Regulation of immune responses by E3 ubiquitin ligase Cbl-b

Abstract

Casitas B lymphoma-b (Cbl-b), a RING finger E3 ubiquitin ligase, has been identified as a critical regulator of adaptive immune responses. Cbl-b is essential for establishing the threshold for T cell activation and regulating peripheral T cell tolerance through various mechanisms. Intriguingly, recent studies indicate that Cbl-b also modulates innate immune responses, and plays a key role in host defense to pathogens and anti-tumor immunity. These studies suggest that targeting Cbl-b may represent a potential therapeutic strategy for the management of human immune-related disorders such as autoimmune diseases, infections, tumors, and allergic airway inflammation. In this review, we summarize the latest developments regarding the roles of Cbl-b in innate and adaptive immunity, and immune-mediated diseases.

Keywords: Cbl-b; Immune-related disorders; Innate and adaptive immune responses; T cell tolerance; Ubiquitination.

Figure 1.. Functional domain structure of Cbl family proteins in mammals.

All three Cbl proteins share an N-terminal tyrosine kinase binding (TKB) region, composed of a four helix (4H) bundle, an EF Hand and an SH2 domain. The TKB domain is connected through a conserved helical linker (L) to a RING finger (RF) domain, which contributes to the E3 ligase activity. The C-terminal region includes proline-rich (PR) motifs, multiple serine and tyrosine phosphorylation sites, and leucine zipper (LZ)/ubiquitin association (UBA) domain. Cbl-3 lacks most of the C-terminal domains of c-Cbl and Cbl-b.

Figure 2.. Ubiquitination pathway related to E3 ubiquitin ligases.

Ubiquitin-activating (E1), ubiquitin-conjugating (E2) and ubiquitin-protein ligase (E3) enzymes are necessary for substrate ubiquitination. The process of ubiquitination is performed through an E1–E2–E3 cascade. The substrate protein can be ubiquitinated on a single or multiple lysine residues, resulting in monoubiquitination or multi-monoubiquitination respectively.

Figure 3.. Cbl-b in T-cell activation and tolerance.

PTEN is inactivated by Nedd4, which targets PTEN for K63-linked polyubiquitination in T cells upon TCR stimulation, and this process can be suppressed by Cbl-b. Nedd4 also ubiquitinates Cbl-b upon TCR stimulation. CD28-mediated inactivation of PTEN increases PIP₃, which recruits Akt, PDK-1 and Vav-1 to the plasma membrane through its association with the PH domains of these molecules. Activated Akt promotes the formation of CARMA1/Bcl-10/MALT1 (CBM) complex via phosphorylating CARMA1. Furthermore, Vav-1 links PKC-θ to PDK-1, then coupling IKKs to the CBM complex. Thus Cbl-b inhibits NF-κB activation through Akt and PKC-θ. In anergic T cells, Cbl-b induces ubiquitination of PLC-γ1 and PKC-θ, thereby inhibiting T cell anergy induction. Cbl-b expression in T cells is regulated via multiple mechanisms, CTLA-4-B7 interaction facilitates Cbl-b expression, while CD28 costimulation potentiates Cbl-b ubiquitination and proteasomal degradation, which is possibly mediated by Nedd4 and PKC-θ, and by auto-ubiquitination.

Figure 4.. Cbl-b in Th2/9 differentiation, iTreg development, and allergic airway inflammation.

Cbl-b selectively associates with Stat-6 and targets Stat-6 for ubiquitination and degradation upon TCR/CD28 and IL-4 stimulation. The negative regulation of Stat-6 by Cbl-b leads to inhibition of Th2/9 responses and allergic airway inflammation. Stub1 initiates Foxp3 ubiquitination, which in turn allows the recruitment of Cbl-b through its UBA domain, thus enhancing Foxp3 ubiquitination. Moreover, Cbl-b inhibits the activation of Akt-2, which phosphorylates Foxo1/Foxo3a, resulting in the exclusion of Foxo1/Foxo3a from the nuclei, whereas Foxo1/Foxo3a is required for the induction of Foxp3. The absence of Cbl-b impairs iTreg development, leading to aberrant Th2/9 response and severe allergic airway inflammation.

Figure 5.. Cbl-b in CLR signaling.

Cbl-b targets CLRs Dectin-1, Dectin-2, Dectin-3, and Syk for K48-linked polyubiquitination, leading to degradation of these molecules. Therefore, the downstream signaling mediated by CARD9/Bcl-10/MALT1 complex is inhibited, which results in the suppression of inflammatory responses against fungal pathogens in the presence of Cbl-b. Moreover, inhibition of Cbl-b by small inhibitory peptides or Cblb-specific siRNA can enhance innate antifungal immunity, thus providing a potential therapeutic strategy for fungal infections.