16 17 ABSTRACT 18 Virus and host factors contribute to cell-to-cell variation in viral infections and determine the 19 outcome of the overall infection. However, the extent of the variability at the single cell level and 20 how it impacts virus-host interactions at a systems level are not well understood. To characterize 21 the dynamics of viral transcription and host responses, we used single-cell RNA sequencing to 22 quantify at multiple time points the host and viral transcriptomes of human A549 cells and primary 23 bronchial epithelial cells infected with influenza A virus. We observed substantial variability of viral 24 transcription between cells, including the accumulation of defective viral genomes (DVGs) that 25 impact viral replication. We show a correlation between DVGs and viral-induced variation of the 26 host transcriptional program and an association between differential induction of innate immune 27 response genes and attenuated viral transcription in subpopulations of cells. These observations 28 at the single cell level improve our understanding of the complex virus-host interplay during 29 influenza infection.30 31 IMPORTANCE 32 Defective influenza virus particles generated during viral replication carry incomplete viral 33 genomes and can interfere with the replication of competent viruses. These defective genomes 34 are thought to modulate disease severity and pathogenicity of the influenza infection. Different 35 defective viral genomes also introduce another source of variation across a heterogeneous cell 36 population. Evaluating the impact of defective virus genomes on host cell responses cannot be 37 fully resolved at the population level, requiring single cell transcriptional profiling. Here we 38 characterized virus and host transcriptomes in individual influenza-infected cells, including that of 39 defective viruses that arise during influenza A virus infection. We established an association 40 between defective virus transcription and host responses and validated interfering and 41 immunostimulatory functions of identified dominant defective virus genome species in vitro. This 42 study demonstrates the intricate effects of defective viral genomes on host transcriptional 43 responses and highlights the importance of capturing host-virus interactions at the single-cell level. 44 45 82 Cell-to-cell variation in virus gene expression. 83 To determine how both the viral and host cell transcriptional programs relate to each other over 84 the course of an influenza infection, we infected two cell types-the adenocarcinomic human 85 alveolar basal epithelial A549 cell line and human primary bronchial epithelial cells HBEpC-at 86 high multiplicity of Infection (MOI; 5) with A/Puerto Rico/8/34(H1N1) (PR8) and performed single 87 cell and bulk RNA-seq expression analyses. A high MOI infection ensures that virtually all the 88 cells can be rapidly infected, promotes the accumulation of DVGs, and consequently enables the 89 characterization of both host response and DVG diversity. We first determi...

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Costimulation pathways play an essential role in T cell activation, differentiation, and regulation. CD155 expressed on antigen-presenting cells (APCs) interacts with TIGIT, an inhibitory costimulatory molecule, and CD226, an activating costimulatory molecule, on T cells. TIGIT and CD226 are expressed at varying levels depending on the T cell subset and activation state. T follicular helper cells in germinal centers (GC-Tfh) in human tonsils express high TIGIT and low CD226. However, the biological role of the CD155/TIGIT/CD226 axis in human Tfh cell biology has not been elucidated. To address this, we analyzed tonsillar CD4+ T cell subsets cultured with artificial APCs constitutively expressing CD155. Here we show that CD226 signals promote the early phase of Tfh cell differentiation in humans. CD155 signals promoted the proliferation of naïve CD4+ T cells and Tfh precursors (pre-Tfh) isolated from human tonsils and upregulated multiple Tfh molecules and decreased IL-2, a cytokine detrimental for Tfh cell differentiation. Blocking CD226 potently inhibited their proliferation and expression of Tfh markers. By contrast, while CD155 signals promoted the proliferation of tonsillar GC-Tfh cells, their proliferation required only weak CD226 signals. Furthermore, attenuating CD226 signals rather increased the expression of CXCR5, ICOS, and IL-21 by CD155-stimulated GC-Tfh cells. Thus, the importance of CD226 signals changes according to the differentiation stage of human Tfh cells and wanes in mature GC-Tfh cells. High TIGIT expression on GC-Tfh may play a role in attenuating the detrimental CD226 signals post GC-Tfh cell maturation.

In BriefHIV evolved a minimalist but robust transcriptional circuit bypassing host regulatory checkpoints; however, the fragility of the circuit in the host phase (which primes HIV for activation) largely affects proviral transcription and fate. Highlights• The host and viral phases of the HIV transcriptional circuit have different functional requirements • HIV evolved a minimalist program to robustly bypass host cell regulatory checkpoints • A mathematical model reveals that the host phase is subject to transcriptional circuit fragility • Host transcriptional circuit fragility influences the viral feedback and latency reversal potential

Background: Tryptophan synthase consists of two subunits, D and E. Two distinct subgroups of E chain exist. The major group (TrpEb_1) includes the well-studied E chain of Salmonella typhimurium. The minor group of E chain (TrpEb_2) is most frequently found in the Archaea. Most of the amino-acid residues important for catalysis are highly conserved between both TrpE subfamilies.