bioRxiv Science⌕ Search

Biology subjects

Wu, C.

Publications and source records attributed to Wu, C..

At least 235 records · Page 13Linked to original sources

ATP Binding Facilitates Target Search of SWR1 Chromatin Remodeler by Promoting One-Dimensional Diffusion on DNA

One-dimensional (1D) target search is a well characterized phenomenon for many DNA binding proteins but is poorly understood for chromatin remodelers. Herein, we characterize the 1D scanning properties of SWR1, a yeast chromatin remodeler that performs histone exchange on +1 nucleosomes adjacent to a nucleosome depleted region (NDR) at promoters. We demonstrate that SWR1 has a kinetic binding preference for DNA of NDR length as opposed to gene-body linker length DNA. Using single and dual color single particle tracking on DNA stretched with optical tweezers, we directly observe SWR1 diffusion on DNA. We found that various factors impact SWR1 scanning, including ATP which promotes diffusion through nucleotide binding rather than ATP hydrolysis. A DNA binding subunit, Swc2, plays an important role in the overall diffusive behavior of the complex, as the subunit in isolation retains similar, although faster, scanning properties as the whole remodeler. ATP-bound SWR1 slides until it encounters a protein roadblock, of which we tested dCas9 and nucleosomes. The median diffusion coefficient, 0.024 m2/sec, in the regime of helical sliding, would mediate rapid encounter of NDR-flanking nucleosomes at length scales found in cells.

biophysics↗

Outbreak.info: A standardized, searchable platform to discover and explore COVID-19 resources and data

To combat the ongoing COVID-19 pandemic, scientists have been conducting research at breakneck speeds, producing over 52,000 peer-reviewed articles within the first year. To address the challenge in tracking the vast amount of new research located in separate repositories, we developed outbreak.info Research Library, a standardized, searchable interface of COVID-19 and SARS-CoV-2 resources. Unifying metadata from sixteen repositories, we assembled a collection of over 350,000 publications, clinical trials, datasets, protocols, and other resources as of October 2022. We used a rigorous schema to enforce consistency across different sources and resource types and linked related resources. Researchers can quickly search the latest research across data repositories, regardless of resource type or repository location, via a search interface, public API, and R package. Finally, we discuss the challenges inherent in combining metadata from scattered and heterogeneous resources and provide recommendations to streamline this process to aid scientific research.

bioinformatics↗

Structures of the Omicron Spike trimer with ACE2 and an anti-Omicron antibody

The Omicron variant of SARS-CoV-2 has rapidly become the dominant infective strain and the focus efforts against the ongoing COVID-19 pandemic. Here we report an extensive set of structures of the Omicron spike trimer by its own or in complex with ACE2 and an anti-Omicron antibody. These structures reveal that most Omicron mutations are located on the surface of the spike protein, which confer stronger ACE2 binding by nearly 10 folds but become inactive epitopes resistant to many therapeutic antibodies. Importantly, both RBD and the closed conformation of the Omicron spike trimer are thermodynamically unstable, with the melting temperature of the Omicron RBD decreased by as much as 7{degrees}C, making the spiker trimer prone to random open conformations. An unusual RBD-RBD interaction in the ACE2-spike complex unique to Omicron is observed to support the open conformation and ACE2 binding, serving the basis for the higher infectivity of Omicron. A broad-spectrum therapeutic antibody JMB2002, which has completed Phase 1 clinical trial, is found to interact with the same two RBDs to inhibit ACE2 binding, in a mode that is distinguished from all previous antibodies, thus providing the structural basis for the potent inhibition of Omicron by this antibody. Together with biochemical data, our structures provide crucial insights into higher infectivity, antibody evasion and inhibition of Omicron.

biophysics↗

Monospecific and bispecific monoclonal SARS-CoV-2 neutralizing antibodies that maintain potency against B.1.617

COVID-19 pathogen SARS-CoV-2 has infected hundreds of millions and caused over 5 million deaths to date. Although multiple vaccines are available, breakthrough infections occur especially by emerging variants. Effective therapeutic options such as monoclonal antibodies (mAbs) are still critical. Here, we report the development, cryo-EM structures, and functional analyses of mAbs that potently neutralize SARS-CoV-2 variants of concern. By high-throughput single cell sequencing of B cells from spike receptor binding domain (RBD) immunized animals, we identified two highly potent SARS-CoV-2 neutralizing mAb clones that have single-digit nanomolar affinity and low-picomolar avidity, and generated a bispecific antibody. Lead antibodies showed strong inhibitory activity against historical SARS-CoV-2 and several emerging variants of concern. We solved several cryo-EM structures at [~]3 [A] resolution of these neutralizing antibodies in complex with prefusion spike trimer ectodomain, and revealed distinct epitopes, binding patterns, and conformations. The lead clones also showed potent efficacy in vivo against authentic SARS-CoV-2 in both prophylactic and therapeutic settings. We also generated and characterized a humanized antibody to facilitate translation and drug development. The humanized clone also has strong potency against both the original virus and the B.1.617.2 Delta variant. These mAbs expand the repertoire of therapeutics against SARS-CoV-2 and emerging variants.

immunology↗

Spontaneous Neuronal Oscillations in the Human Insula are Hierarchically Organized Traveling Waves

The insula plays a fundamental role in a wide range of adaptive human behaviors, but its electrophysiological dynamics are poorly understood. Here we used human intracranial electroencephalographic recordings to investigate the electrophysiological properties and hierarchical organization of spontaneous neuronal oscillations within the insula. We analyzed the neuronal oscillations of the insula directly and found that rhythms in the theta and beta frequency oscillations are widespread and spontaneously present. These oscillations are largely organized along the anterior-posterior axis of the insula. Both the left and right insula showed anterior-to-posterior decreasing gradients for the power of oscillations in the beta frequency band. The left insula also showed a posterior-to-anterior decreasing frequency gradient and an anterior-to-posterior decreasing power gradient in the theta frequency band. In addition to measuring the power of these oscillations, we also examined the phase of these signals across simultaneous recording channels and found that the insula oscillations in the theta and beta bands are traveling waves. The strength of the traveling waves in each frequency was positively correlated with the amplitude of each oscillation. However, the theta and beta traveling waves were uncoupled to each other in terms of phase and amplitude, which suggested that insula traveling waves in the theta and beta bands operate independently. Our findings provide new insights into the spatiotemporal dynamics and hierarchical organization of neuronal oscillations within the insula, which, given its rich connectivity with widespread cortical regions, indicates that oscillations and traveling waves have an important role in intra- and inter-insula communication.

neuroscience↗

NAD metabolism modulates mitochondrial function and inflammation and prevents progression of diabetic kidney disease

Diabetes mellitus is the leading cause of cardiovascular and renal disease in the United States. In spite of the beneficial interventions available for patients with diabetes, there remains a need for additional therapeutic targets and therapies in diabetic kidney disease (DKD). Inflammation and oxidative stress are increasingly recognized as important causes of renal diseases. Inflammation is closely associated with mitochondrial damage. The molecular connection between inflammation and mitochondrial metabolism remains to be elucidated. Recently, nicotinamide adenine nucleotide (NAD+) metabolism has been found to regulate immune function and inflammation. In the present studies we tested the hypothesis that enhancing NAD metabolism could prevent inflammation in and progression of DKD. We found that treatment of db/db mice with type 2 diabetes with nicotinamide riboside (NR) prevented several manifestations of kidney dysfunction (i.e., albuminuria, increased urinary kidney injury marker-1 (KIM1) excretion and pathologic changes). These effects were associated with decreased inflammation, at least in part via inhibiting the activation of the cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) signaling pathway. An antagonist of the serum stimulator of interferon genes (STING) and whole-body STING deletion in diabetic mice showed similar renoprotection. Further analysis found that NR increased SIRT3 activity and improved mitochondrial function, which led to decreased mitochondrial DNA damage, a trigger for mitochondrial DNA leakage which activates the cGAS-STING pathway. Overall, these data show that NR supplementation boosted NAD metabolism to augment mitochondrial function, reducing inflammation and thereby preventing progression of diabetic kidney disease.

pathology↗

Viral genome-capsid core senses host environments to prime RNA release

Viruses are metastable macromolecular assemblies containing a nucleic acid core packaged by capsid proteins that are primed to disassemble in host-specific environments leading to genome release and replication. The mechanism of how viruses sense environmental changes associated with host entry to prime them for disassembly is unknown. We have applied a combination of mass spectrometry, cryo-EM, and simulation-assisted structure refinement to Turnip crinkle virus (TCV), which serves as a model non-enveloped icosahedral virus (Triangulation number = 3, 180 copies/icosahedron). Our results reveal genomic RNA tightly binds a subset of viral coat proteins to form a stable RNA-capsid core which undergoes conformational switching in response to host-specific environmental changes. These changes include: i) Depletion of Ca2+ which triggers viral particle expansion ii) Increase in osmolytes further disrupt interactions of outer coat proteins from the RNA-capsid core to promote complete viral disassembly. A cryo-EM structure of the expanded particle shows that RNA is asymmetrically extruded from a single 5-fold axis during disassembly. The genomic RNA:capsid protein interactions confer metastability to the TCV capsid and drive release of RNA from the disassembling virion within the plant host cell. AUTHOR SUMMARYRNA viruses including coronaviruses, dengue, influenza, and HIV are a significant threat to human health. These viral particles are finely tuned to undergo complex conformational changes that allow for response to varied environments. Turnip crinkle virus (TCV) serves as an excellent model for studying RNA virus dynamics. Since TCV is non-enveloped and has no post-translational modifications, we can specifically investigate the contributions of RNA to viral dynamics. Genomic RNA is not a passive entity but plays a crucial and previously uncharacterized role in viral disassembly. Our results reveal that the genomic RNA-capsid core serves as an environmental sensor and undergoes conformational switching in response to host cell conditions.

plant biology↗

SARS-CoV-2 RdRp is a versatile enzyme with proofreading activity and ability to incorporate NHC into RNA by using diphosphate form molnupiravir as a substrate

The coronavirus disease 2019 (COVID-19) has been ravaging throughout the world for more than two years and has severely impaired both human health and the economy. The causative agent, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) employs the viral RNA-dependent RNA polymerase (RdRp) complex for genome replication and transcription, making RdRp an appealing target for antiviral drug development. Here, we reveal that RdRp can recognize and utilize nucleoside diphosphates (NDPs) as a substrate to synthesize RNAs with an efficiency of about two thirds of using nucleoside triphosphates (NTPs) as a substrate. NDPs incorporation is also template-specific and has high fidelity. Moreover, RdRp can incorporate {beta}-d-N4-hydroxycytidine (NHC) into RNA while using diphosphate form molnupiravir (MDP) as a substrate. We also observed that MDP is a better substrate for RdRp than the triphosphate form molnupiravir (MTP).

molecular biology↗

Low frequency somatic copy number alterations in normal human lymphocytes revealed by large scale single-cell whole genome profiling

Genomic-scale somatic copy number alterations in healthy humans are difficult to investigate because of low occurrence rates and the structural variations stochastic natures. Using a Tn5-transposase assisted single-cell whole genome sequencing method, we sequenced over 20,000 single lymphocytes from 16 individuals. Then, with the scale increased to a few thousand single cells per individual, we found that about 7.5% of the cells had large-size copy number alterations. Trisomy 21 was the most prevalent aneuploid event among all autosomal copy number alterations, while monosomy X occurred most frequently in over-30-year-old females. In the monosomy X single cells from individuals with phased genomes and identified X-inactivation ratios in bulk, the inactive X Chromosomes were lost more often than were the active ones.

genomics↗

Generation and network analysis of an RNA-seq transcriptional atlas for the rat.

The laboratory rat is an important model for biomedical research. To generate a comprehensive rat transcriptomic atlas, we curated and down-loaded 7700 rat RNA-seq datasets from public repositories, down-sampled them to a common depth and quantified expression. Data from 590 rat tissues and cells, averaged from each Bioproject, can be visualised and queried at http://biogps.org/ratatlas. Gene correlation network (GCN) analysis revealed clusters of transcripts that were tissue or cell-type restricted and contained transcription factors implicated in lineage determination. Other clusters were enriched for transcripts associated with biological processes. Many of these clusters overlap with previous data from analysis of other species whilst some (e.g. expressed specifically in immune cells, retina/pineal gland, pituitary and germ cells) are unique to these data. GCN on large subsets of the data related specifically to liver, nervous system, kidney, musculoskeletal system and cardiovascular system enabled deconvolution of cell-type specific signatures. The approach is extensible and the dataset can be used as a point of reference from which to analyse the transcriptomes of cell types and tissues that have not yet been sampled. Sets of strictly co-expressed transcripts provide a resource for critical interpretation of single cell RNA-seq data.

genomics↗

Polarization and cell-fate decision facilitated by the adaptor Ste50 in Saccharomyces cerevisiae

In response to pheromone, many proteins localize on the plasma membrane of yeast cell to reform it into a polarized shmoo structure. The adaptor protein Ste50, long known as a pheromone signal enhancer critical for shmoo polarization, has never been explored systematically for its localization and function in the polarization process. Time-lapse single-cell imaging and quantitation shown here characterizes Ste50p involvement in the establishment of cell polarity. We found early localization of Ste50p patches on the cell cortex that mark the point of shmoo initiation, these polarity sites move, and patches remain associated with the growing shmoo tip in a pheromone concentration timedependent manner until shmoo maturation. By quantitative analysis we show that polarization corelates with the rising levels of Ste50p, enabling rapid individual cell responses to pheromone that corresponds to a critical level of Ste50p at the initial G1 phase. We exploited the quantitative differences in the pattern of Ste50p expression to corelate with the cell-cell phenotypic heterogeneity showing Ste50p involvement in the cellular differentiation choice. Taken together, these findings present spatiotemporal localization of Ste50p during yeast polarization, suggesting that Ste50p is a component of the polarisome, and plays a critical role in regulating the polarized growth of shmoo during pheromone response.

cell biology↗

Kinetic principles underlying pioneer function of GAGA transcription factor in live cells

How pioneer factors interface with chromatin to promote accessibility for transcription control is poorly understood in vivo. Here, we directly visualize chromatin association by the prototypical GAGA pioneer factor (GAF) in live Drosophila hemocytes. Single-particle tracking reveals that the majority of GAF is chromatin-bound, with a stable-binding fraction showing nucleosome-like confinement residing on chromatin for over 2 minutes, far longer than the dynamic range of most transcription factors. These kinetic properties require the full complement of GAFs DNA-binding, multimerization and intrinsically disordered domains, and are autonomous from recruited chromatin remodelers NURF and PBAP, whose activities primarily benefit GAFs neighbors such as HSF. Evaluation of GAF kinetics together with its endogenous abundance indicates that despite on-off dynamics, GAF constitutively and fully occupies chromatin targets, thereby providing a temporal mechanism that sustains open chromatin for transcriptional responses to homeostatic, environmental, and developmental signals.

molecular biology↗

Coordinated DNA and Histone Dynamics Drive Accurate Histone H2A.Z Exchange

Nucleosomal histone H2A is exchanged for its variant H2A.Z by the SWR1 chromatin remodeler, but the mechanism and timing of histone exchange remain unclear. Here, we quantify DNA and histone dynamics during histone exchange in real-time using a three-color single-molecule FRET assay. We show that SWR1 operates with timed precision to unwrap DNA with large displacement from one face of the nucleosome, remove H2A-H2B from the same face, and rewrap DNA, all within 2.3 seconds. Such productive DNA unwrapping requires full SWR1 activation and differs from unproductive, smaller-scale DNA unwrapping caused by SWR1 binding alone. On an asymmetrically positioned nucleosome, SWR1 intrinsically senses long-linker DNA to preferentially exchange H2A.Z on the distal face as observed in vivo. The displaced H2A-H2B dimer remains briefly associated with the SWR1-nucleosome complex and is dissociated by histone chaperones. These findings reveal how SWR1 coordinates DNA unwrapping with histone dynamics to rapidly and accurately place H2A.Z at physiological sites on chromatin. One-Sentence SummaryMulticolor single-molecule FRET reveals how SWR1 unwraps DNA to exchange nucleosomal H2A-H2B for H2A.Z-H2B.

biophysics↗

BioThings SDK: a toolkit for building high-performance data APIs in biomedical research

SummaryTo meet the increased need of making biomedical resources more accessible and reusable, Web APIs or web services have become a common way to disseminate knowledge sources. The BioThings APIs are a collection of high-performance, scalable, annotation as a service APIs that automate the integration of biological annotations from disparate data sources. This collection of APIs currently includes MyGene.info, MyVariant.info, and MyChem.info for integrating annotations on genes, variants, and chemical compounds, respectively. These APIs are used by both individual researchers and application developers to simplify the process of annotation retrieval and identifier mapping. Here, we describe the BioThings Software Development Kit (SDK), a generalizable and reusable toolkit for integrating data from multiple disparate data sources and creating high-performance APIs. This toolkit allows users to easily create their own BioThings APIs for any data type of interest to them, as well as keep APIs up-to-date with their underlying data sources. Availability and implementationThe BioThings SDK is built in Python and released via PyPI (https://pypi.org/project/biothings/). Its source code is hosted at its github repository (https://github.com/biothings/biothings.api).

bioinformatics↗

Cellular perception of growth rate and the mechanistic origin of bacterial growth laws

Bacteria organize many activities according to their grow rate via the ppGpp signaling system. Yet it is not clear how this signaling system "knows" how fast cells grow. Through quantitative experiments, we show that ppGpp responds inversely to the rate of translational elongation in E. coli. Together with its roles in inhibiting ribosome biogenesis and activity, ppGpp closes a key regulatory circuit that enables the cell to perceive and control the rate of its growth across conditions. The celebrated linear growth law relating the ribosome content and growth rate emerges as a consequence of keeping a supply of ribosome reserves while maintaining elongation rate in slow growth conditions. Further analysis suggests the elongation rate itself is detected by sensing the ratio of dwelling and translocating ribosomes, a strategy employed to collapse the complex, high-dimensional dynamics of the molecular processes underlying cell growth to perceive the physiological state of the whole.

systems biology↗

Regulation of drought tolerance in Arabidopsis involves PLATZ4-mediated transcriptional suppression of PIP2;8

PLATZ transcription factors play important roles in plant growth, development, and biotic and abiotic stress responses. However, how PLATZ regulates plant drought tolerance and ABA sensitivity remains largely unknown. Here, we show that PLATZ4 increases drought tolerance and ABA sensitivity in Arabidopsis thaliana by suppressing the expression of PIP2;8, DEG3, At1g33840 and At3g01345, while upregulating the expression of PR1, ABI3, ABI4 and ABI5. Moreover, PLATZ4 directly binds A/T-rich sequences within the PIP2;8 promoter but not ABI3, ABI4 and ABI5 promoters. Consistent with this, PIP2;8 acts epistatically to PLATZ4. Furthermore, the aquaporin activity of PIP2;8 was confirmed in Xenopus laevis oocytes and yeast cells in response to osmotic stress. Analysis of water loss of seedlings overexpressing PIP2;8 or lacking PIP2;8 function indicated that PIP2;8-mediated water flow is particularly active in response to drought stress in planta. Accordingly, stomatal closure was enhanced in pip2;8 mutants but reduced in PIP2;8-overexpressing plants. In platz4 mutant and PLATZ4-overexpressing plants, water loss and stomatal closure changed oppositely to those in pip2;8 mutants and PIP2;8-overexpressing plants, respectively. In addition, the interaction between PLATZ4 and AITR6 was confirmed by several assays, and the binding of PIP2;8 promoter by PLATZ4 was strengthened by an interaction with AITR6. Collectively, our findings reveal that PLATZ4 interacts with AITR6 to increase ABA sensitivity and drought tolerance in Arabidopsis by upregulating expression of PR1, ABI3, ABI4 and ABI5 while inhibiting the expression of PIP2;8 and associated genes. One sentence summaryPLATZ4 interacted with AITR6 to increase drought tolerance and ABA sensitivity in Arabidopsis thaliana by directly bind A/T-rich sequences within the plasma membrane aquaporin (PIP)2;8 promoter.

plant biology↗

Single-cell transcriptomics reveals the identity and regulators of human mast cell progenitors

Mast cell accumulation is a hallmark of a number of diseases including allergic asthma and systemic mastocytosis. IgE-mediated crosslinking of the Fc{varepsilon}RI receptors causes mast cell activation and contributes to disease pathogenesis. The mast cell lineage is one of the least studied among the hematopoietic cell lineages and there are still controversies about the identity of the mast cell progenitor, i.e., whether Fc{varepsilon}RI expression appears during the hematopoietic progenitor stage or in maturing mast cells. Here, we used single-cell transcriptomics to reveal a temporal association between the appearance of Fc{varepsilon}RI and the mast cell gene signature in CD34+ hematopoietic progenitors. In agreement with these data, the Fc{varepsilon}RI+ hematopoietic progenitors formed morphologically, phenotypically and functionally mature mast cells in long-term culture assays. Single-cell transcriptomics analysis further revealed the expression patterns of prospective cytokine receptors regulating mast cell progenitor development. Culture assays showed that IL-3 and IL-5 promoted disparate effects on progenitor cell proliferation and survival, respectively, whereas IL-33 caused robust Fc{varepsilon}RI downregulation. Taken together, we have demonstrated that Fc{varepsilon}RI appears during the hematopoietic progenitor stage of mast cell differentiation and that external stimuli may regulate the Fc{varepsilon}RI expression. Thus, the results resolve the controversy regarding the appearance of Fc{varepsilon}RI during mast cell development. One-sentence summarySingle-cell analysis of human hematopoiesis uncovers the stage at which Fc{varepsilon}RI appears during mast cell differentiation and reveals disparate effects of IL-3, IL-5 and IL-33 on mast cell progenitor proliferation, survival, and suppression of Fc{varepsilon}RI expression.

cell biology↗

Dcp2 C-terminal Cis-Binding Elements Control Selective Targeting of the Decapping Enzyme by Forming Distinct Decapping Complexes

A single Dcp1-Dcp2 decapping enzyme targets diverse classes of yeast mRNAs for decapping-dependent 5 to 3 decay, but the molecular mechanisms controlling selective mRNA targeting by the enzyme remain elusive. Through extensive genetic analyses we uncover cis-regulatory elements in the Dcp2 C-terminal domain that control selective targeting of the decapping enzyme by forming distinct decapping complexes. Two Upf1-binding motifs target the decapping enzyme to NMD substrates, and a single Edc3-binding motif targets both Edc3 and Dhh1 substrates. Pat1-binding leucine-rich motifs target Edc3 and Dhh1 substrates under selective conditions. Although it functions as a unique targeting component of specific complexes, Edc3 is a common component of multiple complexes. Xrn1 also has a specific Dcp2 binding site, allowing it to be directly recruited to decapping complexes. Collectively, our results demonstrate that Upf1, Edc3, and Pat1 function as regulatory subunits of the holo-decapping enzyme, controlling both its targeting specificity and enzymatic activation. HighlightsLoss of Dcp2 cis-binding elements causes selective stabilization of distinct decapping substrates Dcp2 cis-binding elements promote the assembly of target-specific decapping complexes in vivo Xrn1 binds to Dcp2, and both Edc3 and Xrn1 are common components of multiple decapping complexes Upf1, Edc3, and Pat1 function as unique targeting subunits of the yeast holo-decapping enzyme

molecular biology↗