Biology subjects

Hansen, J. M.

Publications and source records attributed to Hansen, J. M..

5 recordsLinked to original sources

bioRxiv2026

Microglial and Neuronal Cross-talk in the Nucleus Accumbens

Microglia are the brain's resident immune cells and rapidly migrate toward sites of neuronal stress and neurotransmitter imbalance. The molecular cues directing microglia movement remain poorly understood, particularly in brain regions involved in reward processing, including the nucleus accumbens (NAc). Dopamine (DA) dysregulation and production of reactive oxygen species (ROS) are hallmarks of substance use disorders and neurodegenerative diseases. Dopamine and adenosine triphosphate (ATP) are co-released in the NAc, but the effects of ROS on ATP/DA co-transmission and microglia surveillance remain unknown. Using multiphoton imaging in brain slices with GFP-labeled microglia, we found that ATP consistently and non-selectively drives microglia chemoattraction, whereas DA chemoattraction only occurs in a subset of microglia through D1 but not D2 receptor activation. Microglia enter a reactive state via LPS, and LPS results in enhanced DA/ATP release measured via fast scan cyclic voltammetry. ROS production also transitions microglia to a reactive state, but inhibits DA release, with mixed effects on ATP release. These findings identify ATP- and DA-mediated mechanisms for NAc microglia chemoattraction and opposing immune dysregulation (via LPS or ROS) of DA terminal function.

neuroscience↗

bioRxiv2023

Unveiling the ultrastructural landscape of native extracellular matrix via lift-out cryo-FIBSEM and cryo-ET

The extracellular matrix (ECM) is a highly hydrated, three-dimensional network composed of various macromolecules and signaling factors. It serves as a structural scaffold for cells and plays an essential role in the regulation of numerous cellular processes, including cell migration, adhesion, and proliferation. Despite its importance in metazoans, structural knowledge is rudimentary on how the components of the matrisome are secreted, remodeled, and interact with each other and with surrounding cells. Specifically, the exact molecular assembly of important ECM fibers, such as fibronectin fibrils, fibrillin microfibrils, or Collagen-VI filaments has remained enigmatic. This is largely due to methodological limitations in specimen preparation for conventional room-temperature electron microscopy (EM). To overcome these limitations, we have developed a cell culture-based 3D-ECM platform compatible with sample thinning by cryo-lift out focused ion beam (FIB) milling and cryo-electron tomography (cryo-ET). Our workflow involves the implementation of cell-derived matrices (CDMs) grown on EM grids, resulting in a highly adaptable and versatile tool to closely mimic ECM environments. This allows us to visualize native ECM and its components for the first time in their fully hydrated, cellular context. Our data reveals an intricate network of ECM fibers and their positioning with respect to matrix-secreting cells. In addition to D-spaced collagen fibers, we visualize previously unresolved fibrous structures, and an amorphous matrix co-assembling in proximity to ECM fibers and delineating the boundary between ECM and empty extra-cellular space. Intra- and extracellular granules presumably represent assembly intermediates of the ECM. Our results add to the structural atlas of the ECM and provide novel insights into ECM secretion, assembly and maintenance.

cell biology↗

bioRxiv2023

Multi-modal cryo-EM reveals trimers of protein A10 to form the palisade layer in poxvirus cores

Poxviruses are among the largest double-stranded DNA viruses with members such as Variola virus, Monkeypox virus and the famous vaccination strain Vaccinia virus (VACV). Knowledge about the structural proteins that form the viral core, found in all infectious poxvirus forms, has remained sparse. While major core proteins have been annotated via indirect experimental evidence, their structures have remained elusive and they could not be assigned to the individual architectural features of the core. Hence, which proteins constitute which layers of the core, such as the so-called palisade layer and the inner core wall has remained enigmatic. Here, we have performed a multi-modal cryo-electron microscopy (cryo-EM) approach to elucidate the structural determinants of the VACV core. In combination with molecular modeling using AlphaFold, we unambiguously identify trimers formed by the cleavage product of A10 as the key component of the palisade layer. This allows us to place previously-obtained descriptions of protein interactions within the core wall into perspective and to provide a substantially revised model of poxvirus core architecture. Importantly, we show that interactions within A10 trimers are likely identical among Poxviridae, implying that our structural observations should be generalizable over most, if not all members of this important virus family. One sentence summarySingle-particle cryo-EM, cryo-electron tomography, and AlphaFold modeling reveal the structural architecture of the poxvirus core and identify trimers of protein A10 as the key component of the palisade layer.

molecular biology↗

bioRxiv2021

Cryo-EM Structures of CTP Synthase Filaments Reveal Mechanism of pH-Sensitive Assembly During Budding Yeast Starvation

Many metabolic enzymes self-assemble into micron-scale filaments to organize and regulate metabolism. The appearance of these assemblies often coincides with large metabolic changes as in development, cancer, and stress. Yeast undergo cytoplasmic acidification upon starvation, triggering the assembly of many metabolic enzymes into filaments. However, it is unclear how these filaments assemble at the molecular level and what their role is in the yeast starvation response. CTP Synthase (CTPS) assembles into metabolic filaments across many species. Here, we characterize in vitro polymerization and investigate in vivo consequences of CTPS assembly in yeast. Cryo-EM structures reveal a pH-sensitive assembly mechanism and highly ordered filament bundles that stabilize an inactive state of the enzyme, features unique to yeast CTPS. Disruption of filaments in cells with non-assembly or hyper-assembly mutations decreases growth rate, reflecting the importance of regulated CTPS filament assembly in homeotstasis.

biochemistry↗

bioRxiv2017

Cryo-EM structure of the bacterial actin AlfA reveals unique assembly and ATP-binding interactions and the absence of a conserved subdomain

Bacterial actins are an evolutionarily diverse family of ATP-dependent filaments built from protomers with a conserved structural fold. Actin-based segregation systems are encoded on many bacterial plasmids and function to partition plasmids into daughter cells. The bacterial actin AlfA segregates plasmids by a mechanism distinct from other partition systems, dependent on its unique dynamic properties. Here, we report the near-atomic resolution cryo-EM structure of the AlfA filament, which reveals a strikingly divergent filament architecture resulting from the loss of a subdomain conserved in all other actins and a novel mode of ATP binding. Its unusual assembly interfaces and nucleotide interactions provide insight into AlfA dynamics, and expand the range of evolutionary variation accessible to actin quaternary structure.\n\nSignificance StatementActin filaments are dynamic cytoskeletal elements that assemble upon ATP binding. Actin homologs are present in all domains of life, and all share a similar three-dimensional structure of the assembling subunit, but evolutionary changes to subunit have generated many different actin filament structures. The filament structure of the bacterial actin AlfA, which positions plasmids - small, circular DNA molecules that encode important genes - to ensure that each daughter cell receives a copy at cell division. AlfA is different from all other actins in two critical ways: it binds to ATP in a unique way, and it is missing a quarter of the conserved structural core. These differences explain unusual AlfA assembly dynamics that underlie its ability to move plasmids.

biophysics↗