Mind the gap from research laboratory to clinic: Challenges and opportunities for next-generation assays in human diseases.

Modern vaccinology has experienced major conceptual and technological advances over the past 30 years. These include atomic-level structures driving immunogen design, new vaccine delivery methods, powerful adjuvants, and novel animal models. In addition, utilizing advanced assays to learn how the immune system senses a pathogen and orchestrates protective immunity has been critical in the design of effective vaccines and therapeutics. The National Institute of Allergy and Infectious Diseases of the National Institutes of Health convened a workshop in September 2020 focused on next generation assays for vaccine development (Table 1). The workshop focused on four critical pathogens: severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) and human immunodeficiency virus (HIV)-which have no licensed vaccines-and tuberculosis (TB) and influenza-both of which are in critical need of improved vaccines. The goal was to share progress and lessons learned, and to identify any commonalities that can be leveraged to design vaccines and therapeutics.

Trans-omics Impact of Thymoproteasome in Cortical Thymic Epithelial Cells.

The thymic function to produce self-protective and self-tolerant T cells is chiefly mediated by cortical thymic epithelial cells (cTECs) and medullary TECs (mTECs). Recent studies including single-cell transcriptomic analyses have highlighted a rich diversity in functional mTEC subpopulations. Because of their limited cellularity, however, the biochemical characterization of TECs, including the proteomic profiling of cTECs and mTECs, has remained unestablished. Utilizing genetically modified mice that carry enlarged but functional thymuses, here we show a combination of proteomic and transcriptomic profiles for cTECs and mTECs, which identified signature molecules that characterize a developmental and functional contrast between cTECs and mTECs. Our results reveal a highly specific impact of the thymoproteasome on proteasome subunit composition in cTECs and provide an integrated trans-omics platform for further exploration of thymus biology.

TCR ITAM multiplicity is required for the generation of follicular helper T-cells.

The T-cell antigen receptor (TCR) complex contains 10 copies of a di-tyrosine Immunoreceptor-Tyrosine-based-Activation-Motif (ITAM) that initiates TCR signalling by recruiting protein tyrosine kinases. ITAM multiplicity amplifies TCR signals, but the importance of this capability for T-cell responses remains undefined. Most TCR ITAMs (6 of 10) are contributed by the CD3ζ subunits. We generated 'knock-in' mice that express non-signalling CD3ζ chains in lieu of wild-type CD3ζ. Here we demonstrate that ITAM multiplicity is important for the development of innate-like T-cells and follicular helper T-cells, events that are known to require strong/sustained TCR-ligand interactions, but is not essential for 'general' T-cell responses including proliferation and cytokine production or for the generation of a diverse antigen-reactive TCR repertoire.

CD5 helps aspiring regulatory T cells ward off unwelcome cytokine advances.

The role of co-receptor molecules in the generation of inducible regulatory T cells (iTregs) remains incompletely defined. In this issue of Immunity, Henderson et al. (2015) show that CD5 regulates iTreg cell induction by rendering emerging iTreg cells refractory to signals mediated by effector-differentiating cytokines.

Human neonatal naive CD4+ T cells have enhanced activation-dependent signaling regulated by the microRNA miR-181a.

Compared with older children and adults, human neonates have reduced and delayed CD4(+) T cell immunity to certain pathogens, but the mechanisms for these developmental differences in immune function remain poorly understood. We investigated the hypothesis that impaired human neonatal CD4(+) T cell immunity was due to reduced signaling by naive CD4(+) T cells following engagement of the αß-TCR/CD3 complex and CD28. Surprisingly, calcium flux following engagement of CD3 was significantly higher in neonatal naive CD4(+) T cells from umbilical cord blood (CB) compared with naive CD4(+) T cells from adult peripheral blood. Enhanced calcium flux was also observed in adult CD4(+) recent thymic emigrants. Neonatal naive CD4(+) T cells also had higher activation-induced Erk phosphorylation. The microRNA miR-181a, which enhances activation-induced calcium flux in murine thymocytes, was expressed at significantly higher levels in CB naive CD4(+) T cells compared with adult cells. Overexpression of miR-181a in adult naive CD4(+) T cells increased activation-induced calcium flux, implying that the increased miR-181a levels of CB naive CD4(+) T cells contributed to their enhanced signaling. In contrast, AP-1-dependent transcription, which is downstream of Erk and required for full T cell activation, was decreased in CB naive CD4(+) T cells compared with adult cells. Thus, CB naive CD4(+) T cells have enhanced activation-dependent calcium flux, indicative of the retention of a thymocyte-like phenotype. Enhanced calcium signaling and Erk phosphorylation are decoupled from downstream AP-1-dependent transcription, which is reduced and likely contributes to limitations of human fetal and neonatal CD4(+) T cell immunity.