Biology subjects

Schoehn, G.

Publications and source records attributed to Schoehn, G..

26 records · Page 2Linked to original sources

bioRxiv2021

Immediate and sustained effects of cobalt-containing pigments on macrophages

Pigments are among the oldest nanoparticulate products known to mankind, and their use in tattoos is also very old. Nowadays, 25% of American people aged 18 to 50 are tattooed, which poses the question of the delayed effects of tattoos. In this article, we investigated three cobalt (Pigment Violet 14 (purple color)) or cobalt alloy pigments (Pigment Blue 28 (blue color), Pigment Green 14 (green color)), and one zinc pigment (Pigment White 4 (white color)) which constitute a wide range of colors found in tattoos. These pigments contain microparticles and a significant proportion of submicroparticles or nanoparticles (in either aggregate or free form). Because of the key role of macrophages in the scavenging of particulate materials, we tested the effects of cobalt- and zinc-based pigments on the J774A.1 macrophage cell line. In order to detect delayed effects, we compared two exposure schemes: acute exposure for 24 hours and an exposure for 24 hours followed by a 3-day post-exposure recovery period. The conjunction of these two schemes allowed for the investigation of the delayed or sustained effects of pigments. All pigments induced functional effects on macrophages, most of which were pigment-dependent. For example, Pigment Green 19, Pigment Blue 28, and Pigment White 4 showed a delayed alteration of the phagocytic capacity of cells. Moreover, all the pigments tested induced a slight but significant increase in tumor necrosis factor secretion. This effect, however, was transitory. Conversely, only Pigment Blue 28 induced both a short and sustained increase in interleukin 6 secretion. Results showed that in response to bacterial stimuli (LPS), the secretion of tumor necrosis factor and interleukin 6 declined after exposure to pigments followed by a recovery period. For chemoattractant cytokines (MCP-1 or MIP-1), delayed effects were observed with a secretion decreased in presence of Pigment Blue 28 and Pigment violet 14, both with or without LPS stimuli. The pigments also induced persisting changes in some important macrophage membrane markers such as CD11b, an integrin contributing to cell adhesion and immunological tolerance. In conclusion, the pigments induced functional disorders in macrophages, which, in some cases, persist long after exposure, even at non-toxic doses. Contribution to the field statementUnlike dyes, which are water-soluble, pigments are water-insoluble and thus viewed as inert coloring substances. However, historical pigments such as lead white or vermilion (mercuric sulfide) have been shown to be toxic, suggesting that pigments inertness may not be complete under biological conditions. Pigments being particulate (nano) materials, they are taken up by professional scavenger cells such as macrophages once they have penetrated into the body. One epitome of this situation is represented by tattooing: the pigments injected with the ink are taken up by dermal macrophages, which life cycle ensures the localized persistence of tattoos over time. Using an in vitro macrophage culture system adapted to study delayed effects, we have investigated the effects of a series of cobalt- and zinc-containing pigments on macrophages. First of all, the toxicity of the pigments correlated well with their solubility in acidic media, i.e. conditions prevailing in the phagolysosomes. Even when used at non-toxic doses, the cobalt and zinc pigments showed immediate and/or delayed effects on macrophage functions such as phagocytosis, adhesion, tissue repair or response to bacterial stimuli. Overall, these results show that some pigments may not be as inert as previously thought, and describe a system to investigate these effects.

immunology↗

bioRxiv2020

Nonclassical nucleation of protein mesocrystals via oriented attachment

Self-assembly of proteins holds great promise for the bottom-up design and production of synthetic biomaterials. In conventional approaches, designer proteins are pre-programmed with specific recognition sites that drive the association process towards a desired organized state. Although proven effective, this approach poses restrictions on the complexity and material properties of the end-state. An alternative, hierarchical approach that has found wide adoption for inorganic systems, relies on the production of crystalline nanoparticles which in turn become the building blocks of a next-level assembly process driven by oriented attachment (OA). As it stands, OA has not been observed for proteins. Here we employ cryoEM in the high nucleation rate limit of protein crystals and map the self-assembly route at molecular resolution. We observe the initial formation of facetted nanocrystals that merge lattices by means of OA alignment well before contact is made, satisfying non-trivial symmetry rules in the process. The OA mechanism yields crystal morphologies that are not attainable through conventional crystallization routes. Based on these insights we revisit a system of protein crystallization that has long been classified as non-classical, but our data is in direct conflict with that conclusion supporting a classical mechanism that implicates OA. These observations raise further questions about past conclusions for other proteins and illustrate the importance of maturation stages after primary nucleation has taken place.

biochemistry↗

bioRxiv2020

DC/L-SIGN recognition of spike glycoprotein promotes SARS-CoV-2 trans-infection and can be inhibited by a glycomimetic antagonist

The efficient spread of SARS-CoV-2 resulted in a pandemic that is unique in modern history. Despite early identification of ACE2 as the receptor for viral spike protein, much remains to be understood about the molecular events behind viral dissemination. We evaluated the contribution of C-type lectin receptors (CLRS) of antigen-presenting cells, widely present in air mucosa and lung tissue. DC-SIGN, L-SIGN, Langerin and MGL bind to diverse glycans of the spike using multiple interaction areas. Using pseudovirus and cells derived from monocytes or T-lymphocytes, we demonstrate that while virus capture by the CLRs examined does not allow direct cell infection, DC/L-SIGN, among these receptors, promote virus transfer to permissive ACE2+ cells. A glycomimetic compound designed against DC-SIGN, enable inhibition of this process. Thus, we described a mechanism potentiating viral capture and spreading of infection. Early involvement of APCs opens new avenues for understanding and treating the imbalanced innate immune response observed in COVID-19 pathogenesis

immunology↗

bioRxiv2020

In-cell quantitative structural imaging of phytoplankton using 3D electron microscopy

Phytoplankton is a minor fraction of the global biomass playing a major role in primary production and climate. Despite improved understanding of phytoplankton diversity and genomics, we lack nanoscale subcellular imaging approaches to understand their physiology and cell biology. Here, we present a complete Focused Ion Beam - Scanning Electron Microscopy (FIB-SEM) workflow (from sample preparation to image processing) to generate nanometric 3D phytoplankton models. Tomograms of entire cells, representatives of six ecologically-successful phytoplankton unicellular eukaryotes, were used for quantitative morphometric analysis. Besides lineage-specific cellular architectures, we observed common features related to cellular energy management: i) conserved cell-volume fractions occupied by the different organelles; ii) consistent plastid-mitochondria interactions, iii) constant volumetric ratios in these energy-producing organelles. We revealed detailed subcellular features related to chromatin organization and to biomineralization. Overall, this approach opens new perspectives to study phytoplankton acclimation responses to abiotic and biotic factors at a relevant biological scale.

plant biology↗

bioRxiv2020

Pre-initiation and elongation structures of full-length La Crosse virus polymerase reveal functionally important conformational changes

Bunyavirales is an order of segmented negative stranded RNA viruses comprising several life-threatening pathogens such as Lassa fever virus (Arenaviridae), Rift Valley Fever virus (Phenuiviridae) and La Crosse virus (LACV, Peribunyaviridae) against which neither specific treatment nor licenced vaccine is available. Replication and transcription of Bunyavirales genome constitute essential reactions of their viral cycle that are catalysed by the virally encoded RNA-dependent RNA polymerase or L protein. Here we describe the complete high-resolution cryo-EM structure of the full-length (FL) LACV-L protein. It reveals the presence of key C-terminal domains, notably the cap-binding domain that undergoes large movements related to its role in transcription initiation and a zinc-binding domain that displays a fold not previously observed. We capture the structure of LACV-L FL in two functionally relevant states, pre-initiation and elongation, that reveal large conformational changes inherent to its function. We uncover the coordinated movement of the polymerase priming loop, lid domain and C-terminal region required for the establishment of a ten-base-pair template-product RNA duplex before strand separation into respective exit tunnels. The revealed structural details and dynamics of functional elements will be instrumental for structure-based development of compounds that inhibit RNA synthesis by the polymerase.

biophysics↗

bioRxiv2020

The binding of palonosetron and other antiemetic drugs to the serotonin 5-HT3 receptor

Inaccurately perceived as niche drugs, antiemetics are key elements of cancer treatment alleviating the most dreaded side effect of chemotherapy. Serotonin 5-HT3 receptor antagonists are the most commonly prescribed class of drugs to control chemotherapy-induced nausea and vomiting (CINV). These antagonists have been clinically successful drugs since the 1980s, yet our understanding of how they operate at the molecular level has been hampered by the difficulty of obtaining structures of drug-receptor complexes. Here, we report the cryo-EM structure of the palonosetron-bound 5-HT3 receptor. We investigate the binding of palonosetron, granisetron, dolasetron, ondansetron, and cilansetron using molecular dynamics, covering the whole set of antagonists used in the clinical practice. The structural and computational results yield detailed atomic insight into the binding modes of the drugs. In light of our data, we establish a comprehensive framework underlying the inhibition mechanism by the -setron drug family.

neuroscience↗

bioRxiv2018

Integrated NMR and cryo-EM atomic-resolution structure determination of a half-megadalton enzyme complex

Atomic-resolution structure determination is the key requirement for understanding protein function. Cryo-EM and NMR spectroscopy both provide structural information, but currently cryo-EM does not routinely give access to atomic-level structural data, and, generally, NMR structure determination is restricted to small (<30 kDa) proteins. We introduce an integrated structure determination approach that simultaneously uses NMR and EM data to overcome the limits of each of these methods. The approach enabled determination of the high-resolution structure of the 468 kDa large dodecameric aminopeptidase TET2 to a precision and accuracy below 1 [A] by combining secondary-structure information obtained from near-complete magic-angle-spinning NMR assignments of the 39 kDa-large subunits, distance restraints from backbone amides and specifically labelled methyl groups, and a 4.1 [A] resolution EM map. The resulting structure exceeds current standards of NMR and EM structure determination in terms of molecular weight and precision. Importantly, the approach is successful even in cases where only medium-resolution (up to 8 [A]) cryo-EM data are available, thus paving avenues for the structure determination of challenging biological assemblies.

biophysics↗

bioRxiv2018

Cryo-EM structure of adenovirus type 3 fibre with desmoglein 2 shows a novel mode of receptor engagement

Attachment of adenovirus (HAd) to host cell is a critical step of infection. This work reports the cryo-electron microscopy (cryo-EM) structure of a non-symmetrical complex smaller than 100kDa formed by the trimeric human adenovirus of type 3 fibre knob (HAd3K) and human desmoglein 2 (DSG2). The structure reveals a unique stoichiometry, shedding light to new adenovirus infection strategies and providing new insights for adenoviral vector development.

microbiology↗