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Updated: Jan 19, 2026
In Vitro Analysis of E3 Ubiquitin Ligase Function
Published on: May 14, 2021
TAM receptors attenuate murine NK-cell responses via E3 ubiquitin ligase Cbl-b
Leilani M Chirino1, Suresh Kumar2, Mariko Okumura1
1Department of Pathology and Laboratory Medicine, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA, USA.
Abstract:
TAM receptors (Tyro3, Axl, and Mer) are receptor tyrosine kinases (RTKs) that are expressed by multiple immune cells including NK cells. Although RTKs typically enhance cellular functions, TAM receptor ligation blocks NK-cell activation. The mechanisms by which RTKs block NK-cell signaling downstream of activating receptors are unknown. In this report, we demonstrate that TAM receptors attenuate NK cell responses via the activity of E3 ubiquitin ligase Casitas B lineage lymphoma b (Cbl-b). Specifically, we show that Tyro3, Axl, and Mer phosphorylate Cbl-b, and Tyro3 ligation activates Cbl-b by phosphorylating tyrosine residues 133 and 363. Ligation of TAM receptors by their ligand Gas6 suppresses activating receptor-stimulated NK-cell functions such as IFN-γ production and degranulation, in a TAM receptor kinase- and Cbl-b-dependent manner. Moreover, Gas6 ligation induces the degradation of LAT1, a transmembrane adaptor protein required for NK cell activating receptor signaling, in WT but not in Cbl-b knock-out NK cells. Together, these results suggest that TAM receptors may attenuate NK-cell function by phosphorylating Cbl-b, which in turn dampens NK-cell activation signaling by promoting the degradation of LAT1. Our data therefore support a mechanism by which RTKs attenuate, rather than stimulate, signaling pathways via the activation of ubiquitin ligases.
Insights
TAM receptors block NK cell activation by activating the E3 ubiquitin ligase Cbl-b. This leads to the degradation of LAT1, dampening NK cell signaling and function.
Area of Science:
- Immunology
- Cellular Signaling
- Receptor Tyrosine Kinases
Background:
- TAM receptors (Tyro3, Axl, Mer) are receptor tyrosine kinases (RTKs) found on immune cells like NK cells.
- RTKs typically enhance cellular functions, but TAM receptors paradoxically block NK cell activation.
- The precise mechanisms of TAM receptor-mediated inhibition of NK cell signaling remain unclear.
Purpose of the Study:
- To elucidate the molecular mechanisms by which TAM receptors attenuate NK cell responses.
- To investigate the role of E3 ubiquitin ligase Casitas B lineage lymphoma b (Cbl-b) in TAM receptor signaling.
- To understand how TAM receptors impact NK cell activation downstream of activating receptors.
Main Methods:
- Phosphorylation analysis of Cbl-b by TAM receptors (Tyro3, Axl, Mer).
- Assessment of NK cell functions (IFN-γ production, degranulation) upon Gas6 ligand stimulation.
- Analysis of LAT1 degradation in wild-type and Cbl-b knock-out NK cells.
Main Results:
- TAM receptors phosphorylate Cbl-b; Tyro3 ligation specifically activates Cbl-b.
- Gas6 ligation suppresses NK cell functions in a TAM receptor kinase- and Cbl-b-dependent manner.
- Gas6 induces LAT1 degradation in wild-type NK cells, but not in Cbl-b knock-out cells.
Conclusions:
- TAM receptors attenuate NK cell function through Cbl-b activation.
- Cbl-b dampens NK cell activation signaling by promoting LAT1 degradation.
- RTKs can attenuate signaling pathways via ubiquitin ligase activation, contrary to typical RTK function.
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