bluuebunny/crossref_metadata_2025 huggingface dataset:
Gene structure, differential exon usage, and expression of the testis long intergenic non-protein coding RNA 1016 in humans reveals isoform-specific roles in controlling biological processes
Authors: Ramos Enrique I. , Yang Barbara , Vasquez Yasmin M. , Lin Ken Y. , Choudhari Ramesh , Gadad Shrikanth S.
Published: March 2021

Long noncoding RNAs (lncRNAs) have emerged as critical regulators of biological processes. The constant expansion of newly-identified lncRNA genes requires that each one be comprehensively annotated to understand its molecular functions. Here, we describe a detailed characterization of the gene which encodes long intergenic non-protein coding RNA 01016 (LINC01016, a.k.a., LncRNA1195) with a focus on its structure, exon usage, and expression in human and macaque tissues. In this study, we show that it is exclusively conserved among non-human primates, suggesting its recent evolution and is expressed and processed into 12 distinct RNAs in testis, cervix, and uterus tissues. Further, we integrate de novo annotation of expressed LINC01016 transcripts and isoform-dependent gene expression analyses to show that human LINC01016 is a multi-exon gene, processed through differential exon usage with isoform-specific functions. Furthermore, in gynecological cancers, such as cervical squamous cell carcinoma and uterine corpus endometrial carcinoma, LINC01016 is downregulated; however, its higher expression is predictive of relapse-free survival in these cancers. Collectively, these analyses reveal that, unlike coding RNAs, lncRNA isoforms are differentially regulated and precisely processed in specific tissues to perform distinct biological roles.

One sentence summary

The distinct molecular role of LINC01016 isoforms reveals intricate biology associated with lncRNA transcription and processing.
Rapid evolutionary turnover of mobile genetic elements drives microbial resistance to viruses
Authors: Hussain Fatima Aysha , Dubert Javier , Elsherbini Joseph , Murphy Mikayla , VanInsberghe David , Arevalo Philip , Kauffman Kathryn , Rodino-Janeiro Bruno Kotska , Polz Martin
Published: March 2021

Although it is generally accepted that viruses (phages) drive bacterial evolution, how these dynamics play out in the wild remains poorly understood. Here we show that the arms race between phages and their hosts is mediated by large and highly diverse mobile genetic elements. These phage-defense elements display exceedingly fast evolutionary turnover, resulting in differential phage susceptibility among clonal bacterial strains while phage receptors remain invariant. Protection afforded by multiple elements is cumulative, and a single bacterial genome can harbor as many as 18 putative phage-defense elements. Overall, elements account for 90% of the flexible genome amongst closely related strains. The rapid turnover of these elements demonstrates that phage resistance is unlinked from other genomic features and that resistance to phage therapy might be as easily acquired as antibiotic resistance.
Genomic signatures of past and present chromosomal instability in the evolution of Barrett’s esophagus to esophageal adenocarcinoma
Authors: Stachler Matthew D. , Bao Chunyang , Tourdot Richard W. , Brunette Gregory J. , Stewart Chip , Sun Lili , Baba Hideo , Watanabe Masayuki , Agoston Agoston , Jajoo Kunal , Davison Jon M. , Nason Katie , Getz Gad , Wang Kenneth K. , Imamura Yu , Odze Robert , Bass Adam J. , Zhang Cheng-Zhong
Published: March 2021

The progression of precancerous lesions to malignancy is often accompanied by increasing complexity of chromosomal alterations but how these alterations arise is poorly understood. Here we performed haplotype-specific analysis of chromosomal copy-number evolution in the progression of Barrett’s esophagus (BE) to esophageal adenocarcinoma (EAC) on multiregional whole-genome sequencing data of BE with dysplasia and microscopic EAC foci. We identified distinct patterns of copy-number evolution indicating multigenerational chromosomal instability that is initiated by cell division errors but propagated only after p53 loss. While abnormal mitosis, including whole-genome duplication, underlies chromosomal copy-number changes, segmental alterations display signatures of successive breakage-fusion-bridge cycles and chromothripsis of unstable dicentric chromosomes. Our analysis elucidates how multigenerational chromosomal instability generates copy-number variation in BE cells, precipitates complex alterations including DNA amplifications, and promotes their independent clonal expansion and transformation. In particular, we suggest sloping copy-number variation as a signature of ongoing chromosomal instability that precedes copy-number complexity.

These findings suggest copy-number heterogeneity in advanced cancers originates from chromosomal instability in precancerous cells and such instability may be identified from the presence of sloping copy-number variation in bulk sequencing data.
Parental-fetal interplay of immune genes leads to intrauterine growth restriction
Authors: Kaur Gurman , Porter Caroline B. M. , Ashenberg Orr , Lee Jack , Riesenfeld Samantha J. , Hofree Matan , Aggelakopoulou Maria , Subramanian Ayshwarya , Kuttikkatte Subita Balaram , Attfield Kathrine E. , Desel Christiane A. E. , Davies Jessica L. , Evans Hayley G. , Avraham-Davidi Inbal , Nguyen Lan T. , Dionne Danielle A. , Neumann Anna E. , Jensen Lise Torp , Barber Thomas R. , Soilleux Elizabeth , Carrington Mary , McVean Gil , Rozenblatt-Rosen Orit , Regev Aviv , Fugger Lars
Published: March 2021

Intrauterine growth restriction (IUGR) of fetuses affects 5-10% of pregnancies and is associated with perinatal morbidity, mortality and long-term health issues. Understanding genetic predisposition to IUGR is challenging, owing to extensive gene polymorphisms, linkage disequilibrium, and maternal and paternal influence. Here, we demonstrate that the inhibitory receptor, KIR2DL1, expressed on maternal uterine natural killer (uNK) cells, in interaction with the paternally-inherited HLA-C*05, an HLA-C group 2 allotype, expressed on fetal trophoblast cells, causes IUGR in a humanised mouse model. Micro-CT imaging of the uteroplacental vasculature revealed reduced uterine spiral artery diameter and increased segment length, increasing fetal blood flow resistance. Single cell RNA-Seq from the maternal-fetal interface highlighted expression programs activated by KIR2DL1-induced IUGR in several placental cell types, including degradation of extracellular matrix components, angiogenesis, and uNK cell communication, suggesting a complex response underlying IUGR. As current IUGR treatments are insufficient, our findings provide important insight for drug development.
Paenibacillussp. strain UY79, isolated from a root nodule ofArachis villosa, displays a broad spectrum of antifungal activity
Authors: Costa Andrés , Corallo Belén , Amarelle Vanesa , Stewart Silvina , Pan Dinorah , Tiscornia Susana , Fabiano Elena
Published: March 2021

A nodule-inhabitingPaenibacillussp. strain (UY79) isolated from wild peanut (Arachis villosa) was screened for its antagonistic activity against diverse fungi and oomycetes (Botrytis cinerea, Fusarium verticillioides, Fusarium oxysporum, Fusarium graminearum, Fusarium semitectum, Macrophomina phaseolina, Phomopsis longicolla, Pythium ultimum, Phytophthora sojae, Rhizoctonia solani, Sclerotium rolfsiiandTrichoderma atroviride). Results obtained show thatPaenibacillussp. UY79 was able to antagonize these fungi/oomycetes and that agar-diffusible metabolites and enzymes, as well as volatile compounds (different from HCN), participate in the antagonism exerted. We found thatPaenibacillussp. strain UY79 did not affect symbiotic association or growth promotion of alfalfa plants when co-inoculated with rhizobia. By whole genome sequence analysis, we determined that strain UY79 is a new species ofPaenibacilluswithin thePaenibacillus polymyxacomplex. Diverse genes putatively involved in biocontrol activity were identified in the UY79 genome. Moreover, according to genome mining and antibiosis assays, strain UY79 would have the capability to modulate the growth of bacteria commonly found in soil/plant communities.

IMPORTANCE

Phytopathogenic fungi and oomycetes are responsible for causing devastating losses in agricultural crops. Therefore, there is an enormous interest in the development of effective and complementary strategies that allow the control of the phytopathogens, reducing the input of agrochemicals in croplands. Discovery of new strains with expanded antifungal activities and with a broad spectrum of action is challenging and of great future impact. Diverse strains belonging to theP. polymyxacomplex have been reported to be effective biocontrol agents. Results presented here show that the novel discovered strain ofPaenibacillussp. presents diverse traits involved in antagonistic activity against a broad spectrum of pathogens and would be a potential and valuable strain to be further assessed for the development of biofungicides.
The Detection ofBergeyella zoohelcumin Young Children after Cat Bite
Authors: Hu Min , Li Hao , Hu Tao , Zhou Shuang-Yan , Yang Ze-Hua , Fan Cai-Lan , Bao Rong , Shi Jian-Guo , Zhao Ke-Bin
Published: March 2021

Bergeyella zoohelcumis an uncommon zoonotic pathogen typically associated with cat or dog bites. Previously, only 14 cases ofB. zoohelcuminfection have been reported. We isolated the bacteria from the face of a 2-year-old girl who was bitten by a cat. The organism was identified by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and 16S rRNA gene sequence, and phylogenetic analysise supported that this isolate was belong toB. zoohelcum. Due to the contradicts of culture characteristics ofB. zoohelcumwhich described in reported literatures, we used different media to culture the bacteria. After 24 h incubation, Colombia blood agar (CBA), Mueller-Hinton agar plate with 5% sheep blood and chocolate agar (CA) grew well, but blood agar (BA) grew well until 72 h. The strain did not grow on McConkey agar (MAC), Mueller-Hinton agar (MHA) and chocolate agar (containing vancomycin). The low detection rate of this strain was related to its harsh growth conditions and the limitations of traditional identification techniques. With the popularization of MALDI-TOF MS and 16S rRNA gene sequence and our further understanding of the fastidious bacteria, we will quickly and accurately identify theB. zoohelcum, meanwhile the detection rate of this bacterium will also be significantly improved. So that clinicians can achieve precise anti-infective treatment according to antimicrobial susceptibility testing.
How unequal vaccine distribution promotes the evolution of vaccine escape
Authors: Gerrish Philip J , Saldaña Fernando , Galeota-Sprung Benjamin , Colato Alexandre , Rodriguez Erika E , Hernández Jorge X Velasco
Published: March 2021

Health officials warn that SARS-CoV-2 vaccines must be uniformly distributed within and among countries if we are to quell the ongoing pandemic. Yet there has been little critical assessment of the underlying reasons for this warning. Here, we explicitly show why vaccine equity is necessary. Perhaps counter-intuitively, we find that vaccine escape mutants are less likely to come from highly vaccinated regions where there is strong selection pressure favoring vaccine escape and more likely to come from neighboring unvaccinated regions where there is no selection favoring escape. Unvaccinated geographic regions thus provide evolutionary reservoirs from which new strains can arise and cause new epidemics within neighboring vaccinated regions and beyond. Our findings have timely implications for vaccine rollout strategies and public health policy.
Estimation of SARS-CoV-2 antibody prevalence through integration of serology and incidence data
Authors: Wang Liangliang , Min Joosung , Doig Renny , Elliott Lloyd T , Colijn Caroline
Published: March 2021

Serology tests for SARS-CoV-2 provide a paradigm for estimating the number of individuals who have had infection in the past (including cases that are not detected by routine testing, which has varied over the course of the pandemic and between jurisdictions). Classical statistical approaches to such estimation do not incorporate case counts over time, and may be inaccurate due to uncertainty about the sensitivity and specificity of the serology test. In this work, we provide a joint Bayesian model for case counts and serological data, integrating uncertainty through priors on the sensitivity and specificity. We also model the Phases of the pandemic with exponential growth and decay. This model improves upon maximum likelihood estimates by conditioning on more data, and by taking into account the epidemiological trajectory. We apply our model to the greater Vancouver area, British Columbia, Canada with data acquired during Phase 1 of the pandemic.
Th17 cell master transcription factor RORC2 regulates HIV-1 gene expression and viral outgrowth
Authors: Wiche Salinas Tomas Raul , Zhang Yuwei , Sarnello Daniele , Zhyvoloup Alexander , Raymond Marchand Laurence , Planas Delphine , Lodha Manivel , Chatterjee Debashree , Karwacz Kasia , Oxenford Sally , Routy Jean-Pierre , Amrine-Madsen Heather , Ancuta Petronela , Fassati Ariberto
Published: March 2021

Among CD4+ T-cells, T helper 17 (Th17) cells are particularly susceptible to HIV-1 infection and are depleted from mucosal sites, which causes damage to the gut barrier resulting in microbial translocation-induced systemic inflammation, a hallmark of disease progression. Furthermore, a proportion of latently infected Th17 cells persist long-term in the gastro-intestinal lymphatic tract, where low-level HIV-1 transcription is observed. This residual viremia contributes to chronic immune activation. Thus, Th17 cells are key players in HIV pathogenesis and viral persistence, however it is unclear why these cells are highly susceptible to HIV-1 infection. Th17 cell differentiation depends on expression of the master transcriptional regulatorRORC2, aretinoic acid-related nuclear hormone receptor that regulates specific transcriptional programs by binding to promoter/enhancer DNA. Here, we report that RORC2 is a key host-cofactor for HIV replication in Th17 cells. We found that specific inhibitors that bind to the RORC2 ligand-binding domain reduced HIV replication in CD4+ T-cells. Depletion of RORC2 inhibited HIV-1 infection, whereas RORC2 overexpression enhanced it. RORC2 was found to promote HIV-1 gene expression. Chromatin immune precipitation revealed that RORC2 binds to the nuclear receptor responsive element (NRRE) in the HIV-1 LTR. In treated HIV-1 patients, RORC2+ CD4 T cells contained more proviral DNA than RORC2− cells. Pharmacological inhibition of RORC2 potently reduced HIV-1 outgrowth in CD4+ T-cells from antiretroviral-treated patients. Altogether, these results provide a new explanation as to why Th17 cells are highly susceptible to HIV-1 infection and point to RORC2 as a cell-specific target for HIV-1 therapy.

Significance statement

HIV-1 infects CD4 T cells and among these, Th17 cells are known to be particularly permissive for virus replication. Infection of Th17 cells is critical for AIDS pathogenesis and viral persistence, however it is not clear why these cells are highly permissive to HIV-1. We found that Th17 cell permissiveness depends on expression of the hormone receptor RORC2, which is the master transcriptional regulator of Th17 cell differentiation. We identify RORC2 as a new, cell-specific host-dependency factor that can be targeted by small molecules. Our results point to RORC2 as a cell-specific target for HIV-1 therapy, an entirely new concept in the field, and suggest HIV-1 might have evolved to exploit RORC2 to promote its own persistence.
Canagliflozin increases adenoma burden in female APCMin/+mice
Authors: Korfhage Justin , Skinner Mary E. , Basu Jookta , Greenson Joel K. , Miller Richard A. , Lombard David B.
Published: March 2021

The diabetes drug canagliflozin acts primarily by inhibiting glucose reuptake by the sodium glucose transporter 2 (SGLT2) in the kidney proximal tubule, thereby lowering serum glucose levels. Canagliflozin also acts on SGLT1, a related transporter responsible for glucose uptake in the small intestine and more distal kidney tubules. Several cancers overexpress SGLT1 and SGLT2, where these transporters fuel tumor metabolism. A recent study by NIA’s Interventions Testing Program (ITP) showed that canagliflozin treatment extends lifespan in male mice. Since cancer is the major cause of death in most mouse strains, including the UM-HET3 strain used by the ITP, this observation suggests that canagliflozin might exert anti-cancer effects in this context. Here, we treated a commonly-used mouse neoplasia model -- the intestinal adenoma-prone APCMin/+strain -- with canagliflozin, to test the effects of drug treatment on tumor burden. Surprisingly, canagliflozin increased the total area of intestine involved by adenomas, an effect that was most marked in the distal intestine and in female mice. Immunohistochemical analysis suggested that canagliflozin may not influence adenoma growth via direct SGLT1/2 inhibition in neoplastic cells themselves. Instead, our results are most consistent with a model whereby canagliflozin aggravates adenoma development by altering the anatomic distribution of intestinal glucose absorption, as evidenced by increases in postprandial GLP-1 levels consistent with delayed glucose absorption. Our results suggest that canagliflozin exacerbates adenomatosis in the APCMin/+model via complex, cell-non-autonomous mechanisms, and hint that sex differences in incretin responses may underlie differential effects of this drug on lifespan.
In vivodissection of Rhoa function in vascular development using zebrafish
Authors: Pillay Laura M. , Yano Joseph J. , Davis Andrew E. , Butler Matthew G. , Barnes Keith A. , Reyes Vanessa L. , Castranova Daniel , Gore Aniket V. , Swift Matthew R. , Iben James R. , Stratman Amber N. , Weinstein Brant M.
Published: March 2021

Rationale

The small monomeric GTPase RHOA acts as a master regulator of signal transduction cascades by activating effectors of cellular signaling, including the Rho-associated protein kinases ROCK1/2. Previousin vitrocell culture studies suggest that RHOA can regulate many critical aspects of vascular endothelial cell (EC) biology, including focal adhesion, stress fiber formation, and angiogenesis. However, the specificin vivoroles of RHOA during vascular development and homeostasis are still not well understood.

Objective

In this study we examine thein vivofunctions of RHOA in regulating vascular development and integrity in zebrafish.

Methods and Results

We use zebrafish RHOA-ortholog (rhoaa) mutants, transgenic embryos expressing wild type, dominant-negative, or constitutively active forms ofrhoaain ECs, and a pharmacologic inhibitor of ROCK1/2 to study thein vivoconsequences of RHOA gain- and loss-of-function in the vascular endothelium. Our findings document roles for RHOA in vascular integrity, developmental angiogenesis, and vascular morphogenesis.

Conclusions

Our results indicate that either too much or too little RHOA activity leads to vascular dysfunctionin vivo.
aPC/PAR1 confers endothelial anti-apoptotic activity via a discrete β-arrestin-2 mediated SphK1-S1PR1-Akt signaling axis
Authors: Molinar-Inglis Olivia , Birch Cierra A. , Nicholas Dequina , Cisneros-Aguirre Metzli , Patwardhan Anand , Chen Buxin , Grimsey Neil J. , Gomez-Menzies Patrick K. , Lin Huilan , Coronel Luisa J. , Lawson Mark A. , Patel Hemal H. , Trejo JoAnn
Published: March 2021

Endothelial dysfunction is associated with multiple vascular diseases and lacks effective treatments. Activated Protein C (aPC) is a promising biotherapeutic that signals via protease-activated receptor-1 (PAR1) to promote diverse cytoprotective responses, including endothelial barrier stabilization, anti-inflammatory and anti-apoptotic activities, which is facilitated by co-receptors. We showed that aPC-activated PAR1 signals preferentially via b-arrestin-2 (b-arr2) and dishevelled-2 (Dvl2) scaffolds rather than G proteins to enhance barrier protection. However, the mechanisms by which aPC/PAR1 promotes other cytoprotective responses are poorly understood. Here we define a novel β-arr2-mediated sphingosine kinase-1 (SphK1)-sphingosine-1-phosphate receptor-1 (S1PR1)-Akt signaling axis that confers aPC/PAR1-mediated protection against cell death. We show that PAR1 and S1PR1 co-exist in caveolin-1-rich microdomains basally and aPC markedly increases S1PR1-caveolin-1 co-association. Moreover, aPC stimulates b-arr2-dependent SphK1 activation independent of Dvl2, which is critical for S1PR1 transactivation. These studies reveal that different aPC/PAR1 cytoprotective responses are mediated by discrete b-arr2-driven signaling pathways in caveolae.
Rapid Evolution of SARS-CoV-2 Challenges Human Defenses
Authors: Duarte Carlos M. , Ketcheson David , Eguíluz Víctor M. , Agustí Susana , Fernández-Gracia Juan , Jamil Tahira , Laiolo Elisa , Gojobori Takashi , Alam Intikhab
Published: March 2021

Evolutionary ecology theory provides an avenue to anticipate the future behavior of SARS-CoV-2. Here we quantify the accelerating evolution of SARS-CoV-2 by tracking the SARS-CoV-2 mutation globally, with a focus on the Receptor Binding Domain (RBD) of the spike protein believed to determine infectivity. We estimate that 384 million people were infected by March 1st, 2021, producing up to 1021copies of the virus, with one new RBD variant appearing for every 600,000 human infections, resulting in approximately three new effective RBD variants produced daily. Doubling the number of RBD variants every 71.67 days followed by selection of the most infective variants challenges our defenses and calls for a shift to anticipatory, rather than reactive tactics.

One-Sentence Summary

Accelerating evolution of SARS-CoV-2 demands formulating universal vaccines and treatments based on big-data simulations of possible new variants.
Dynamic phase separation of the androgen receptor and its coactivators to regulate gene expression
Authors: Zhang Fan , Wong Samantha , Lee Joseph , Lingadahalli Shreyas , Wells Christopher , Saxena Neetu , Sanchez Christophe , Sun Bei , Parra-Nuñez Ana Karla , Chan Novia , Bui Jennifer M. , Wang Yuzhuo , Rennie Paul S. , Lack Nathan , Cherkasov Artem , Gleave Martin , Gsponer Jörg , Lallous Nada
Published: March 2021

Numerous cancers, including prostate cancer (PCa), are addicted to transcription programs driven by superenhancers (SEs). The transcription of genes at SEs is enabled by the formation of phase-separated condensates by transcription factors and co-activators with intrinsically disordered regions. The androgen receptor (AR), main oncogenic driver in PCa, contains large disordered regions and is co-recruited with the co-activator MED1 to SEs to promote oncogenic programs. In this work, we show that dynamic AR-rich, liquid-like foci form in PCa models upon androgen stimulation and correlate with AR transcriptional activity. The co-activator MED1 plays an essential role in the formation of AR foci while AR antagonists hinder their formation. These results suggest that enhanced compartmentalization of AR and co-activators at SEs may play an important role in the activation of oncogenic transcription programs in PCa. A better understanding of the assembly and the regulation of these AR-rich compartments may provide novel therapeutic options.
Decoding regulatory specificity of human ribosomal proteins
Authors: Luan Yizhao , Tang Nan , Yang Jiaqi , Chen Congying , Liu Shuting , Cheng Chichi , Wang Yan , Guo Ya-nan , Wang Hongwei , Xiang Mengqing , Ju Rong , Xie Zhi
Published: March 2021

Human ribosomes, made of around 80 ribosomal proteins (RPs) and four ribosomal RNAs, have long been thought as uniform passive protein-making factories with little regulatory function. Recently, accumulating evidence showed heterogeneity of RP composition in ribosomes responsible for regulating gene expression in development and tumorigenesis. However, a comprehensive understanding of regulatory spectrum of RPs is still lacking. In this study, we conducted a systematic survey of regulatory specificity of human RPs on global gene expression. We quantified translational and transcriptional changes of gene expression upon deficiency of 75 RPs, including 44 from the large subunit (60S) and 31 from the small subunit (40S), by ribosomal profiling and RNA sequencing analysis. We showed that RP deficiency induced diverse expression changes, particularly at the translational level. RPs were subjected to co-translational regulation under ribosomal stress where deficiency of the 60S and the 40S RPs had opposite effects on the two subunits. RP deficiency perturbed expression of genes related to cell cycle, cellular metabolism, signal transduction and development. Deficiency of RPs from the 60S led to a greater repression effect on cell growth than that from the 40S by perturbing P53 signaling and cell cycle pathways. To demonstrate functional specificity of RPs, we showed thatRPS8deficiency stimulated cellular apoptosis andRPL13orRPL18deficiency promoted cellular senescence. We also showed thatRPL11andRPL15played important roles in retina development and angiogenesis, respectively. Overall, our study demonstrated a widespread regulatory role of RPs in controlling cellular activity, providing an important resource which can offer novel insights into ribosome regulation in human diseases and cancer.
Validating Pore Size Estimates in a Complex Microfibre Environment on a Human MRI System
Authors: Huang Chu-Chung , Hsu Chih-Chin Heather , Zhou Feng-Lei , Kusmia Slawomir , Drakesmith Mark , Parker Geoff J.M. , Lin Ching-Po , Jones Derek K.
Published: March 2021

Purpose

Recent advances in diffusion-weighted MRI provide ‘restricted diffusion signal fraction’ and restricting pore size estimates. Materials based on co-electrospun oriented hollow cylinders have been introduced to provide validation for such methods. This study extends this work, exploring accuracy and repeatability using an extended acquisition on a 300 mT/m gradient human MRI scanner, in substrates closely mimicking tissue, i.e., non-circular cross-sections, intra-voxel fibre crossing, intra-voxeldistributionsof pore-sizes and smaller pore-sizes overall.

Methods

In a single-blind experiment, diffusion-weighted data were collected from a biomimetic phantom on a 3T Connectom system using multiple gradient directions/diffusion times. Repeated scans established short-term and long-term repeatability. The total scan time (54 minutes) matched similar protocols used in human studies. The number of distinct fibre populations was estimated using spherical deconvolution, and median pore size estimated through the combination of CHARMED and AxCaliber3D framework. Diffusion-based estimates were compared with measurements derived from scanning electron microscopy.

Results

The phantom contained substrates with different orientations, fibre configurations and pore size distributions. Irrespective of one or two populations within the voxel, the pore-size estimates (~5μm) and orientation-estimates showed excellent agreement with the median values of pore-size derived from scanning electron microscope and phantom configuration. Measurement repeatability depended on substrate complexity, with lower values seen in samples containing crossing-fibres. Sample-level repeatability was found to be good.

Conclusion

While no phantom mimics tissue completely, this study takes a step closer to validating diffusion microstructure measurements for usein vivoby demonstrating the ability to quantify microgeometry in relatively complex configurations.
The formation of the bacterial RNA polymerase–promoter open complex involves a branched pathway
Authors: Malinen Anssi M. , Bakermans Jacob , Aalto-Setälä Emil , Blessing Martin , Bauer David L.V. , Parilova Olena , Belogurov Georgiy A. , Dulin David , Kapanidis Achillefs N.
Published: March 2021

The expression of most bacterial genes commences with the binding of RNA polymerase (RNAP)–σ70holoenzyme to the promoter DNA. This initial RNAP–promoter closed complex undergoes a series of conformational changes, including the formation of a transcription bubble on the promoter and the loading of template DNA strand into the RNAP active site; these changes lead to the catalytically active open complex (RPO) state. Recent cryo-electron microscopy studies have provided detailed structural insight on the RPOand putative intermediates on its formation pathway. Here, we employ single-molecule fluorescence microscopy to interrogate the conformational dynamics and reaction kinetics during real-time RPOformation. We find that the RPOpathway is branched, generating RPOcomplexes with different stabilities. The RNAP cleft loops, and especially the β’ rudder, stabilise the transcription bubble. The RNAP interactions with the promoter upstream sequence (beyond −35) stimulate transcription bubble nucleation and tune the reaction path towards stable forms of the RPO. The mechanistic heterogeneity of the RPOpathway may be a prerequisite for its regulation since such heterogeneity allows the amplification of small promoter sequence or transcription-factor-dependent changes in the free energy profile of the RPOpathway to large differences in transcription efficiency.
Learning brain dynamics for decoding and predicting individual differences
Authors: Pessoa Luiz , Limbachia Chirag , Misra Joyneel , Surampudi Srinivas Govinda , Venkatesh Manasij , Jaja Joseph
Published: March 2021

Insights from functional Magnetic Resonance Imaging (fMRI), as well as recordings of large numbers of neurons, reveal that many cognitive, emotional, and motor functions depend on the multivariate interactions of brain signals. Todecodebrain dynamics, we propose an architecture based on recurrent neural networks to uncover distributed spatiotemporal signatures. We demonstrate the potential of the approach using human fMRI data during movie-watching data and a continuous experimental paradigm. The model was able to learn spatiotemporal patterns that supported 15-way movie-clip classification (~90%) at the level of brain regions, and binary classification of experimental conditions (~60%) at the level of voxels. The model was also able to learn individual differences in measures of fluid intelligence and verbal IQ at levels comparable or better than existing techniques. We propose a dimensionality reduction approach that uncovers low-dimensional trajectories and captures essential informational (i.e., classification related) properties of brain dynamics. Finally, saliency maps were employed to characterize brain-region/voxel importance, and uncovered how dynamic but consistent changes in fMRI activation influenced decoding performance. When applied at the level of voxels, our framework implements a dynamic version of multivariate pattern analysis. We believe our approach provides a powerful framework for visualizing, analyzing, and discovering dynamic spatially distributed brain representations during naturalistic conditions.1

Author summary

Brain signals are inherently dynamic and evolve in both space and time as a function of cognitive or emotional task condition or mental state. To characterize brain dynamics, we employed an architecture based on recurrent neural networks, and applied it to functional magnetic resonance imaging data from humans watching movies or during continuous experimental conditions. The model learned spatiotemporal patterns that allowed it to correctly classify which clip a participant was watching based entirely on data from other participants; the model also learned a binary classification of experimental conditions at the level of voxels. We developed a dimensionality reduction approach that uncovered low-dimensional “trajectories” and captured essential information properties of brain dynamics. When applied at the level of voxels, our framework implements a dynamic version of multivariate pattern analysis. We believe our approach provides a powerful framework for visualizing, analyzing, and discovering dynamic spatially distributed brain representations during naturalistic conditions.
Regenerating vascular mural cells in zebrafish fin blood vessels are not derived from pre-existing ones and differentially requirepdgfrbsignaling for their development
Authors: Leonard Elvin V. , Figueroa Ricardo J. , Bussmann Jeroen , Lawson Nathan D. , Amigo Julio D. , Siekmann Arndt F.
Published: March 2021

Vascular networks are comprised of endothelial cells and mural cells, which include pericytes and smooth muscle cells. It is well established that new endothelial cells are derived from pre-existing ones during the angiogenic phase of blood vessel growth. By contrast, mural cell ontogeny is less clear with an ongoing debate whether mural cells possess mesenchymal stem cell properties. To elucidate the mechanisms controlling mural cell recruitment during development and tissue regeneration, we studied the formation of zebrafish caudal fin arteries. Mural cells showed morphological heterogeneity: cells colonizing arteries proximal to the body wrapped around them, while those in more distal regions extended protrusions along the proximo-distal vascular axis. Despite these differences, both cell populations expressed platelet-derived growth factor receptor beta (Pdgfrb) and the smooth muscle cell marker myosin heavy chain 11a (Myh11a). Loss of Pdgfrb signalling during development or tissue regeneration resulted in a substantial decrease in mural cells at the vascular front, while those proximal to the body were less affected. Using lineage tracing, we demonstrate that precursor cells located in periarterial regions of the caudal fin and expressing Pgdfrb can give rise to mural cells, while in regeneration newly formed mural cells were not derived from pre-existing ones. Together, our findings reveal conserved roles for pdgfrb signalling in development and regeneration, while at the same time illustrating a limited capacity of mural cells to self-renew or contribute to other cell types during tissue regeneration.
Consensus tetratricopeptide repeat proteins are complex superhelical nanosprings
Authors: Synakewicz Marie , Eapen Rohan S. , Perez-Riba Albert , Bauer Daniela , Weißl Andreas , Fischer Gerhard , Hyvönen Marko , Rief Matthias , Itzhaki Laura S. , Stigler Johannes
Published: March 2021

Tandem-repeat proteins comprise small secondary structure motifs that stack to form one-dimensional arrays with distinctive mechanical properties that are proposed to direct their cellular functions. Here, we use single-molecule optical tweezers to study the folding of consensus-designed tetratricopeptide repeats (CTPRs) — superhelical arrays of short helix-turn-helix motifs. We find that CTPRs display a spring-like mechanical response in which individual repeats undergo rapid equilibrium fluctuations between folded and unfolded conformations. We rationalise the force response using Ising models and dissect the folding pathway of CTPRs under mechanical load, revealing how the repeat arrays form from the centre towards both termini simultaneously. Strikingly, we also directly observe the protein’s superhelical tertiary structure in the force signal. Using protein engineering, crystallography and single-molecule experiments, we show how the superhelical geometry can be altered by carefully placed amino-acid substitutions and examine how these sequence changes affect intrinsic repeat stability and inter-repeat coupling. Our findings provide the means to dissect and modulate repeat-protein stability and dynamics, which will be essential for researchers to understand the function of natural repeat proteins and to exploit artificial repeats proteins in nanotechnology and biomedical applications.

Significance statement

Repetition of biological building blocks is crucial to modulating and diversifying structure and function of biomolecules across all organisms. In tandem-repeat proteins, the linear arrangement of small structural motifs leads to the formation of striking supramolecular shapes. Using a combination of single-molecule biophysical techniques and modelling approaches, we dissect the spring-like nature of a designed repeat protein and demonstrate how its shape and mechanics can be manipulated by design. These novel insights into the biomechanical and biochemical characteristics of this protein class give us a methodological basis from which to understand the biological functions of repeat proteins and to exploit them in nanotechnology and biomedicine.
mTORC2 contributes to murine systemic autoimmunity
Authors: Zhou Xian , Qi Haiyu , Li Meilu , Li Yanfeng , Zhu Xingxing , Amin Shreyasee , Alexander Mariam , Diadhiou Catherine , Davidson Anne , Zeng Hu
Published: March 2021

The development of many systemic autoimmune diseases, including systemic lupus erythematosus, is associated with overactivation of the type I interferon (IFN) pathway, lymphopenia, and increased follicular helper T (Tfh) cell differentiation. However, the cellular and molecular mechanisms underlying these immunological perturbations remain incompletely understood. Here we show that the mechanistic target of rapamycin complex 2 (mTORC2) promotes Tfh differentiation and disrupts Treg homeostasis. Inactivation of mTORC2 in total T cells, but not in Tregs, greatly ameliorated the immunopathology in a systemic autoimmunity mouse model. This was associated with reduced Tfh differentiation, B cell activation, and reduced T cell glucose metabolism. Finally, we show that type I IFN can synergize with TCR ligation to activate mTORC2 in T cells, which partially contributes to T cell lymphopenia. These data indicate that mTORC2 may act as downstream of type I IFN, TCR, and costimulatory receptor ICOS, to promote glucose metabolism, Tfh differentiation, and T cell lymphopenia, but not to suppress Treg function in systemic autoimmunity. Our results suggest that mTORC2 might be a rational target for systemic autoimmunity treatment.
Cell trajectory modeling identifies a primitive trophoblast state defined by BCAM enrichment
Authors: Shannon Matthew , Baltayeva Jennet , Castellana Barbara , Wächter Jasmin , Yoon Samantha , Treissman Jenna , Le Hoa T. , Lavoie Pascal M. , Beristain Alexander G.
Published: March 2021

In early placental development, progenitor cytotrophoblasts (CTBs) differentiate along one of two cellular trajectories: the villous or extravillous pathways. CTBs committed to the villous pathway fuse with neighboring CTBs to form the outer multinucleated syncytiotrophoblast (SCT), while CTBs committed to the extravillous pathway differentiate into invasive extravillous trophoblasts (EVT). Unfortunately, little is known about the processes controlling human CTB progenitor maintenance and differentiation. To address this, we established a single cell RNA sequencing (scRNA-seq) dataset from first trimester placentas to identify cell states important in trophoblast progenitor establishment, renewal, and differentiation. Multiple distinct trophoblast states were identified, representing progenitor CTBs, column CTBs, SCT precursors, and EVT. Lineage trajectory analysis identified a progenitor origin that was reproduced in human trophoblast stem cell organoids. Heightened expression of basal cell adhesion molecule (BCAM) defined this primitive state, where BCAM enrichment or gene silencing resulted in enhanced or diminished organoid growth. Together, this work describes at high-resolution trophoblast heterogeneity within the first trimester, resolves gene networks within human CTB progenitors, and identifies BCAM as a primitive progenitor marker and possible regulator.

Summary Statement

Lineage trajectory modeling identifies multiple human progenitor trophoblast states and defines trophoblast differentiation kinetics, where BCAM-expressing progenitors demonstrate enhanced regenerative ability.
A second specificity-determining loop in Class A sortases: Biochemical characterization of natural sequence variation in chimeric SrtA enzymes
Authors: Piper Isabel M. , Struyvenberg Sarah A. , Valgardson Jordan D. , Johnson D. Alex , Gao Melody , Johnston Katherine , Svendsen Justin E. , Kodama Hanna M. , Hvorecny Kelli L. , Antos John M. , Amacher Jeanine F.
Published: March 2021

Gram-positive bacteria contain sortase enzymes on their cell surfaces that catalyze transpeptidation reactions critical for proper cellular function.In vitro, sortases are used in sortase-mediated ligation (SML) reactions for a variety of protein engineering applications. Historically, sortase A fromStaphylococcus aureus(saSrtA) has been the enzyme of choice for SML reactions. However, the stringent specificity of saSrtA for the sequence motif LPXTG limits its uses. Here, we use principal component analysis to identify a structurally conserved loop with a high degree of variability in all classes of sortases. We investigate the contribution of this β7-β8 loop, located between the catalytic cysteine and arginine residues and immediately adjacent to the target binding cleft, by designing and testing chimeric sortase enzymes. Our chimeras utilize natural sequence variation of Class A sortases from 8 species engineered into the SrtA sequence fromStreptococcus pneumoniae(spSrtA). While some of our chimeric enzymes mimic the activity and selectivity of the wild-type protein from which the loop sequence is derived (e.g., that of saSrtA), others result in chimeric spSrtA enzymes able to accommodate a range of residues in the final position of the substrate motif (LPXTX). Using mutagenesis, structural, and sequence analyses, we identify three interactions facilitated by β7-β8 loop residues that appear to be broadly characteristic of Class A sortase enzymes. These studies provide the foundation for a deeper understanding of sortase target selectivity and can expand the sortase toolbox for future SML applications.
Antisense RNAs during early vertebrate development are divided in groups with distinct features
Authors: Pillay Sanjana , Takahashi Hazuki , Carninci Piero , Kanhere Aditi
Published: April 2021

Long noncoding RNAs or lncRNAs are a class of non-protein-coding RNAs that are >200 nt in length. Almost 50% of lncRNAs during zebrafish development are transcribed in an antisense direction to a protein-coding gene. However, the role of these natural antisense transcripts (NATs) during development remains enigmatic. To understand NATs in early vertebrate development, we took a computational biology approach and analyzed existing as well as novel data sets. Our analysis indicates that zebrafish NATs can be divided into two major classes based on their coexpression patterns with respect to the overlapping protein-coding genes. Group 1 NATs have characteristics similar to maternally deposited RNAs in that their levels decrease as development progresses. Group 1 NAT levels are negatively correlated with that of overlapping sense-strand protein-coding genes. Conversely, Group 2 NATs are coexpressed with overlapping protein-coding genes. In contrast to Group 1, which is enriched in genes involved in developmental pathways, Group 2 protein-coding genes are enriched in housekeeping functions. Group 1 NATs also show larger overlap and higher complementarity with the sense-strand mRNAs compared to other NATs. In addition, our transcriptomics data, quantifying RNA levels from cytoplasmic and nuclear compartments, indicates that Group 1 NATs are more abundant in the cytosol. Based on their expression pattern, cytosolic nature, and their higher complementarity to the overlapping developmental mRNAs, we speculate that Group 1 NATs function post-transcriptionally to silence spurious expression of developmental genes.
Building code amendment and building resilience: perspective of building code users in New Zealand
Authors: Nwadike Amarachukwu Nnadozie , Wilkinson Suzanne
Published: March 2021

Purpose

The process followed in amending building code creates problems for code users within the building industry. These problems include the need and frequency of changes made to building code, access to updated documents, method of communication, amendment interval and amendment pathway. This study aims to explore the viewpoints of building code users regarding building code amendments in New Zealand.

Design/methodology/approach

Using a closed-ended questionnaire survey, this paper examined the New Zealand building code amendments by evaluating the views of experienced and relevant stakeholders within the research area.

Findings

A high proportion (50.90%) of the survey participants agreed to a three-years building code amendment cycle, as against the current biannual Amendment practiced in New Zealand. Findings from the study affirmed the necessity for building code amendment and the support for free amended building code documents to the public and other building standards. The study concludes with strong support to the use of intensive research and learning gained from disasters in building code amendment in New Zealand. Implementing the code users opinions encourages disaster resilience through effective application of the building code requirements in design and construction.

Originality/value

The contribution from this study offered a unique insight into the perspectives of building code users on building code amendment in New Zealand and ways of incorporating the findings in the building code later updates to improve disaster resilience in the built environment.
Enabling lean through human capacity building: an investigation of small and medium contractors
Authors: Ranadewa K.A.T.O. , Y.G. Sandanayake Y.G. Sandanayake , Siriwardena Mohan
Published: March 2021

Purpose

This paper investigates the lean enabling human capacities and develops a framework integrating individual, organisational and environmental level strategies to build human capacities for successful lean implementation of small and medium contractors (SMCs) in Sri Lanka.

Design/methodology/approach

An interpretivism stance is adopted, and a qualitative research approach is used. The data collection technique adopted is semi-structured interviews. In total, 24 experts with experience in lean implementation of SMCs were interviewed, and data were analysed through code based content analysis using NVivo10.

Findings

Team working skills, critical thinking, leadership, communication skills, work ethics, knowledge and positive attitudes were identified as lean enabling human capacities for SMCs. The framework developed in this study provides individual, organisational and environmental level strategies that can be used to build human capacities necessary for enabling lean in construction SMCs.

Practical implications

The study will be beneficial to construction SMCs, academics, researchers and government institutions in developing countries, which share socio-economic, demographic or cultural traits similar to Sri Lanka.

Originality/value

A novel lean enabling human capacity building framework is developed with the strategies required for building those capacities in order to accelerate the lean implementation in construction SMCs. This contributes to the body of knowledge as it uncovers individual, organisational and environmental level strategies for enabling lean through human capacity building in Sri Lankan SMCs.