Nur77-Tempo mice reveal T cell steady state antigen recognition

Abstract

In lymphocytes, <i>Nr4a</i> gene expression is specifically regulated by antigen receptor signalling, making them ideal targets for use as distal T cell receptor (TCR) reporters. <i>Nr4a3</i>-Timer of cell kinetics and activity (Tocky) mice are a ground-breaking tool to report TCR-driven <i>Nr4a3</i> expression using Fluorescent Timer protein (FT). FT undergoes a time-dependent shift in its emission spectrum following translation, allowing for the temporal reporting of transcriptional events. Our recent work suggested that <i>Nr4a1</i>/Nur77 may be a more sensitive gene to distal TCR signals compared to <i>Nr4a3</i>, so we, therefore, generated Nur77-Timer-rapidly-expressed-in-lymphocytes (Tempo) mice that express FT under the regulation of Nur77. We validated the ability of Nur77-Tempo mice to report TCR and B cell receptor signals and investigated the signals regulating Nur77-FT expression. We found that Nur77-FT was sensitive to low-strength TCR signals, and its brightness was graded in response to TCR signal strength. Nur77-FT detected positive selection signals in the thymus, and analysis of FT expression revealed that positive selection signals are often persistent in nature, with most thymic Treg expressing FT Blue. We found that active TCR signals in the spleen are low frequency, but CD69⁺ lymphoid T cells are enriched for FT Blue⁺ Red⁺ T cells, suggesting frequent TCR signalling. In non-lymphoid tissue, we saw a dissociation of FT protein from CD69 expression, indicating that tissue residency is not associated with tonic TCR signals. Nur77-Tempo mice, therefore, combine the temporal dynamics from the Tocky innovation with increased sensitivity of <i>Nr4a1</i> to lower TCR signal strengths.