In severe viral pneumonia, including Coronavirus disease 2019 (COVID-19), the viral replication phase is often followed by hyperinflammation, which can lead to acute respiratory distress syndrome, multi-organ failure, and death. We previously demonstrated that alpha-1 adrenergic receptor (⍺-AR) antagonists can prevent hyperinflammation and death in mice. Here, we conducted retrospective analyses in two cohorts of patients with acute respiratory distress (ARD, n = 18,547) and three cohorts with pneumonia (n = 400,907). Federated across two ARD cohorts, we find that patients exposed to ⍺-AR antagonists, as compared to unexposed patients, had a 34% relative risk reduction for mechanical ventilation and death (OR = 0.70, p = 0.021). We replicated these methods on three pneumonia cohorts, all with similar effects on both outcomes. All results were robust to sensitivity analyses. These results highlight the urgent need for prospective trials testing whether prophylactic use of ⍺-AR antagonists ameliorates lower respiratory tract infection-associated hyperinflammation and death, as observed in COVID-19.

For a more complete understanding of molecular mechanisms, it is important to study macromolecules and their assemblies in the broader context of the cell. This context can be visualized at nanometer resolution in three dimensions (3D) using electron cryo-tomography, which requires tilt series to be recorded and computationally aligned, currently limiting throughput. Additionally, the high-resolution signal preserved in the raw tomograms is currently limited by a number of technical difficulties, leading to an increased false-positive detection rate when using 3D template matching to find molecular complexes in tomograms. We have recently described a 2D template matching approach that addresses these issues by including high-resolution signal preserved in single-tilt images. A current limitation of this approach is the high computational cost that limits throughput. We describe here a GPU-accelerated implementation of 2D template matching in the image processing software TEM that allows for easy scaling and improves the accessibility of this approach. We apply 2D template matching to identify ribosomes in images of frozen-hydrated cells with high precision and sensitivity, demonstrating that this is a versatile tool for visual proteomics and structure determination. We benchmark the results with 3D template matching of tomograms acquired on identical sample locations and identify strengths and weaknesses of both techniques, which offer complementary information about target localization and identity.

Fold-switching proteins challenge the one-sequence-one-structure paradigm by adopting multiple stable folds. Nevertheless, it is uncertain whether fold switchers are naturally pervasive or rare exceptions to the well-established rule. To address this question, we developed a predictive method and applied it to the NusG superfamily of >15,000 transcription factors. We predicted that a substantial population (25%) of the proteins in this family switch folds. Circular dichroism and nuclear magnetic resonance spectroscopies of 10 sequence-diverse variants confirmed our predictions. Thus, we leveraged family-wide predictions to determine both conserved contacts and taxonomic distributions of fold-switching proteins. Our results indicate that fold switching is pervasive in the NusG superfamily and that the single-fold paradigm significantly biases structure-prediction strategies.

Hippocampal place cells contribute to mammalian spatial navigation and memory formation. Numerous models have been proposed to explain the location-specific firing of this cognitive representation, but the pattern of excitatory synaptic input leading to place firing is unknown, leaving no synaptic-scale explanation of place coding. Here we used resonant scanning two-photon microscopy to establish the pattern of synaptic glutamate input received by CA1 place cells in behaving mice. During traversals of the somatic place field, we found increased excitatory dendritic input, mainly arising from inputs with spatial tuning overlapping the somatic field, and functional clustering of this input along the dendrites over ~10 µm. These results implicate increases in total excitatory input and co-activation of anatomically clustered synaptic input in place firing. Since they largely inherit their fields from upstream synaptic partners with similar fields, many CA1 place cells appear to be part of multi-brain-region cell assemblies forming representations of specific locations.