DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target

Crystallization and preliminary X-ray diffraction analysis of the cytosolic domain of a cation diffusion facilitator family protein

Selenomethionine-substituted crystals of the cytosolic domain of the cation diffusion facilitator family protein from T. maritima diffracted X-rays to 2.5 Å and belonged to space group R32, with unit-cell parameters a = b = 97.7, c = 83.4 Å

Novel Mg ²⁺ binding sites in the cytoplasmic domain of the MgtE Mg ²⁺ channels revealed by X-ray crystal structures

MgtE is a Mg 2+-selective channel regulated by the intracellular Mg 2+ concentration. MgtE family proteins are highly conserved in all domains of life and contribute to cellular Mg 2+ homeostasis. In humans, mutations in the SLC41 proteins, the eukaryotic counterparts of the bacterial MgtE, are known to be associated with various diseases. The first MgtE structure from a thermophilic bacterium, Thermus thermophilus, revealed that MgtE forms a homodimer consisting of transmembrane and cytoplasmic domains with a plug helix connecting the two and that the cytoplasmic domain possesses multiple Mg 2+ binding sites. Structural and electrophysiological analyses revealed that the dissociation of Mg 2+ ions from the cytoplasmic domain induces structural changes in the cytoplasmic domain, leading to channel opening. Thus, previous works showed the importance of MgtE cytoplasmic Mg 2+ binding sites. Nevertheless, due to the limited structural information on MgtE from different species, the conservation and diversity of the cytoplasmic Mg 2+ binding site in MgtE family proteins remain unclear. Here, we report crystal structures of the Mg 2+-bound MgtE cytoplasmic domains from two different bacterial species, Chryseobacterium hispalense and Clostridiales bacterium, and identify multiple Mg 2+ binding sites, including ones that were not observed in the previous MgtE structure. These structures reveal the conservation and diversity of the cytoplasmic Mg 2+ binding site in the MgtE family proteins

Crystallization and preliminary X-ray diffraction analysis of the cytosolic domain of the Mg2+ transporter MgtE

Crystals of the cytosolic domain of the Mg2+ transporter MgtE from T. thermophilus in the presence and absence of Mg2+ diffracted X-rays to 2.3 and 3.5 Å resolution, respectively, and belong to space groups P6522 (a = b = 57.7, c = 317.6 Å) and P212121 (a = 77.0, b = 100.3, c = 100.3 Å), respectively

Crystallization and preliminary X-ray diffraction analysis of the truncated cytosolic domain of the iron transporter FeoB

The truncated cytosolic domain of the iron transporter FeoB from T. maritima was overexpressed, purified and crystallized. Four native or SeMet crystal forms in a nucleotide-free state or in complex with either GDP or GMPPNP diffracted to resolutions of between 1.5 and 2.1 Å

Crystallization and preliminary X-ray diffraction analysis of the full-length Mg2+ transporter MgtE

Selenomethionine-substituted crystals of the Mg2+ transporter MgtE from T. thermophilus diffracted to 3.5 Å resolution and belonged to space group C2221, with unit-cell parameters a = 118.3, b = 134.9, c = 366.2 Å

Antithetic effects of agonists and antagonists on the structural fluctuations of TRPV1 channel

Transient receptor potential vanilloid member 1 (TRPV1) is a heat and capsaicin receptor that allows cations to permeate and cause pain. As the molecular basis for temperature sensing, the heat capacity (ΔCp) model [D. E. Clapham, C. Miller, Proc. Natl. Acad. Sci. U.S.A. 108, 19492–19497 (2011).] has been proposed and experimentally supported. Theoretically, heat capacity is proportional to a variance in enthalpy, presumably related to structural fluctuation; however, the fluctuation of TRPV1 has not been directly visualized. In this study, we directly visualized single-molecule structural fluctuations of the TRPV1 channels in a lipid bilayer with the ligands resiniferatoxin (agonist, 1,000 times hotter than capsaicin) and capsazepine (antagonist) by high-speed atomic force microscopy. We observed the structural fluctuations of TRPV1 in an apo state and found that RTX binding enhances structural fluctuations, while CPZ binding suppresses fluctuations. These ligand-dependent differences in structural fluctuation would play a key role in the gating of TRPV1

Mg2+-sensing mechanism of Mg2+ transporter MgtE probed by molecular dynamics study

Proper regulation of the intracellular ion concentration is essential to maintain life and is achieved by ion transporters that transport their substrates across the membrane in a strictly regulated manner. MgtE is a Mg2+ transporter that may function in the homeostasis of the intracellular Mg2+ concentration. A recent crystallographic study revealed that its cytosolic domain undergoes a Mg2+-dependent structural change, which is proposed to gate the ion-conducting pore passing through the transmembrane domain. However, the dynamics of Mg2+ sensing, i.e., how MgtE responds to the change in the intracellular Mg2+ concentration, remained elusive. Here we performed molecular dynamics simulations of the MgtE cytosolic domain. The simulations successfully reproduced the structural changes of the cytosolic domain upon binding or releasing Mg2+, as well as the ion selectivity. These results suggested the roles of the N and CBS domains in the cytosolic domain and their respective Mg2+ binding sites. Combined with the current crystal structures, we propose an atomically detailed model of Mg2+ sensing by MgtE