Biology subjects

Yu, Z.

Publications and source records attributed to Yu, Z..

At least 235 records · Page 13Linked to original sources

bioRxiv2022

Structure of the human UBR5 E3 ubiquitin ligase

The human UBR5 (also known as EDD) is a single polypeptide chain HECT-type E3 ubiquitin ligase essential for embryonic development in mammals. Although widely expressed, UBR5 is markedly amplified and overexpressed in breast, ovarian, prostate, gastric and pancreatic cancers. Dysregulated UBR5 functions like an oncoprotein to promote cancer growth and metastasis, making UBR5 a potential target for therapeutics. Unexpectedly, we found that human UBR5 assembles a dimer and a tetramer in solution. We determined the dimer structure at 2.8 [A] and the tetramer structure at 3.5 [A] average resolution. UBR5 is a crescent shaped molecule with a seven-bladed {beta}-propeller and two small {beta}-barrel domains (SBB1/2) at the N-terminal region, a catalytic HECT domain at the C-terminus, and an extended helical scaffold and an N-degron-recognizing UBR box in the middle. The dimer is assembled as a stable head-to-tail dimer via extensive interactions in the middle helical scaffold region. The tetramer is assembled via SBB2-SBB2 interaction from two face-to-face dimers, forming a large cage with all four catalytic HECT domains facing the central cavity. Importantly, the N-terminal region of one subunit and the HECT of the other form an "intermolecular jaw" in the dimer. Using enzymatic and cellular assays, we showed that the jaw-lining residues are important for function, suggesting that the intermolar jaw functions to recruit ubiquitin loaded E2 to UBR5 for the transthiolation reaction. Further work is needed to understand how oligomerization regulates the UBR5 ligase activity. This work provides a framework for structure-based anticancer drug development against the distinctive HECT E3 ligase and contributes to a growing appreciation of E3 ligase diversity.

biochemistry↗

bioRxiv2022

De novo protein identification in mammalian sperm using high-resolution in situ cryo-electron tomography

Understanding molecular mechanisms of cellular pathways requires knowledge of the identities of participating proteins, their cellular localization and their 3D structures. Contemporary workflows typically require multiple techniques to identify target proteins, track their localization using fluorescence microscopy, followed by in vitro structure determination. To identify mammal-specific sperm proteins and understand their functions, we developed a visual proteomics workflow to directly address these challenges. Our in situ cryo-electron tomography and subtomogram averaging provided 6.0 [A] resolution reconstructions of axonemal microtubules and their associated proteins. The well-resolved secondary and tertiary structures allowed us to computationally match, in an unbiased manner, novel densities in our 3D reconstruction maps with 21,615 AlphaFold2-predicted protein models of the mouse proteome. We identified Tektin 5, CCDC105 and SPACA9 as novel microtubule inner proteins that form an extensive network crosslinking the lumen of microtubule and existing proteins. Additional biochemical and mass spectrometry analyses helped validate potential candidates. The novel axonemal sperm structures identified by this approach form an extensive interaction network within the lumen of microtubules, suggesting they have a role in the mechanical and elastic properties of the microtubule filaments required for the vigorous beating motions of flagella.

biophysics↗

bioRxiv2022

Low Molecular Weight Hyaluronan Inhibits Lung Epithelial Ion Channels by Activating Calcium-Sensing Receptor

Herein, we tested the hypothesis low molecular weight hyaluronan (LMW-HA) inhibits lung epithelial ion transport in-vivo, ex-vivo, and in-vitro by activating the calcium-sensing receptor (CaSR). Intranasal instillation of LMW-HA (150g/ml) to C57BL/6 mice inhibited their alveolar fluid clearance (AFC) by 75%, increased the epithelial lining fluid (ELF) thickness threefold, and lung wet/dry (W/D) ratio by 20% 24hrs later. Incubation of lung slices from mouse and human lungs with 150g/ml LMW-HA decreased the open probability (Po) of ENaC in ATII cell by more than 50% in 4hrs, inhibited amiloride sensitive short circuit current (SCC) 4hrs post exposure, and Cl- current through CFTR by more than 70%, and Na,K-ATPase current by 66% at 24hrs. In all cases the inhibitory effect of LMW-HA on lung epithelial ion transport in vivo, ex vivo, and in vitro preparations were reversed by the administration of 1M of NPS2143, a CaSR inhibitor, or 150g/ml HMW-HA. In HEK-293 cells co-transfected with CaSR and the calcium sensitive Cl- channel TMEM16-A, LMW-HA activated an inward Cl- current. These data are the first demonstration of the inhibitory effects of LMW-HA on lung epithelial ion and water transport, and are due to the activation of CaSR and its downstream signaling cascades.

physiology↗

bioRxiv2022

Heterologous boost with mRNA vaccines against SARS-CoV-2 Delta/Omicron variants following an inactivated whole-virus vaccine

The coronavirus SARS-CoV-2 has mutated quickly and caused significant global damage. This study characterizes two mRNA vaccines ZSVG-02 (Delta) and ZSVG-02-O (Omicron BA.1), and associating heterologous prime-boost strategy following the prime of a most widely administrated inactivated whole-virus vaccine (BBIBP-CorV). The ZSVG-02-O induces neutralizing antibodies that effectively cross-react with Omicron subvariants following an order of BA.1>BA.2>BA.4/5. In naive animals, ZSVG-02 or ZSVG-02-O induce humoral responses skewed to the vaccines targeting strains, but cellular immune responses cross-react to all variants of concern (VOCs) tested. Following heterologous prime-boost regimes, animals present comparable neutralizing antibody levels and superior protection across all VOCs. Single-boost only generated ancestral and omicron dual-responsive antibodies, probably by "recall" and "reshape" the prime immunity. New Omicron-specific antibody populations, however, appeared only following the second boost with ZSVG-02-O. Overall, our results support a heterologous boost with ZSVG-02-O, providing the best protection against current VOCs in inactivated virus vaccine- primed populations.

molecular biology↗

bioRxiv2022

The SARS-CoV-2 accessory protein Orf3a is not an ion channel, but does interact with trafficking proteins

The severe acute respiratory syndrome associated coronavirus 2 (SARS-CoV-2) and SARS-CoV-1 accessory protein Orf3a colocalizes with markers of the plasma membrane, endocytic pathway, and Golgi apparatus. Some reports have led to annotation of both Orf3a proteins as a viroporin. Here we show that neither SARS-CoV-2 nor SARS-CoV-1 form functional ion conducting pores and that the conductances measured are common contaminants in overexpression and with high levels of protein in reconstitution studies. Cryo-EM structures of both SARS-CoV-2 and SARS-CoV-1 Orf3a display a narrow constriction and the presence of a basic aqueous vestibule, which would not favor cation permeation. We observe enrichment of the late endosomal marker Rab7 upon SARS-CoV-2 Orf3a overexpression, and co-immunoprecipitation with VPS39. Interestingly, SARS-CoV-1 Orf3a does not cause the same cellular phenotype as SARS-CoV-2 Orf3a and does not interact with VPS39. To explain this difference, we find that a divergent, unstructured loop of SARS-CoV-2 Orf3a facilitates its binding with VPS39, a HOPS complex tethering protein involved in late endosome and autophagosome fusion with lysosomes. We suggest that the added loop enhances SARS-CoV-2 Orf3a ability to co-opt host cellular trafficking mechanisms for viral exit or host immune evasion.

biophysics↗

bioRxiv2022

KMer-Node2Vec: Learning Vector Representations of K-mers from the K-meGraph

Learning low-dimensional continuous vector representation for short k-mers divided from long DNA sequences is key to DNA sequence modeling that can be utilized in many bioinformatics investigations, such as DNA sequence classification and retrieval. DNA2Vec is the most widely used method for DNA sequence embedding. However, it poorly scales to large data sets due to its extremely long training time in kmer embedding. In this paper, we propose a novel efficient graph-based kmer embedding method, named Kmer-Node2Vec, to tackle this concern. Our method converts the large DNA corpus into one kmer co-occurrence graph and extracts kmer relation on the graph by random walks to learn fast and high-quality kmer embedding. Extensive experiments show that our method is faster than DNA2Vec by 29 times for training on a 4GB data set, and on par with DNA2Vec in terms of task-specific accuracy of sequence retrieval and classification.

bioinformatics↗

bioRxiv2022

TMEM100, a Lung-Specific Endothelium Gene

The heterogeneity of endothelium across different organs was recently explored using single-cell RNA-sequencing analysis. Compared to other organs, the lung exhibits a distinct structure composed of a thin layer of capillary for efficient gas exchange. In this study, we demonstrate that Tmem100 is a lung-specific endothelium gene.

physiology↗

bioRxiv2022

Fat Extract Modulates Calcium Signaling and Protects Against Hyperactive Osteoclastogenesis in Bone Remodeling with Antioxidant Capacity

Osteoclasts are cells which are primarily involved in bone remodeling and osteolytic bone diseases. Hyperactive osteoclastogenesis leads to pathological bone loss and microarchitectural deterioration, particularly in postmenopausal osteoporosis. However, given the limitations of current first-line osteoclast inhibitors, there is an urgent need for a novel antiresorptive agent with higher efficiency and fewer side effects. Cell-free fat extract (CEFFE) is the liquid fraction obtained from human adipose tissues, which are enriched with a variety of cytokines and growth factors. This study aims to explore its pharmaceutical effect on hyperactive osteoclast formation in vivo and in vitro. CEFFE exhibits excellent potentials to attenuate osteoclast-associated bone loss in an ovariectomy (OVX) mouse model and to inhibit RANKL-induced osteoclastogenesis in primary bone marrow-derived monocytes. Furthermore, the cationic protein fraction of CEFFE (CEFFE-Cation) is identified as the main inhibitory component in osteoclast formation assay. Excessive reactive oxygen species (ROS) production is the main cause of osteoclast overactivation. According to LC-MS/MS analysis, the CEFFE-Cation fraction mainly consists of various antioxidant enzymes, extracellular matrix, and secreted cytokines, which endow it with a superior antioxidant capacity. Ca2+ signaling contributes to osteoclast maturation. In addition to scavenging ROS, CEFFE-Cation is also capable of mitigating Ca2+ oscillation, calcineurin activation, and subsequent NFATc1 nuclear translocation during osteoclastogenesis. Overall, this study elucidates the promising translational potential of CEFFE as a next-generation personalized antiresorptive agent for osteolytic bone disease treatment.

pathology↗

bioRxiv2022

APC and P53 mutations synergize to create a therapeutic vulnerability to NOTUM inhibition in advanced colorectal cancer

Colorectal cancer (CRC) is a leading cause of cancer-related deaths globally, with the majority of cases initiated by inactivation of the APC tumor suppressor. This results in the constitutive transcriptional activation of the canonical WNT signal transduction pathway effector {beta}-Catenin, along with induction of WNT feedback inhibitors, including the extracellular palmitoleoyl-protein carboxylesterase NOTUM. Here, we show that NOTUM retains cell-autonomous tumor suppressive activity in APC-null adenomatous lesions despite constitutive {beta}-Catenin activation. Strikingly, we find that NOTUM becomes an obligate oncogene upon subsequent P53 inactivation during the adenoma-adenocarcinoma transition, and that these phenotypes are WNT-independent, resulting from differential activity of NOTUM upon its enzymatic targets Glypican 1 and 4 in early vs. late-stage disease, respectively. Ultimately, preclinical mouse models of CRC and human tumoroid cultures demonstrate that pharmacological inhibition of NOTUM is highly effective in arresting primary adenocarcinoma growth and inhibiting metastatic colonization of distal organs. The finding that a single agent targeting an extracellular enzyme is effective in treating highly aggressive tumors make NOTUM a novel therapeutic vulnerability in advanced colorectal adenocarcinomas.

cancer biology↗

bioRxiv2022

Kinetic and Thermodynamic Insights on Agonist Interactions with the Parathyroid Hormone Receptor-1 from a New NanoBRET assay

Polypeptides that activate the parathyroid hormone receptor-1 (PTHR1) are important in human physiology and medicine. Most previous studies of peptide binding to this receptor have involved displacement of a radiolabeled ligand. We report a new assay format based on bioluminescence resonance energy transfer (BRET). Fusion of a nanoluciferase (nLuc) unit to the N-terminus of the PTHR1 allows direct detection of binding by an agonist peptide bearing a tetramethylrhodamine (TMR) unit. Affinity measurements from the BRET assay align well with results previously obtained via radioligand displacement. The BRET assay offers substantial operational benefits relative to affinity measurements involving radioactive compounds. The convenience of the new assay allowed us to explore several questions raised by earlier reports. For example, we show that although the first two residues of PTH(1-34) (the drug teriparatide) are critical for PTHR1 activation, these two residues contribute little or nothing to affinity. Comparisons among the well-studied agonists PTH(1-34), PTHrP(1-34) and "long-acting PTH" (LA-PTH) reveal that the high affinity of LA-PTH arises largely from a diminished rate constant for dissociation relative to the other two.

biochemistry↗

bioRxiv2022

The metabolic influence of the core ciliate Entodinium caudatum within the rumen microbiome

Protozoa comprise a major fraction of the microbial biomass in the rumen microbiome, of which the entodiniomorphs (order: Entodiniomorphida) and holotrichs (order: Vestibuliferida) are consistently observed to be dominant across a diverse genetic and geographical range of ruminant hosts. Despite the apparent core role that protozoal species exert, their major biological and metabolic contributions to rumen function remain largely undescribed in vivo. Here, we have leveraged (meta)genome-centric metaproteomes from rumen fluid samples originating from both cattle and goats fed diets with varying inclusion levels of lipids and starch, to detail the specific metabolic niches that protozoa occupy in the context of their microbial co-habitants. Initial proteome estimations via total protein counts and label-free quantification highlight that entodiniomorph species Entodinium and Epidinium as well as the holotrichs Dasytricha and Isotricha comprise an extensive fraction of the total rumen metaproteome. Proteomic detection of protozoal metabolism such as hydrogenases (Dasytricha, Isotricha, Epidinium, Enoploplastron), carbohydrate-active enzymes (Epidinium, Diplodinium, Enoploplastron, Polyplastron), microbial predation (Entodinium) and volatile fatty acid production (Entodinium and Epidinium) was observed at increased levels in high methane-emitting animals. Despite certain protozoal species having well-established reputations for digesting starch, they were unexpectedly less detectable in low methane emitting-animals fed high starch diets, which were instead dominated by propionate/succinate-producing bacterial populations suspected of being resistant to predation irrespective of host. Finally, we reaffirmed our abovementioned observations in geographically independent datasets, thus illuminating the substantial metabolic influence that under-explored eukaryotic populations have in the rumen, with greater implications for both digestion and methane metabolism.

microbiology↗

bioRxiv2022

Pan-genome inversion index reveals evolutionary insights into the subpopulation structure of Asian rice (Oryza sativa)

Understanding and exploiting genetic diversity is a key factor for the productive and stable production of rice. Utilizing 16 high-quality genomes that represent the subpopulation structure of Asian rice (O. sativa), plus the genomes of two close relatives (O. rufipogon and O. punctata), we built a pan-genome inversion index of 1,054 non-redundant inversions that span an average of [~] 14% of the O. sativa cv. Nipponbare reference genome sequence. Using this index we estimated an inversion rate of 1,100 inversions per million years in Asian rice, which is 37 to 73 times higher than previously estimated for plants. Detailed analyses of these inversions showed evidence of their effects on gene regulation, recombination rate, linkage disequilibrium and agronomic trait performance. Our study uncovers the prevalence and scale of large inversions ([≥] 100 kb) across the pan-genome of Asian rice, and hints at their largely unexplored role in functional biology and crop performance.

genomics↗

bioRxiv2022

Multivalency, autoinhibition, and protein disorder in the regulation of interactions of dynein intermediate chain with dynactin and the nuclear distribution protein

Cytoplasmic dynein plays crucial roles in the intracellular transport of organelles and other cargoes. Central to dynein function is the intrinsically disordered N-terminal domain of dynein intermediate chain (IC), which binds the three dimeric dynein light chains at multivalent sites, and dynactin p150Glued and nuclear distribution protein (NudE) at overlapping sites. The disorder in IC has hindered cryo-electron microscopy and X-ray crystallography studies of its structure and interactions. Here we use a suite of biophysical methods to reveal how multivalent binding of the three light chains regulate IC interactions with p150Glued and NudE. Using the N-terminal domain or the full-length IC from Chaetomium thermophilum, a tractable species to interrogate IC interactions, we identify a significant reduction in ICs binding affinity for p150Glued and a loss of binding to NudE in contrast to the tight binding observed with small IC constructs. We attribute this difference to autoinhibition caused by strong long-range intramolecular interactions that cover ICs N-terminal single -helix, the site for p150Glued and NudE binding. Reconstitution of IC subcomplexes demonstrate that autoinhibition is differentially regulated by light chains binding underscoring their importance both in assembly and organization of IC, and in selection between multiple binding partners at the same site.

biophysics↗

bioRxiv2022

Chemical genetic screening identifies nalacin as an inhibitor of GH3 amido synthetase for auxin conjugation

Root system is critical for plant growth and development. To develop plant growth regulators functioning in root development, we performed a phenotype-based chemical screen in Arabidopsis and identified a chemical, nalacin, that mimicked the effects of auxin on root development. Genetic, pharmacological and biochemical approaches demonstrated that nalacin exerts its auxin-like activities by inhibiting indole-3-acetic acid (IAA) conjugation that is mediated by Gretchen Hagen 3 (GH3) acyl acid amido synthetases. The crystal structure of Arabidopsis GH3.6 in complex with D4 (a derivative of nalacin) together with docking simulation analysis revealed the molecular basis of the inhibition of group II GH3 by nalacin. Sequence alignment analysis indicated broad bioactivities of nalacin and D4 as inhibitors of GH3s in vascular plants, which were confirmed, at least, in tomato and rice. In summary, our work identifies nalacin as a potent inhibitor of IAA conjugation mediated by group II GH3 that plays versatile roles in hormone-regulated plant development and has potential applications in both basic research and agriculture.

plant biology↗

bioRxiv2022

BEN: a generalizable Brain Extraction Net for multimodal MRI data from rodents, nonhuman primates, and humans

Accurate brain tissue extraction on magnetic resonance imaging (MRI) data is crucial for analyzing brain structure and function. While several conventional tools have been optimized to handle human brain data, there have been no generalizable methods to extract brain tissues for multimodal MRI data from rodents, nonhuman primates, and humans. Therefore, developing a flexible and generalizable method for extracting whole brain tissue across species would allow researchers to analyze and compare experiment results more efficiently. Here, we propose a domain-adaptive and semi-supervised deep neural network, named the Brain Extraction Net (BEN), to extract brain tissues across species, MRI modalities, and MR scanners. We have evaluated BEN on 18 independent datasets, including 783 rodent MRI scans, 246 nonhuman primate MRI scans, and 4,601 human MRI scans, covering five species, four modalities, and six MR scanners with various magnetic field strengths. Compared to conventional toolboxes, the superiority of BEN is illustrated by its robustness, accuracy, and generalizability. Our proposed method not only provides a generalized solution for extracting brain tissue across species but also significantly improves the accuracy of atlas registration, thereby benefiting the downstream processing tasks. As a novel fully automated deep-learning method, BEN is designed as an open-source software to enable high-throughput processing of neuroimaging data across species in preclinical and clinical applications.

neuroscience↗

bioRxiv2022

Greater alteration of gut microbiota occurs in childhood obesity than adulthood obesity

The childrens gut microbiota, associated with the development of obesity, is in maturation. The impact of obesity on the gut microbiota in childhood could have a more significant effect than on adulthood and eventually be lifelong lasting, but it has been rarely studied. Aimed to discover the difference in gut microbiota between obese children and adults, we collected and re-analyzed published data from National Center for Biotechnology Information (NCBI). We found that the gut microbiota had a significant difference between children with and without obesity, but this did not happen in adults. Based on the random forest model, childhood obesity is more challenging to predict using gut microbiota than adulthood obesity. Our results suggest that gut microbiota in childhood is more easily affected than in adulthood. Hence, early intervention for childhood obesity is essential to improve childrens health and lifelong gut microbiota-related health.

microbiology↗

bioRxiv2022

Structure of nucleosome-bound human PBAF complex

BAF and PBAF are mammalian SWI/SNF family chromatin remodeling complexes that possess multiple histone/DNA-binding subunits and create nucleosome-depleted/free regions for transcription activation. Despite structural studies of nucleosome-bound human BAF and yeast SWI/SNF family complexes, it remains elusive how PBAF-nucleosome complex is organized. Here we determined structure of 13-subunit human PBAF in complex with acetylated nucleosome in ADP-BeF3-bound state. Four PBAF-specific subunits work together with nine BAF/PBAF-shared subunits to generate PBAF-specific modular organization, distinct from that of BAF at various regions. PBAF-nucleosome structure reveals six histone-binding domains and four DNA-binding domains/modules, the majority of which directly bind histone/DNA. This multivalent nucleosome-binding pattern, not observed in previous studies, suggests that PBAF may integrate comprehensive chromatin information to target genomic loci for function. Our study reveals molecular organization of subunits and histone/DNA-binding domains/modules in PBAF-nucleosome complex and provides a framework to understand chromatin targeting of SWI/SNF family complexes.

molecular biology↗

bioRxiv2022

In situ cryo-electron tomography reveals the asymmetric architecture of mammalian sperm axonemes

The flagella of mammalian sperm display non-planar, asymmetric beating, in contrast to the planar, symmetric beating of flagella from sea urchin sperm and unicellular organisms. The molecular basis of this difference is unclear. Here, we perform in situ cryo-electron tomography of mouse and human sperm axonemes, providing the highest resolution structural information to date. Our subtomogram averages reveal mammalian sperm- specific protein complexes within the outer microtubule doublets, the radial spokes and nexin-dynein regulatory complexes. The locations and structures of these complexes suggest potential roles in enhancing the mechanical strength of mammalian sperm axonemes and regulating dynein-based axonemal bending. Intriguingly, we find that each of the nine outer microtubule doublets is decorated with a distinct combination of sperm- specific complexes. We propose that this asymmetric distribution of proteins differentially regulates the sliding of each microtubule doublet and may underlie the asymmetric beating of mammalian sperm.

biophysics↗