changes the type of myosin heavy chain in myotubes. These results suggest ATRA increases GADD34 expression via transcriptional and post-transcriptional regulation in myoblasts, which changes muscle fiber type in myotubes.
Dalpiciclib and Pyrotinib Exert Synergistic Antitumor Effects in Triple Positive Breast Cancer
Authors: Bu Jiawen , Zhang Yixiao , Niu Nan , Bi Kewei , Sun Lisha , Qiao Xinbo , Wang Yimin , Zhang Yinan , Jiang Xiaofan , Wang Dan , Ma Qingtian , Li Huajun , Liu Caigang
Published: October 2021

Background

The therapeutic benefit of the standard combination of anti-HER2 and chemotherapy in triple-positive breast cancer (TPBC) is limited even after the addition of endocrine therapy to the regimen. Therefore, treatment optimization is required urgently.

Methods

Through the drug sensitivity test, the drug combination efficacy of anti-HER2 drug, endocrine drug and CDK4/6 inhibitor to BT474 cells were tested. The underlying molecular mechanisms were investigated using immunofluorescence, western blot analysis, immunohistochemical staining and cell cycle analysis. Potential biomarker which may indicate the responsiveness to drug treatment in triple positive breast cancer was selected out using RNA-sequence and tested using immunohistochemical staining.

Results

We found that pyrotinib combined with dalpiciclib showed better efficacy than pyrotinib combined with tamoxifen in BT474 cells. Degradation of HER2 could enhance ER nuclear transportation, whereas cell cycle blockers could reverse this process. This may be the underlying mechanism by which the addition of dalpiciclib was more beneficial than the addition of pyrotinib plus tamoxifen. Furthermore, CALML5 was revealed to be a potential indicator of responsiveness to anti-HER2 therapy plus CDK4/6 inhibition in triple positive breast cancer.

Conclusion

Our study provided evidence for the introduction of CDK4/6 inhibitor in the treatment of TPBC and indicated that the combination of anti-HER2 therapy and cell cycle blockers may be a better strategy for TPBC treatment.

Funding

This study was supported by the National Natural Science Foundation of China (#U20A20381, #81872159)
The zinc-binding motif in tankyrases is required for the structural integrity and proper function of the catalytic domain
Authors: Sowa Sven T. , Lehtiö Lari
Published: October 2021

Tankyrases are ADP-ribosylating enzymes that regulate many physiological processes in the cell and they are therefore possible drug targets for cancer and fibrotic diseases. The catalytic ADP-ribosyl-transferase domain of tankyrases contains a unique zinc-binding motif of unknown function. Recently, this motif was suggested to be involved in the catalytic activity of tankyrases. In this work, we set out to study the effect of the zinc-binding motif on activity, stability and structure of human tankyrases. We generated mutants of human TNKS1 and TNKS2 abolishing the zinc-binding capabilities and characterized the proteins biochemically and biophysically in vitro. We further generated a crystal structure of TNKS2, in which the zinc ion was oxidatively removed. Our work shows that the zinc-binding motif in tankyrases is a crucial structural element which is particularly important for the structural integrity of the acceptor site. While mutation of the motif rendered TNKS1 inactive likely due to introduction of major structural defects, the TNKS2 mutant remained active and displayed a different activity profile compared to the wild type.
Detection of Cell-Cell Interactions via Photocatalytic Cell Tagging
Authors: Oslund Rob C. , Reyes-Robles Tamara , White Cory H. , Tomlinson Jake H. , Crotty Kelly A. , Bowman Edward P. , Chang Dan , Peterson Vanessa M. , Li Lixia , Frutos Silvia , Vila-Perelló Miquel , Vlerick David , Cromie Karen , Perlman David H. , O’Hara Samantha D. , Roberts Lee R. , Piizzi Grazia , Hett Erik C. , Hazuda Daria J. , Fadeyi Olugbeminiyi O.
Published: October 2021

Cell-cell interactions drive essential biological processes critical to cell and tissue development, function, pathology, and disease outcome. The growing appreciation of immune cell interactions within disease environments has led to significant efforts to develop protein- and cell-based therapeutic strategies. A better understanding of these cell-cell interactions will enable the development of effective immunotherapies. However, characterizing these complex cellular interactions at molecular resolution in their native biological contexts remains challenging. To address this, we introduce photocatalytic cell tagging (PhoTag), a modality agnostic platform for profiling cell-cell interactions. Using photoactivatable flavin-based cofactors, we generate phenoxy radical tags for targeted labeling at the cell surface. Through various targeting modalities (e.g. MHC-Multimer, antibody, single domain antibody (VHH)) we deliver a flavin photocatalyst for cell tagging within monoculture, co-culture, and peripheral blood mononuclear cells. PhoTag enables highly selective tagging of the immune synapse between an immune cell and an antigen-presenting cell through targeted labeling at the cell-cell junction. This allowed for the ability to profile gene expression-level differences between interacting and bystander cell populations. Given the modality agnostic and spatio-temporal nature of PhoTag, we envision its broad utilization to detect and profile intercellular interactions within an immune synapse and other confined cellular regions for any biological system.
Slow TCA flux implies low ATP production in tumors
Authors: Bartman Caroline R. , Shen Yihui , Lee Won Dong , TeSlaa Tara , Jankowski Connor S.R. , Wang Lin , Yang Lifeng , Roichman Asael , Bhatt Vrushank , Lan Taijin , Hu Zhixian , Xing Xi , Lu Wenyun , Guo Jessie Yanxiang , Rabinowitz Joshua D.
Published: October 2021

The tricarboxylic acid (TCA) cycle oxidizes carbon substrates to carbon dioxide, with the resulting high energy electrons fed into the electron transport chain to produce ATP by oxidative phosphorylation. Healthy tissues derive most of their ATP from oxidative metabolism, and the remainder from glycolysis. The corresponding balance in tumors remains unclear. Tumors upregulate aerobic glycolysis (the Warburg effect), yet they also typically require an intact TCA cycle and electron transport chain1–6. Recent studies have measured which nutrients contribute carbon to the tumor TCA metabolites7,8, but not tumor TCA flux: how fast the cycle turns. Here, we develop and validate an in vivo dynamic isotope tracing-mass spectrometry strategy for TCA flux quantitation, which we apply to all major mouse organs and to five tumor models. We show that, compared to the tissue of origin, tumor TCA flux is markedly suppressed. Complementary glycolytic flux measurements confirm tumor glycolysis acceleration, but the majority of tumor ATP is nevertheless made aerobically, and total tumor ATP production is suppressed compared to healthy tissues. In murine pancreatic cancer, this is accommodated by downregulation of the major energy-using pathway in the healthy exocrine pancreas, protein synthesis. Thus, instead of being hypermetabolic as commonly assumed, tumors apparently make ATP at a lower than normal rate. We propose that, as cells de-differentiate into cancer, they eschew ATP-intensive processes characteristic of the host tissue, and that the resulting suppressed ATP demand contributes to the Warburg effect and facilitates cancer growth in the nutrient-poor tumor microenvironment.
Correcting Methylation Calls in Clinically Relevant Low-Mappability Regions
Authors: Kumar Caiden M. , Ryan Devon P. , Langhorst Bradley W.
Published: October 2021

DNA methylation is an important component in vital biological functions such as embryonic development, carcinogenesis, and heritable regulation. Accurate methods to assess genomic methylation status are crucial to its effective use in many scenarios, especially in the detection and diagnosis of disease. Methylation aligners, such as Bismark and bwa-meth, frequently assign significantly higher MapQ values than can be supported by the uniqueness of the region reads are mapped to. These incorrectly high MapQs result in inappropriate methylation calling in repetitive regions. We observe reads that should map to separate locations (possibly having different methylation states) actually end up mapping to the same locus, causing apparent mixed methylation at such loci. Methylation calling can be improved by using Bismap mappability data to filter out insufficiently unique reads. However, simply filtering out Cs in insufficiently unique regions is not adequate as it is prone to over-filtering Cs in small mappability dips. These Cs can in fact often be called using reads anchored in a nearby mappable region. We have created a new feature for the MethylDackel methylation caller to perform read-based filtering. This new methylation calling method resolves some of the apparent mixed methylation to either 0% or 100% methylation and removes many unsupportable methylation calls. We examined methylation calls with and without read-based filtering in or near the 7830 genes containing ClinVar variants in a methylation sequencing data set from the NA12878 cell line. Use of this improved method corrected 41,143 mixed methylation Cs to 0% methylation, and 22,345 to 100% methylation throughout the genome.