Single cell tuning of Myc expression by antigen receptor signal strength and interleukin‐2 in T lymphocytes

Abstract

Myc controls the metabolic reprogramming that supports effector T cell differentiation. The expression of Myc is regulated by the T cell antigen receptor (TCR) and pro-inflammatory cytokines such as interleukin-2 (IL-2). We now show that the TCR is a digital switch for Myc mRNA and protein expression that allows the strength of the antigen stimulus to determine the frequency of T cells that express Myc. IL-2 signalling strength also directs Myc expression but in an analogue process that fine-tunes Myc quantity in individual cells via post-transcriptional control of Myc protein. Fine-tuning Myc matters and is possible as Myc protein has a very short half-life in T cells due to its constant phosphorylation by glycogen synthase kinase 3 (GSK3) and subsequent proteasomal degradation. We show that Myc only accumulates in T cells exhibiting high levels of amino acid uptake allowing T cells to match Myc expression to biosynthetic demands. The combination of digital and analogue processes allows tight control of Myc expression at the population and single cell level during immune responses.

Suboptimal T-cell receptor signaling compromises protein translation, ribosome biogenesis, and proliferation of mouse CD8 T cells

“…The results are in agreement with our prior understanding of Myc regulation in T cells, which is driven by TCR signals upon antigen encounter and maintained by IL-2 signaling at later stages (30). It is likely that weakly stimulated T cells are unable to produce sufficient IL-2 to sustain expression of Myc and, consequently, its target genes involved in ribosome biogenesis.…”

“…STAT5 is activated by IL-2 and has been shown to regulate the expression of various cyclins and CDKs required for cell cycle progression through the G1 and S phases (52). IL-2 signaling is also very important for maintaining Myc translation because Myc protein, but not mRNA, is lost when cytotoxic T cells (CTLs) are incubated with the JAK inhibitor tofacitinib (30,53). Preston et al showed that proinflammatory cytokines are required to maintain Myc protein abundance because, in T cells, these cytokines sustain expression of amino acid transporters, which in turn support the amino acid uptake required to replenish rapidly turning over Myc protein (30).…”

“…Initially Myc expression in T cells is driven exclusively by TCR engagement (30), and, at 24 h, Myc was upregulated more strongly in N4-stimulated than in G4-stimulated cells and was not restored to N4 levels by addition of exogenous IL-2. By 72 h, however, when continued Myc expression in T cells is largely sustained by cytokine signaling (27,30), the addition of IL-2 to G4 cells had a significant rescue effect on Myc abundance (Fig. 7B).…”

“…7A), with the percentage of G0/1, S, and G2/M cells comparable with WT cells without exogenous IL-2. Because IL-2 is required for sustained expression of Myc in T cells (27,30) and many key genes involved in rRNA processing and ribosome assembly are targets of Myc regulation (31, 32), we measured by FACS the intracellular level of Myc and one of its targets, fibrillarin, a site-specific methyltransferase required for prerRNA processing. Initially Myc expression in T cells is driven exclusively by TCR engagement (30), and, at 24 h, Myc was upregulated more strongly in N4-stimulated than in G4-stimulated cells and was not restored to N4 levels by addition of exogenous IL-2.…”

“…Because IL-2 is required for sustained expression of Myc in T cells (27,30) and many key genes involved in rRNA processing and ribosome assembly are targets of Myc regulation (31, 32), we measured by FACS the intracellular level of Myc and one of its targets, fibrillarin, a site-specific methyltransferase required for prerRNA processing. Initially Myc expression in T cells is driven exclusively by TCR engagement (30), and, at 24 h, Myc was upregulated more strongly in N4-stimulated than in G4-stimulated cells and was not restored to N4 levels by addition of exogenous IL-2. By 72 h, however, when continued Myc expression in T cells is largely sustained by cytokine signaling (27,30), the addition of IL-2 to G4 cells had a significant rescue effect on Myc abundance (Fig.…”