Kinetics of cytokine receptor trafficking determine signaling and functional selectivity

Martínez-Fábregas, Jonathan; Wilmes, Stephan; Wang, L.; Hafer, Maximillian; Pöhler, Elizabeth; Lokau, Juliane; Garbers, Christoph; Cozzani, Adeline; Piehler, Jacob; Kazemian, Majid; Mitra, Suman; Moraga, Ignacio

doi:10.1101/638031

Cited by 11 publications

(33 citation statements)

References 76 publications

(84 reference statements)

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“…We postulated that the weak binding affinity that IL-10 exhibits for IL-10Rβ critically contributed to its functional fitness, by limiting the range of concentrations at which IL-10 elicits its full immunomodulatory potential. Here, we engineered variants of IL-10 with enhanced affinity for IL-10Rβ to investigate whether the stability of the IL- reported that IL-6, another STAT3 activating cytokine, promotes strong STAT3 binding to chromatin but poor gene expression (30). Our study was done in resting monocytes.…”

Section: Discussionmentioning

confidence: 99%

“…Overall these observations suggest that in addition to receptor binding affinity, the stoichiometry of the IL-10-receptor complex helps to fine-tune IL-10 bioactivity potencies. We showed that the number of phospho-tyrosines available in cytokine receptor intracellular domains critically contributes to defining signalling identity by cytokines (30). IL-6 variants that triggered partial phosphorylation of Tyr residues available in the gp130 intracellular domain exhibited a biased STAT3 versus STAT1 activation (30).…”

Section: Discussionmentioning

confidence: 99%

“…Monomeric wild type IL-10 (25), monomeric high affinity variants and IL-10Rα ectodomain (amino acids 22-235) were cloned and expressed as described in (30). Briefly, protein sequences were cloned into the pAcGP67-A vector (BD Biosciences) in frame with an N-terminal gp67 signal sequence, driving protein secretion, and a C-terminal hexahistidine tag.…”

Section: Materials and Methods: Protein Expression And Purificationmentioning

confidence: 99%

“…These nanobodies were conjugated to photostable organic fluorophores ATTO Rho11 and ATTO 643, which are suitable for simultaneous dual-colour single molecule tracking of IL-10Rα and IL-10Rβ in the plasma membrane of live cells as shown for other cytokine receptor systems (30)(31)(32) ( Fig. 2a and fig.…”

Section: Enhanced Il-10rβ Binding Affinity Increases Receptor Heterodmentioning

confidence: 99%

“…The yeast surface display protocol was adapted from previous protocols (30,61). To create an IL-10 yeast display library, the monomeric IL-10 gene (25) was subject to error-prone PCR as previously described (28).…”

Section: Yeast Display Librarymentioning

confidence: 99%

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Engineered IL-10 variants elicit potent immunomodulatory effects at low ligand doses

et al. 2020

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Interleukin-10 (IL-10) is a dimeric cytokine with both immunosuppressive and immunostimulatory activities; however, IL-10–based therapies have shown only marginal clinical benefits. Here, we explored whether the stability of the IL-10 receptor complex contributes to the immunomodulatory potency of IL-10. We generated an IL-10 mutant with enhanced affinity for its IL-10Rβ receptor using yeast surface display. Compared to the wild-type cytokine, the affinity-enhanced IL-10 variants recruited IL-10Rβ more efficiently into active cell surface signaling complexes and triggered greater STAT1 and STAT3 activation in human monocytes and CD8+ T cells. These effects, in turn, led to more robust induction of IL-10–mediated gene expression programs at low ligand concentrations in both human cell subsets. IL-10–regulated genes are involved in monocyte energy homeostasis, migration, and trafficking and in CD8+ T cell exhaustion. At nonsaturating doses, IL-10 did not induce key components of its gene expression program, which may explain its lack of efficacy in clinical settings. Our engineered IL-10 variant showed a more robust bioactivity profile than that of wild-type IL-10 at low doses in monocytes and CD8+ T cells. Moreover, CAR-modified T cells expanded with the engineered IL-10 variant displayed superior cytolytic activity than those expanded with wild-type IL-10. Our study provides insights into how IL-10 receptor complex stability fine-tunes IL-10 biology and opens new opportunities to revitalize failed IL-10 therapies.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Materials and Methods: Protein Expression And Purificationmentioning

confidence: 99%

Section: Enhanced Il-10rβ Binding Affinity Increases Receptor Heterodmentioning

confidence: 99%

Section: Yeast Display Librarymentioning

confidence: 99%

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Engineered IL-10 variants elicit potent immunomodulatory effects at low ligand doses

et al. 2020

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Making sense of IL‐6 signalling cues in pathophysiology

et al. 2021

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Unravelling the molecular mechanisms that account for functional pleiotropy is a major challenge for researchers in cytokine biology. Cytokine–receptor cross‐reactivity and shared signalling pathways are considered primary drivers of cytokine pleiotropy. However, reports epitomized by studies of Jak‐STAT cytokine signalling identify interesting biochemical and epigenetic determinants of transcription factor regulation that affect the delivery of signal‐dependent cytokine responses. Here, a regulatory interplay between STAT transcription factors and their convergence to specific genomic enhancers support the fine‐tuning of cytokine responses controlling host immunity, functional identity, and tissue homeostasis and repair. In this review, we provide an overview of the signalling networks that shape the way cells sense and interpret cytokine cues. With an emphasis on the biology of interleukin‐6, we highlight the importance of these mechanisms to both physiological processes and pathophysiological outcomes.

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The discriminatory power of the T cell receptor

Huhn

Wilson

Merwe

et al. 2020

Preprint

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T cells use their T-cell receptors (TCRs) to discriminate between lower-affinity self and higher-affinity non-self pMHC antigens. The strength of this discrimination and the mechanisms that produce it remain controversial. Although a large number of mouse and human TCRs have now been characterised, they have not been used to precisely quantitate discrimination. Here, we systematically quantify the discrimination of TCRs using a discrimination power (α). Early influential studies on three mouse TCRs suggested that discrimination was nearly perfect (α ~ 9.0). In striking contrast, our analysis of published data on other mouse and human TCRs, and more recent data on the original mouse TCRs, produced significantly lower discrimination (α = 2.0). Although not perfect, the discriminatory power of TCR was greater than that of conventional receptors such as cytokine receptors, GPCRs, RTKs, and CARs (α ≤ 1). The revised discriminatory power of the TCR is readily explained by a kinetic proofreading mechanisms, and accounts for the ability of low affinity self-antigens to stimulate autoimmune and anti-tumour T cell responses.

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Kinetics of cytokine receptor trafficking determine signaling and functional selectivity

Cited by 11 publications

References 76 publications

Engineered IL-10 variants elicit potent immunomodulatory effects at low ligand doses

Engineered IL-10 variants elicit potent immunomodulatory effects at low ligand doses

Making sense of IL‐6 signalling cues in pathophysiology

The discriminatory power of the T cell receptor

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