A5040600416- Nuclear Structure and Function
- RNA Research and Splicing
- RNA modifications and cancer
- RNA and protein synthesis mechanisms
- Lysosomal Storage Disorders Research
- Cellular transport and secretion
- Mitochondrial Function and Pathology
- Advanced Electron Microscopy Techniques and Applications
- Proteoglycans and glycosaminoglycans research
- Trypanosoma species research and implications
- Autophagy in Disease and Therapy
- Enzyme Structure and Function
- Glycosylation and Glycoproteins Research
- Genomics and Chromatin Dynamics
- Ubiquitin and proteasome pathways
- Microtubule and mitosis dynamics
- RNA Interference and Gene Delivery
- Advanced Fluorescence Microscopy Techniques
- Bacteriophages and microbial interactions
- Caveolin-1 and cellular processes
- Chromosomal and Genetic Variations
- Viral gastroenteritis research and epidemiology
- SARS-CoV-2 and COVID-19 Research
- Genetics and Neurodevelopmental Disorders
- Electron and X-Ray Spectroscopy Techniques
University of Michigan
2021-2025
Thermo Fisher Scientific (United States)
2024
Ann Arbor Center for Independent Living
2024
Michigan United
2023
European Molecular Biology Laboratory
2015-2022
Max Planck Institute of Biophysics
2020-2022
European Molecular Biology Laboratory
2020
Max Planck Institute of Molecular Physiology
2013-2017
The spike protein (S) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is required for cell entry and the primary focus vaccine development. In this study, we combined cryo-electron tomography, subtomogram averaging, molecular dynamics simulations to structurally analyze S in situ. Compared with recombinant S, viral was more heavily glycosylated occurred mostly closed prefusion conformation. We show that stalk domain contains three hinges, giving head unexpected orientational...
Blueprint for a macromolecular machine Nuclear pore complexes (NPCs) consist of around 1000 protein subunits, are embedded in the membrane that surrounds nucleus, and regulate transport between nucleus cytoplasm. Although overall shape NPCs is known, details this complex have been obscure. Now, Lin et al. reconstituted components, determined interactions them, fitted them into tomographic reconstruction. Kosinski provided an architectural map inner ring pore. Science , issue pp....
INTRODUCTION The eukaryotic nucleus pro-tects the genome and is enclosed by two membranes of nuclear envelope. Nuclear pore complexes (NPCs) perforate envelope to facilitate nucleocytoplasmic transport. With a molecular weight ∼120 MDa, human NPC one larg-est protein complexes. Its ~1000 proteins are taken in multiple copies from set about 30 distinct nucleoporins (NUPs). They can be roughly categorized into classes. Scaf-fold NUPs contain folded domains form cylindrical scaffold...
Rapid structural analysis of purified proteins and their complexes has become increasingly common thanks to key methodological advances in cryo-electron microscopy (cryo-EM) associated data processing software packages. In contrast, analogous cells via tomography (cryo-ET) remains challenging due critical technical bottlenecks, including low-throughput sample preparation imaging, laborious methods. Here, we describe a rapid situ cryo-ET workflow that results the routine determination sub-nm...
The partitioning of cellular components between the nucleus and cytoplasm is defining feature eukaryotic life. nuclear pore complex (NPC) selectively gates transport macromolecules these compartments, but it unknown whether surveillance mechanisms exist to reinforce this function. By leveraging in situ cryo-electron tomography image native environment Chlamydomonas reinhardtii, we observed that 26S proteasomes crowd around NPCs. Through a combination subtomogram averaging nanometer-precision...
Abstract Nuclear pore complexes (NPCs) span the nuclear envelope and mediate nucleocytoplasmic exchange. They are a hallmark of eukaryotes deeply rooted in evolutionary origin cellular compartmentalization. NPCs have an elaborate architecture that has been well studied vertebrates. Whether this is unique or varies significantly other eukaryotic kingdoms remains unknown, predominantly due to missing situ structural data. Here, we report algal NPC from early branching eukaryote Chlamydomonas...
Abstract Cryo electron tomography with subsequent subtomogram averaging is a powerful technique to structurally analyze macromolecular complexes in their native context. Although close atomic resolution principle can be obtained, it not clear how individual experimental parameters contribute the attainable resolution. Here, we have used immature HIV-1 lattice as benchmarking sample optimize for averaging. We systematically tested various such order of projections, different angular...
Rotavirus assembly is a complex process that involves the stepwise acquisition of protein layers in distinct intracellular locations to form fully assembled particle. Understanding and visualization has been hampered by inaccessibility unstable intermediates. We characterize pathway group A rotaviruses observed situ within cryo-preserved infected cells through use cryoelectron tomography cellular lamellae. Our findings demonstrate viral polymerase VP1 recruits genomes during particle...
Kinetochores are macromolecular assemblies that connect chromosomes to spindle microtubules (MTs) during mitosis. The metazoan-specific ≈800-kD ROD–Zwilch–ZW10 (RZZ) complex builds a fibrous corona assembles on mitotic kinetochores before MT attachment promote chromosome alignment and robust assembly checkpoint signaling. In this study, we combine biochemical reconstitutions, single-particle electron cryomicroscopy, cross-linking mass spectrometry, structural modeling build complete model of...
Nuclear pore complexes (NPCs) are large macromolecular machines that mediate the traffic between nucleus and cytoplasm. In vertebrates, each NPC consists of ∼1000 proteins, termed nucleoporins, has a mass more than 100 MDa. While pseudo-atomic static model central scaffold recently been assembled by integrating data from isolated proteins complexes, many structural components still remain elusive due to enormous size flexibility NPC. Here, we explored power three-dimensional (3D)...
Abstract The spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is required for cell entry and the major focus vaccine development. We combine cryo electron tomography, subtomogram averaging molecular dynamics simulations to structurally analyze S in situ . Compared recombinant S, viral more heavily glycosylated occurs predominantly a closed pre-fusion conformation. show that stalk domain contains three hinges give globular unexpected orientational freedom....
The T andem of pore domain in a W eak Inward R ectifying K+ channel 2 (TWIK-2; KCNK6) is member the Two-Pore Domain K + (K 2P ) family, which associated with pulmonary hypertension, lung injury, and inflammation. structure regulatory mechanisms TWIK-2 remain largely unknown. Here, we present cryo-electron microscopy (cryo-EM) human at ∼3.7 Å highlight its conserved unique features. Using automated whole-cell patch clamp recordings, demonstrate that gating voltage-dependent insensitive to...
Abstract Nuclear pore complexes (NPCs) mediate nucleocytoplasmic transport. Their intricate 120 MDa architecture remains incompletely understood. Here, we report a near-complete structural model of the human NPC scaffold with explicit membrane and in multiple conformational states. We combined AI-based structure prediction situ cellulo cryo-electron tomography integrative modeling. show that linker Nups spatially organize within across subcomplexes to establish higher-order structure....
Accurate chromosome segregation during mitosis and meiosis is crucial for cellular organismal viability. Kinetochores connect chromosomes with spindle microtubules are essential segregation. These large protein scaffolds emerge from the centromere, a specialized region of enriched histone H3 variant CENP-A. In most eukaryotes, kinetochore core consists centromere-proximal constitutive centromere-associated network (CCAN), which binds CENP-A contains 16 subunits, centromere-distal Knl1...
Significance K63 ubiquitination of ribosomes serves as a key regulator protein production during cellular exposure to oxidative stress. Defining the structural and functional mechanisms translation regulation would support current understanding critical reprogramming eukaryotic gene expression. Our paper presents an examination structure ubiquitinated ribosomes, revealing that this modification structurally destabilizes proteins involved in binding factors is required trap at...
Inter-organellar communication is critical for cellular metabolic homeostasis. One of the most abundant inter-organellar interactions are those at endoplasmic reticulum and mitochondria contact sites (ERMCS). However, a detailed understanding mechanisms governing ERMCS regulation their roles in metabolism limited by lack tools that permit temporal induction reversal. Through unbiased screening approaches, we identified fedratinib, an FDA-approved drug, dramatically increases abundance...
Sirtuins are NAD+-dependent protein deacetylases regulating metabolism, stress responses and ageing processes. Among the seven mammalian Sirtuins, Sirt1 is physiologically best-studied isoform. It regulates nuclear functions such as chromatin remodelling gene transcription, it appears to mediate beneficial effects of a low calorie diet which can partly be mimicked by activating polyphenol resveratrol. The molecular details domain architecture regulation, however, little understood. has...
Cryogenic electron microscopy (cryo-EM) has become a widely used tool for determining the protein structure. Despite recent technical advances, sample preparation remains major bottleneck several reasons, including denaturation at air-water interface, presence of preferred orientations, nonuniform ice layers, etc. Graphene, two-dimensional allotrope carbon consisting single atomic layer, recently gained attention as near-ideal support film cryo-EM that can overcome these challenges because...
Degradation of heparan sulfate (HS), a glycosaminoglycan (GAG) comprised repeating units N-acetylglucosamine and glucuronic acid, begins in the cytosol is completed lysosomes. Acetylation terminal non-reducing amino group α-D-glucosamine HS essential for its complete breakdown into monosaccharides free sulfate. Heparan-α-glucosaminide N-acetyltransferase (HGSNAT), resident lysosomal membrane, catalyzes this acetylation reaction by accepting transferring acetyl from cytosolic acetyl-CoA to...