Publications by authors named "Eric Xu"

222 Publications

Structural basis for inhibition of the SARS-CoV-2 RNA polymerase by suramin.

Nat Struct Mol Biol 2021 Mar 5. Epub 2021 Mar 5.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China.

The COVID-19 pandemic caused by nonstop infections of SARS-CoV-2 has continued to ravage many countries worldwide. Here we report that suramin, a 100-year-old drug, is a potent inhibitor of the SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) and acts by blocking the binding of RNA to the enzyme. In biochemical assays, suramin and its derivatives are at least 20-fold more potent than remdesivir, the currently approved nucleotide drug for treatment of COVID-19. The 2.6 Å cryo-electron microscopy structure of the viral RdRp bound to suramin reveals two binding sites. One site directly blocks the binding of the RNA template strand and the other site clashes with the RNA primer strand near the RdRp catalytic site, thus inhibiting RdRp activity. Suramin blocks viral replication in Vero E6 cells, although the reasons underlying this effect are likely various. Our results provide a structural mechanism for a nonnucleotide inhibitor of the SARS-CoV-2 RdRp.
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http://dx.doi.org/10.1038/s41594-021-00570-0DOI Listing
March 2021

Structural insights into the human D1 and D2 dopamine receptor signaling complexes.

Cell 2021 Feb 10;184(4):931-942.e18. Epub 2021 Feb 10.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China; University of Chinese Academy of Sciences, Beijing 100049, China; School of Life Science and Technology, ShanghaiTech University, Shanghai 201210, China. Electronic address:

The D1- and D2-dopamine receptors (D1R and D2R), which signal through G and G, respectively, represent the principal stimulatory and inhibitory dopamine receptors in the central nervous system. D1R and D2R also represent the main therapeutic targets for Parkinson's disease, schizophrenia, and many other neuropsychiatric disorders, and insight into their signaling is essential for understanding both therapeutic and side effects of dopaminergic drugs. Here, we report four cryoelectron microscopy (cryo-EM) structures of D1R-G and D2R-G signaling complexes with selective and non-selective dopamine agonists, including two currently used anti-Parkinson's disease drugs, apomorphine and bromocriptine. These structures, together with mutagenesis studies, reveal the conserved binding mode of dopamine agonists, the unique pocket topology underlying ligand selectivity, the conformational changes in receptor activation, and potential structural determinants for G protein-coupling selectivity. These results provide both a molecular understanding of dopamine signaling and multiple structural templates for drug design targeting the dopaminergic system.
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http://dx.doi.org/10.1016/j.cell.2021.01.027DOI Listing
February 2021

Kinetics-Driven Drug Design Strategy for Next-Generation Acetylcholinesterase Inhibitors to Clinical Candidate.

J Med Chem 2021 Feb 11;64(4):1844-1855. Epub 2021 Feb 11.

State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Shanghai 201203, People's Republic of China.

The acetylcholinesterase (AChE) inhibitors remain key therapeutic drugs for the treatment of Alzheimer's disease (AD). However, the low-safety window limits their maximum therapeutic benefits. Here, a novel kinetics-driven drug design strategy was employed to discover new-generation AChE inhibitors that possess a longer drug-target residence time and exhibit a larger safety window. After detailed investigations, compound was identified as a highly potent, highly selective, orally bioavailable, and brain preferentially distributed AChE inhibitor. Moreover, it significantly ameliorated cognitive impairments in different mouse models with a lower effective dose than donepezil. The X-ray structure of the cocrystal complex provided a precise binding mode between and AChE. Besides, the data from the phase I trials demonstrated that had good safety, tolerance, and pharmacokinetic profiles at all preset doses in healthy volunteers, providing a solid basis for its further investigation in phase II trials for the treatment of AD.
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http://dx.doi.org/10.1021/acs.jmedchem.0c01863DOI Listing
February 2021

Structures of the human dopamine D3 receptor-G complexes.

Mol Cell 2021 Jan 27. Epub 2021 Jan 27.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China; University of Chinese Academy of Sciences, Beijing 100049, China; School of Life Science and Technology, ShanghaiTech University, 201210 Shanghai, China. Electronic address:

The dopamine system, including five dopamine receptors (D1R-D5R), plays essential roles in the central nervous system (CNS), and ligands that activate dopamine receptors have been used to treat many neuropsychiatric disorders. Here, we report two cryo-EM structures of human D3R in complex with an inhibitory G protein and bound to the D3R-selective agonists PD128907 and pramipexole, the latter of which is used to treat patients with Parkinson's disease. The structures reveal agonist binding modes distinct from the antagonist-bound D3R structure and conformational signatures for ligand-induced receptor activation. Mutagenesis and homology modeling illuminate determinants of ligand specificity across dopamine receptors and the mechanisms for G protein coupling. Collectively our work reveals the basis of agonist binding and ligand-induced receptor activation and provides structural templates for designing specific ligands to treat CNS diseases targeting the dopaminergic system.
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http://dx.doi.org/10.1016/j.molcel.2021.01.003DOI Listing
January 2021

Structures of the glucocorticoid-bound adhesion receptor GPR97-G complex.

Nature 2021 01 6;589(7843):620-626. Epub 2021 Jan 6.

Department of Physiology and Pathophysiology, School of Basic Medical Sciences, Peking University, Key Laboratory of Molecular Cardiovascular Science, Ministry of Education, Beijing, China.

Adhesion G-protein-coupled receptors (GPCRs) are a major family of GPCRs, but limited knowledge of their ligand regulation or structure is available. Here we report that glucocorticoid stress hormones activate adhesion G-protein-coupled receptor G3 (ADGRG3; also known as GPR97), a prototypical adhesion GPCR. The cryo-electron microscopy structures of GPR97-G complexes bound to the anti-inflammatory drug beclomethasone or the steroid hormone cortisol revealed that glucocorticoids bind to a pocket within the transmembrane domain. The steroidal core of glucocorticoids is packed against the 'toggle switch' residue W, which senses the binding of a ligand and induces activation of the receptor. Active GPR97 uses a quaternary core and HLY motif to fasten the seven-transmembrane bundle and to mediate G protein coupling. The cytoplasmic side of GPR97 has an open cavity, where all three intracellular loops interact with the G protein, contributing to the high basal activity of GRP97. Palmitoylation at the cytosolic tail of the G protein was found to be essential for efficient engagement with GPR97 but is not observed in other solved GPCR complex structures. Our work provides a structural basis for ligand binding to the seven-transmembrane domain of an adhesion GPCR and subsequent G protein coupling.
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http://dx.doi.org/10.1038/s41586-020-03083-wDOI Listing
January 2021

Single cell analysis reveals distinct immune landscapes in transplant and primary sarcomas that determine response or resistance to immunotherapy.

Nat Commun 2020 12 17;11(1):6410. Epub 2020 Dec 17.

Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, NC, 27708, USA.

Immunotherapy fails to cure most cancer patients. Preclinical studies indicate that radiotherapy synergizes with immunotherapy, promoting radiation-induced antitumor immunity. Most preclinical immunotherapy studies utilize transplant tumor models, which overestimate patient responses. Here, we show that transplant sarcomas are cured by PD-1 blockade and radiotherapy, but identical treatment fails in autochthonous sarcomas, which demonstrate immunoediting, decreased neoantigen expression, and tumor-specific immune tolerance. We characterize tumor-infiltrating immune cells from transplant and primary tumors, revealing striking differences in their immune landscapes. Although radiotherapy remodels myeloid cells in both models, only transplant tumors are enriched for activated CD8+ T cells. The immune microenvironment of primary murine sarcomas resembles most human sarcomas, while transplant sarcomas resemble the most inflamed human sarcomas. These results identify distinct microenvironments in murine sarcomas that coevolve with the immune system and suggest that patients with a sarcoma immune phenotype similar to transplant tumors may benefit most from PD-1 blockade and radiotherapy.
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http://dx.doi.org/10.1038/s41467-020-19917-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7746723PMC
December 2020

A unique hormonal recognition feature of the human glucagon-like peptide-2 receptor.

Cell Res 2020 Dec 25;30(12):1098-1108. Epub 2020 Nov 25.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, 201203, China.

Glucagon-like peptides (GLP-1 and GLP-2) are two proglucagon-derived intestinal hormones that mediate distinct physiological functions through two related receptors (GLP-1R and GLP-2R) which are important drug targets for metabolic disorders and Crohn's disease, respectively. Despite great progress in GLP-1R structure determination, our understanding on the differences of peptide binding and signal transduction between these two receptors remains elusive. Here we report the electron microscopy structure of the human GLP-2R in complex with GLP-2 and a G heterotrimer. To accommodate GLP-2 rather than GLP-1, GLP-2R fine-tunes the conformations of the extracellular parts of transmembrane helices (TMs) 1, 5, 7 and extracellular loop 1 (ECL1). In contrast to GLP-1, the N-terminal histidine of GLP-2 penetrates into the receptor core with a unique orientation. The middle region of GLP-2 engages with TM1 and TM7 more extensively than with ECL2, and the GLP-2 C-terminus closely attaches to ECL1, which is the most protruded among 9 class B G protein-coupled receptors (GPCRs). Functional studies revealed that the above three segments of GLP-2 are essential for GLP-2 recognition and receptor activation, especially the middle region. These results provide new insights into the molecular basis of ligand specificity in class B GPCRs and may facilitate the development of more specific therapeutics.
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http://dx.doi.org/10.1038/s41422-020-00442-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7785020PMC
December 2020

Structural basis for activation of the growth hormone-releasing hormone receptor.

Nat Commun 2020 10 15;11(1):5205. Epub 2020 Oct 15.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 201203, Shanghai, China.

Growth hormone-releasing hormone (GHRH) regulates the secretion of growth hormone that virtually controls metabolism and growth of every tissue through its binding to the cognate receptor (GHRHR). Malfunction in GHRHR signaling is associated with abnormal growth, making GHRHR an attractive therapeutic target against dwarfism (e.g., isolated growth hormone deficiency, IGHD), gigantism, lipodystrophy and certain cancers. Here, we report the cryo-electron microscopy (cryo-EM) structure of the human GHRHR bound to its endogenous ligand and the stimulatory G protein at 2.6 Å. This high-resolution structure reveals a characteristic hormone recognition pattern of GHRH by GHRHR, where the α-helical GHRH forms an extensive and continuous network of interactions involving all the extracellular loops (ECLs), all the transmembrane (TM) helices except TM4, and the extracellular domain (ECD) of GHRHR, especially the N-terminus of GHRH that engages a broad set of specific interactions with the receptor. Mutagenesis and molecular dynamics (MD) simulations uncover detailed mechanisms by which IGHD-causing mutations lead to the impairment of GHRHR function. Our findings provide insights into the molecular basis of peptide recognition and receptor activation, thereby facilitating the development of structure-based drug discovery and precision medicine.
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http://dx.doi.org/10.1038/s41467-020-18945-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7567103PMC
October 2020

Structure of nucleosome-bound DNA methyltransferases DNMT3A and DNMT3B.

Nature 2020 10 23;586(7827):151-155. Epub 2020 Sep 23.

Center for Epigenetics, Van Andel Institute, Grand Rapids, MI, USA.

CpG methylation by de novo DNA methyltransferases (DNMTs) 3A and 3B is essential for mammalian development and differentiation and is frequently dysregulated in cancer. These two DNMTs preferentially bind to nucleosomes, yet cannot methylate the DNA wrapped around the nucleosome core, and they favour the methylation of linker DNA at positioned nucleosomes. Here we present the cryo-electron microscopy structure of a ternary complex of catalytically competent DNMT3A2, the catalytically inactive accessory subunit DNMT3B3 and a nucleosome core particle flanked by linker DNA. The catalytic-like domain of the accessory DNMT3B3 binds to the acidic patch of the nucleosome core, which orients the binding of DNMT3A2 to the linker DNA. The steric constraints of this arrangement suggest that nucleosomal DNA must be moved relative to the nucleosome core for de novo methylation to occur.
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http://dx.doi.org/10.1038/s41586-020-2747-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7540737PMC
October 2020

RNA-dependent RNA polymerase: Structure, mechanism, and drug discovery for COVID-19.

Biochem Biophys Res Commun 2021 01 4;538:47-53. Epub 2020 Sep 4.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, 201203, China; University of Chinese Academy of Sciences, Beijing, 100049, China; School of Life Science and Technology, ShanghaiTech University, Shanghai, 201210, China. Electronic address:

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has rapidly become a global pandemic. Although great efforts have been made to develop effective therapeutic interventions, only the nucleotide analog remdesivir was approved for emergency use against COVID-19. Remdesivir targets the RNA-dependent RNA polymerase (RdRp), an essential enzyme for viral RNA replication and a promising drug target for COVID-19. Recently, several structures of RdRp in complex with substrate RNA and remdesivir were reported, providing insights into the mechanisms of RNA recognition by RdRp. These structures also reveal the mechanism of RdRp inhibition by nucleotide inhibitors and offer a molecular template for the development of RdRp-targeting drugs. This review discusses the recognition mechanism of RNA and nucleotide inhibitor by RdRp, and its implication in drug discovery.
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http://dx.doi.org/10.1016/j.bbrc.2020.08.116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7473028PMC
January 2021

Cryo-EM structure of an activated VIP1 receptor-G protein complex revealed by a NanoBiT tethering strategy.

Nat Commun 2020 08 17;11(1):4121. Epub 2020 Aug 17.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, 201203, China.

Vasoactive intestinal polypeptide receptor (VIP1R) is a widely expressed class B G protein-coupled receptor and a drug target for the treatment of neuronal, metabolic, and inflammatory diseases. However, our understanding of its mechanism of action and the potential of drug discovery targeting this receptor is limited by the lack of structural information of VIP1R. Here we report a cryo-electron microscopy structure of human VIP1R bound to PACAP27 and Gs heterotrimer, whose complex assembly is stabilized by a NanoBiT tethering strategy. Comparison with other class B GPCR structures reveals that PACAP27 engages VIP1R with its N-terminus inserting into the ligand binding pocket at the transmembrane bundle of the receptor, which subsequently couples to the G protein in a receptor-specific manner. This structure has provided insights into the molecular basis of PACAP27 binding and VIP receptor activation. The methodology of the NanoBiT tethering may help to provide structural information of unstable complexes.
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http://dx.doi.org/10.1038/s41467-020-17933-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7431577PMC
August 2020

Comparison and validation of seven white matter hyperintensities segmentation software in elderly patients.

Neuroimage Clin 2020 22;27:102357. Epub 2020 Jul 22.

Institut du Cerveau et de la Moelle épinière, ICM, F-75013 Paris, France; Inserm, U 1127, F-75013 Paris, France; CNRS, UMR 7225, F-75013 Paris, France; Sorbonne Université, F-75013 Paris, France; Inria Paris, Aramis Project-Team, F-75013 Paris, France; AP-HP, Hôpital de la Pitié-Salpêtrière, Department of Neurology, Institut de la Mémoire et de la Maladie d'Alzheimer (IM2A), F-75013 Paris, France.

Background: Manual segmentation is currently the gold standard to assess white matter hyperintensities (WMH), but it is time consuming and subject to intra and inter-operator variability.

Purpose: To compare automatic methods to segment white matter hyperintensities (WMH) in the elderly in order to assist radiologist and researchers in selecting the most relevant method for application on clinical or research data.

Material And Methods: We studied a research dataset composed of 147 patients, including 97 patients from the Alzheimer's Disease Neuroimaging Initiative (ADNI) 2 database and 50 patients from ADNI 3 and a clinical routine dataset comprising 60 patients referred for cognitive impairment at the Pitié-Salpêtrière hospital (imaged using four different MRI machines). We used manual segmentation as the gold standard reference. Both manual and automatic segmentations were performed using FLAIR MRI. We compared seven freely available methods that produce segmentation mask and are usable by a radiologist without a strong knowledge of computer programming: LGA (Schmidt et al., 2012), LPA (Schmidt, 2017), BIANCA (Griffanti et al., 2016), UBO detector (Jiang et al., 2018), W2MHS (Ithapu et al., 2014), nicMSlesion (with and without retraining) (Valverde et al., 2019, 2017). The primary outcome for assessing segmentation accuracy was the Dice similarity coefficient (DSC) between the manual and the automatic segmentation software. Secondary outcomes included five other metrics.

Results: A deep learning approach, NicMSlesion, retrained on data from the research dataset ADNI, performed best on this research dataset (DSC: 0.595) and its DSC was significantly higher than that of all others. However, it ranked fifth on the clinical routine dataset and its performance severely dropped on data with artifacts. On the clinical routine dataset, the three top-ranked methods were LPA, SLS and BIANCA. Their performance did not differ significantly but was significantly higher than that of other methods.

Conclusion: This work provides an objective comparison of methods for WMH segmentation. Results can be used by radiologists to select a tool.
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http://dx.doi.org/10.1016/j.nicl.2020.102357DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7394967PMC
July 2020

Structural insights into the activation of GLP-1R by a small molecule agonist.

Cell Res 2020 Dec 28;30(12):1140-1142. Epub 2020 Jul 28.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, 201203, China.

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http://dx.doi.org/10.1038/s41422-020-0384-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7784854PMC
December 2020

GABA receptor pas de deux: insights from high-resolution structures.

Authors:
Yi Jiang H Eric Xu

Cell Res 2020 Aug;30(8):631-632

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 201203, Shanghai, China.

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http://dx.doi.org/10.1038/s41422-020-0373-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7395756PMC
August 2020

The Long Noncoding RNA Promotes Sarcoma Metastasis by Regulating RNA Splicing Pathways.

Mol Cancer Res 2020 10 19;18(10):1534-1544. Epub 2020 Jun 19.

Department of Radiation Oncology, Duke University Medical Center, Durham, North Carolina.

Soft-tissue sarcomas (STS) are rare malignancies showing lineage differentiation toward diverse mesenchymal tissues. Half of all high-grade STSs develop lung metastasis with a median survival of 15 months. Here, we used a genetically engineered mouse model that mimics undifferentiated pleomorphic sarcoma (UPS) to study the molecular mechanisms driving metastasis. High-grade sarcomas were generated with Cre recombinase technology using mice with conditional mutations in and (KP) genes. After amputation of the limb bearing the primary tumor, mice were followed for the development of lung metastasis. Using RNA-sequencing of matched primary KP tumors and lung metastases, we found that the long noncoding RNA (lncRNA) Nuclear Enriched Abundant Transcript 1 () is significantly upregulated in lung metastases. Furthermore, RNA ISH of human UPS showed that is upregulated within a subset of lung metastases compared with paired primary UPS. Remarkably, CRISPR/Cas9-mediated knockout of suppressed the ability of KP tumor cells to colonize the lungs. To gain insight into the underlying mechanisms by which the lncRNA promotes sarcoma metastasis, we pulled down RNA and used mass spectrometry to identify interacting proteins. Interestingly, most interacting proteins are involved in RNA splicing regulation. In particular, KH-Type Splicing Regulatory Protein (KHSRP) interacts with and is associated with poor prognosis of human STS. Moreover, depletion of KHSRP suppressed the ability of KP tumor cells to colonize the lungs. Collectively, these results suggest that and its interacting proteins, which regulate RNA splicing, are involved in mediating sarcoma metastasis. IMPLICATIONS: Understanding that lncRNA promotes sarcoma metastasis, at least in part, through interacting with the RNA splicing regulator KHSRP may translate into new therapeutic approaches for sarcoma.
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http://dx.doi.org/10.1158/1541-7786.MCR-19-1170DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7541426PMC
October 2020

Small-molecule inhibitor targeting orphan nuclear receptor COUP-TFII for prostate cancer treatment.

Sci Adv 2020 May 29;6(18):eaaz8031. Epub 2020 Apr 29.

Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX 77030, USA.

The orphan nuclear receptor COUP-TFII is expressed at a low level in adult tissues, but its expression is increased and shown to promote progression of multiple diseases, including prostate cancer, heart failure, and muscular dystrophy. Suppression of COUP-TFII slows disease progression, making it an intriguing therapeutic target. Here, we identified a potent and specific COUP-TFII inhibitor through high-throughput screening. The inhibitor specifically suppressed COUP-TFII activity to regulate its target genes. Mechanistically, the inhibitor directly bound to the COUP-TFII ligand-binding domain and disrupted COUP-TFII interaction with transcription regulators, including FOXA1, thus repressing COUP-TFII activity on target gene regulation. Through blocking COUP-TFII's oncogenic activity in prostate cancer, the inhibitor efficiently exerted a potent antitumor effect in xenograft mouse models and patient-derived xenograft models. Our study identified a potent and specific COUP-TFII inhibitor that may be useful for the treatment of prostate cancer and possibly other diseases.
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http://dx.doi.org/10.1126/sciadv.aaz8031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7190335PMC
May 2020

Structural basis for inhibition of the RNA-dependent RNA polymerase from SARS-CoV-2 by remdesivir.

Science 2020 06 1;368(6498):1499-1504. Epub 2020 May 1.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.

The pandemic of coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has become a global crisis. Replication of SARS-CoV-2 requires the viral RNA-dependent RNA polymerase (RdRp) enzyme, a target of the antiviral drug remdesivir. Here we report the cryo-electron microscopy structure of the SARS-CoV-2 RdRp, both in the apo form at 2.8-angstrom resolution and in complex with a 50-base template-primer RNA and remdesivir at 2.5-angstrom resolution. The complex structure reveals that the partial double-stranded RNA template is inserted into the central channel of the RdRp, where remdesivir is covalently incorporated into the primer strand at the first replicated base pair, and terminates chain elongation. Our structures provide insights into the mechanism of viral RNA replication and a rational template for drug design to combat the viral infection.
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http://dx.doi.org/10.1126/science.abc1560DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7199908PMC
June 2020

Molecular mechanisms of fentanyl mediated β-arrestin biased signaling.

PLoS Comput Biol 2020 04 10;16(4):e1007394. Epub 2020 Apr 10.

Center for Epigenetics, Van Andel Research Institute, Grand Rapids, Michigan, United States of America.

The development of novel analgesics with improved safety profiles to combat the opioid epidemic represents a central question to G protein coupled receptor structural biology and pharmacology: What chemical features dictate G protein or β-arrestin signaling? Here we use adaptively biased molecular dynamics simulations to determine how fentanyl, a potent β-arrestin biased agonist, binds the μ-opioid receptor (μOR). The resulting fentanyl-bound pose provides rational insight into a wealth of historical structure-activity-relationship on its chemical scaffold. Following an in-silico derived hypothesis we found that fentanyl and the synthetic opioid peptide DAMGO require M153 to induce β-arrestin coupling, while M153 was dispensable for G protein coupling. We propose and validate an activation mechanism where the n-aniline ring of fentanyl mediates μOR β-arrestin through a novel M153 "microswitch" by synthesizing fentanyl-based derivatives that exhibit complete, clinically desirable, G protein biased coupling. Together, these results provide molecular insight into fentanyl mediated β-arrestin biased signaling and a rational framework for further optimization of fentanyl-based analgesics with improved safety profiles.
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http://dx.doi.org/10.1371/journal.pcbi.1007394DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7176292PMC
April 2020

Synthetic antibodies against BRIL as universal fiducial marks for single-particle cryoEM structure determination of membrane proteins.

Nat Commun 2020 03 27;11(1):1598. Epub 2020 Mar 27.

Department of Biochemistry and Molecular Biology, The University of Chicago, Chicago, IL, USA.

We propose the concept of universal fiducials based on a set of pre-made semi-synthetic antibodies (sABs) generated by customized phage display selections against the fusion protein BRIL, an engineered variant of apocytochrome b562a. These sABs can bind to BRIL fused either into the loops or termini of different GPCRs, ion channels, receptors and transporters without disrupting their structure. A crystal structure of BRIL in complex with an affinity-matured sAB (BAG2) that bound to all systems tested delineates the footprint of interaction. Negative stain and cryoEM data of several examples of BRIL-membrane protein chimera highlight the effectiveness of the sABs as universal fiducial marks. Taken together with a cryoEM structure of sAB bound human nicotinic acetylcholine receptor, this work demonstrates that these anti-BRIL sABs can greatly enhance the particle properties leading to improved cryoEM outcomes, especially for challenging membrane proteins.
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http://dx.doi.org/10.1038/s41467-020-15363-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7101349PMC
March 2020

Identification and structural insight of an effective PPARγ modulator with improved therapeutic index for anti-diabetic drug discovery.

Chem Sci 2020 Feb 21;11(8):2260-2268. Epub 2020 Jan 21.

Guangzhou Municipal and Guangdong Provincial Key Laboratory of Protein Modification and Degradation & Molecular Target and Clinical Pharmacology , State Key Laboratory of Respiratory Disease , School of Pharmaceutical Sciences & the Fifth Affiliated Hospital , Guangzhou Medical University , Guangzhou , Guangdong 511436 , China . Email:

Peroxisome proliferator-activated receptor γ (PPARγ) is a key regulator of glucose homeostasis and lipid metabolism, and an important target for the development of modern anti-diabetic drugs. However, current PPARγ-targeting anti-diabetic drugs such as classical thiazolidinediones (TZDs) are associated with undesirable side effects. To address this concern, we here describe the structure-based design, synthesis, identification and detailed and characterization of a novel, decanoic acid (DA)-based and selective PPARγ modulator (SPPARγM), VSP-77, especially (S)-VSP-77, as the potential "hit" for the development of improved and safer anti-diabetic therapeutics. We have also determined the co-crystal structure of the PPARγ ligand-binding domain (LBD) in complex with two molecules of (S)-VSP-77, which reveal a previously undisclosed allosteric binding mode. Overall, these findings not only demonstrate the therapeutic advantage of (S)-VSP-77 over current TZD drugs and representative partial agonist INT131, but also provide a rational basis for the development of future SPPARγMs as safe and highly efficacious anti-diabetic drugs.
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http://dx.doi.org/10.1039/c9sc05487aDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7059199PMC
February 2020

Structural basis of Fusarium myosin I inhibition by phenamacril.

PLoS Pathog 2020 03 12;16(3):e1008323. Epub 2020 Mar 12.

Key Laboratory of Pesticide, College of Plant Protection, Nanjing Agricultural University, Nanjing, China.

Fusarium is a genus of filamentous fungi that includes species that cause devastating diseases in major staple crops, such as wheat, maize, rice, and barley, resulting in severe yield losses and mycotoxin contamination of infected grains. Phenamacril is a novel fungicide that is considered environmentally benign due to its exceptional specificity; it inhibits the ATPase activity of the sole class I myosin of only a subset of Fusarium species including the major plant pathogens F. graminearum, F. asiaticum and F. fujikuroi. To understand the underlying mechanisms of inhibition, species specificity, and resistance mutations, we have determined the crystal structure of phenamacril-bound F. graminearum myosin I. Phenamacril binds in the actin-binding cleft in a new allosteric pocket that contains the central residue of the regulatory Switch 2 loop and that is collapsed in the structure of a myosin with closed actin-binding cleft, suggesting that pocket occupancy blocks cleft closure. We have further identified a single, transferable phenamacril-binding residue found exclusively in phenamacril-sensitive myosins to confer phenamacril selectivity.
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http://dx.doi.org/10.1371/journal.ppat.1008323DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7100991PMC
March 2020

Structure of formylpeptide receptor 2-G complex reveals insights into ligand recognition and signaling.

Nat Commun 2020 02 14;11(1):885. Epub 2020 Feb 14.

Laboratory for GPCR Biology, Department of Pharmacology and Chemical Biology, University of Pittsburgh School of Medicine, University of Pittsburgh, Pittsburgh, PA, 15261, USA.

Formylpeptide receptors (FPRs) as G protein-coupled receptors (GPCRs) can recognize formylpeptides derived from pathogens or host cells to function in host defense and cell clearance. In addition, FPRs, especially FPR2, can also recognize other ligands with a large chemical diversity generated at different stages of inflammation to either promote or resolve inflammation in order to maintain a balanced inflammatory response. The mechanism underlying promiscuous ligand recognition and activation of FPRs is not clear. Here we report a cryo-EM structure of FPR2-G signaling complex with a peptide agonist. The structure reveals a widely open extracellular region with an amphiphilic environment for ligand binding. Together with computational docking and simulation, the structure suggests a molecular basis for the recognition of formylpeptides and a potential mechanism of receptor activation, and reveals conserved and divergent features in G coupling. Our results provide a basis for understanding the molecular mechanism of the functional promiscuity of FPRs.
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http://dx.doi.org/10.1038/s41467-020-14728-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7021761PMC
February 2020

Fasting-induced FGF21 signaling activates hepatic autophagy and lipid degradation via JMJD3 histone demethylase.

Nat Commun 2020 02 10;11(1):807. Epub 2020 Feb 10.

Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, Urbana, IL, 61801, USA.

Autophagy is essential for cellular survival and energy homeostasis under nutrient deprivation. Despite the emerging importance of nuclear events in autophagy regulation, epigenetic control of autophagy gene transcription remains unclear. Here, we report fasting-induced Fibroblast Growth Factor-21 (FGF21) signaling activates hepatic autophagy and lipid degradation via Jumonji-D3 (JMJD3/KDM6B) histone demethylase. Upon FGF21 signaling, JMJD3 epigenetically upregulates global autophagy-network genes, including Tfeb, Atg7, Atgl, and Fgf21, through demethylation of histone H3K27-me3, resulting in autophagy-mediated lipid degradation. Mechanistically, phosphorylation of JMJD3 at Thr-1044 by FGF21 signal-activated PKA increases its nuclear localization and interaction with the nuclear receptor PPARα to transcriptionally activate autophagy. Administration of FGF21 in obese mice improves defective autophagy and hepatosteatosis in a JMJD3-dependent manner. Remarkably, in non-alcoholic fatty liver disease patients, hepatic expression of JMJD3, ATG7, LC3, and ULK1 is substantially decreased. These findings demonstrate that FGF21-JMJD3 signaling epigenetically links nutrient deprivation with hepatic autophagy and lipid degradation in mammals.
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http://dx.doi.org/10.1038/s41467-020-14384-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7010817PMC
February 2020

Molecular Basis for Hormone Recognition and Activation of Corticotropin-Releasing Factor Receptors.

Mol Cell 2020 02 30;77(3):669-680.e4. Epub 2020 Jan 30.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China; University of Chinese Academy of Sciences, Beijing 100049, China. Electronic address:

Corticotropin-releasing factor (CRF) and the three related peptides urocortins 1-3 (UCN1-UCN3) are endocrine hormones that control the stress responses by activating CRF1R and CRF2R, two members of class B G-protein-coupled receptors (GPCRs). Here, we present two cryoelectron microscopy (cryo-EM) structures of UCN1-bound CRF1R and CRF2R with the stimulatory G protein. In both structures, UCN1 adopts a single straight helix with its N terminus dipped into the receptor transmembrane bundle. Although the peptide-binding residues in CRF1R and CRF2R are different from other members of class B GPCRs, the residues involved in receptor activation and G protein coupling are conserved. In addition, both structures reveal bound cholesterol molecules to the receptor transmembrane helices. Our structures define the basis of ligand-binding specificity in the CRF receptor-hormone system, establish a common mechanism of class B GPCR activation and G protein coupling, and provide a paradigm for studying membrane protein-lipid interactions for class B GPCRs.
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http://dx.doi.org/10.1016/j.molcel.2020.01.013DOI Listing
February 2020

Toward a Structural Understanding of Class B GPCR Peptide Binding and Activation.

Mol Cell 2020 02 30;77(3):656-668.e5. Epub 2020 Jan 30.

Drug Discovery Biology and Department of Pharmacology, Monash Institute of Pharmaceutical Sciences, Monash University, Parkville 3052, VIC, Australia; School of Pharmacy, Fudan University, Shanghai 201203, China. Electronic address:

Class B G protein-coupled receptors (GPCRs) are important therapeutic targets for major diseases. Here, we present structures of peptide and Gs-bound pituitary adenylate cyclase-activating peptide, PAC1 receptor, and corticotropin-releasing factor (CRF), (CRF1) receptor. Together with recently solved structures, these provide coverage of the major class B GPCR subfamilies. Diverse orientations of the extracellular domain to the receptor core in different receptors are at least partially dependent on evolutionary conservation in the structure and nature of peptide interactions. Differences in peptide interactions to the receptor core also influence the interlinked TM2-TM1-TM6/ECL3/TM7 domain, and this is likely important in their diverse signaling. However, common conformational reorganization of ECL2, linked to reorganization of ICL2, modulates G protein contacts. Comparison between receptors reveals ICL2 as a key domain forming dynamic G protein interactions in a receptor- and ligand-specific manner. This work advances our understanding of class B GPCR activation and Gs coupling.
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http://dx.doi.org/10.1016/j.molcel.2020.01.012DOI Listing
February 2020

Cryo-EM Structure of the Human Cannabinoid Receptor CB2-G Signaling Complex.

Cell 2020 02 30;180(4):645-654.e13. Epub 2020 Jan 30.

Department of Pharmaceutical Sciences, Computational Chemical Genomics Screen Center, School of Pharmacy, and NIDA National Center of Excellence for Computational Drug Abuse Research, University of Pittsburgh, Pittsburgh, PA 15261, USA; Drug Discovery Institute and Departments of Computational Biology and of Structural Biology, University of Pittsburgh, Pittsburgh, PA 15261, USA. Electronic address:

Drugs selectively targeting CB2 hold promise for treating neurodegenerative disorders, inflammation, and pain while avoiding psychotropic side effects mediated by CB1. The mechanisms underlying CB2 activation and signaling are poorly understood but critical for drug design. Here we report the cryo-EM structure of the human CB2-G signaling complex bound to the agonist WIN 55,212-2. The 3D structure reveals the binding mode of WIN 55,212-2 and structural determinants for distinguishing CB2 agonists from antagonists, which are supported by a pair of rationally designed agonist and antagonist. Further structural analyses with computational docking results uncover the differences between CB2 and CB1 in receptor activation, ligand recognition, and G coupling. These findings are expected to facilitate rational structure-based discovery of drugs targeting the cannabinoid system.
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http://dx.doi.org/10.1016/j.cell.2020.01.007DOI Listing
February 2020

Crystal structure of heliorhodopsin 48C12.

Cell Res 2020 01 26;30(1):88-90. Epub 2019 Dec 26.

Laboratory of Receptor Structure and Signaling, HIT Center for Life Sciences, Harbin Institute of Technology, Harbin, Heilongjiang, 150001, China.

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http://dx.doi.org/10.1038/s41422-019-0266-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6951262PMC
January 2020

Maternal risk factors for adverse outcome in asphyxiated newborns treated with hypothermia: parity and labor duration matter.

J Matern Fetal Neonatal Med 2019 Dec 26:1-9. Epub 2019 Dec 26.

Division of Newborn Medicine, Department of Pediatrics, Montreal Children's Hospital, McGill University, Montreal, Canada.

Perinatal asphyxia remains a frequent cause of neonatal mortality and long-term neurodevelopmental sequelae, despite the introduction of therapeutic hypothermia. Specific maternal characteristics may predispose asphyxiated newborns treated with hypothermia to worse outcome. To investigate the possible association between specific maternal factors and adverse outcome in asphyxiated newborns treated with hypothermia. We conducted a retrospective review of our database of 215 asphyxiated newborns treated with hypothermia from 2008 to 2015. We collected maternal characteristics including parity and labor duration, and we defined as death and/or brain injury. We compared the maternal characteristics between the asphyxiated newborns who developed adverse outcome and those who did not. Asphyxiated newborns born to nulliparous mothers had a significantly higher risk of adverse outcome (61%), compared to asphyxiated newborns born from primiparous (19%) and multiparous (20%) mothers ( .002). Labor duration was longer in nulliparous mothers ( .04). Among mothers who delivered vaginally, labor duration was significantly longer in newborns developing adverse outcome ( .04). In multivariable analysis, parity was confirmed as an independent predictor of adverse outcome in all newborns, but labor duration showed a borderline non-significant association with adverse outcome ( .051) only in newborns born vaginally. Labor duration beyond 12 h of life was associated with maximal sensitivity and specificity in detecting asphyxiated newborns at an increased risk of adverse outcome despite hypothermia treatment (AUC 0.62,  .044). Newborns with evidence of perinatal asphyxia, born to nulliparous mothers, and especially to those in whom the duration of labor has been prolonged, might be at higher risk of death or brain injury despite the use of therapeutic hypothermia.
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http://dx.doi.org/10.1080/14767058.2019.1706472DOI Listing
December 2019

A complex structure of arrestin-2 bound to a G protein-coupled receptor.

Cell Res 2019 Dec 27;29(12):971-983. Epub 2019 Nov 27.

The CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, 201203, China.

Arrestins comprise a family of signal regulators of G-protein-coupled receptors (GPCRs), which include arrestins 1 to 4. While arrestins 1 and 4 are visual arrestins dedicated to rhodopsin, arrestins 2 and 3 (Arr2 and Arr3) are β-arrestins known to regulate many nonvisual GPCRs. The dynamic and promiscuous coupling of Arr2 to nonvisual GPCRs has posed technical challenges to tackle the basis of arrestin binding to GPCRs. Here we report the structure of Arr2 in complex with neurotensin receptor 1 (NTSR1), which reveals an overall assembly that is strikingly different from the visual arrestin-rhodopsin complex by a 90° rotation of Arr2 relative to the receptor. In this new configuration, intracellular loop 3 (ICL3) and transmembrane helix 6 (TM6) of the receptor are oriented toward the N-terminal domain of the arrestin, making it possible for GPCRs that lack the C-terminal tail to couple Arr2 through their ICL3. Molecular dynamics simulation and crosslinking data further support the assembly of the Arr2‒NTSR1 complex. Sequence analysis and homology modeling suggest that the Arr2‒NTSR1 complex structure may provide an alternative template for modeling arrestin-GPCR interactions.
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http://dx.doi.org/10.1038/s41422-019-0256-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6951264PMC
December 2019