CTLA-4 trafficking and surface expression

Abstract

The T-cell co-receptor cytotoxic T-cell antigen 4 (CTLA-4) has a strong inhibitory role as shown by the lymphoproliferative phenotype of CTLA-4-deficient mice. Despite its potent effects on T-cell function, CTLA-4 is primarily an intracellular antigen whose surface expression is tightly regulated by restricted trafficking to the cell surface and rapid internalisation. Recently, several signalling molecules such as Trim, PLD, ARF-1 and TIRC7 have been described to be involved in the transport of CTLA-4 to the cell surface. Minor changes in surface expression levels have major effects on the outcome of T-cell activation. Optimal regulation of CTLA-4 surface expression is crucial for the balance of stimulatory and inhibitory signals to maximize protective immune responses while maintaining immunological tolerance and preventing autoimmunity.

Figure 1. Cytotoxic T-cell antigen 4 (CTLA-4) function at the T cell–antigen-presenting cell (APC) interface.

Both CD28 and the inhibitory co-receptor CTLA-4 bind to B7–1 (CD80) and B7–2 (CD86) molecules on APCs. The cytoplasmic tail of CTLA-4 contains two tyrosine-based motifs. The YVKM motif in the intracellular tail constitutes a binding site for the lipid kinase PI3K, the phosphatases PP2A and SHP2 and the clathrin adaptor proteins AP-1 and AP-2.

Figure 2. Cytotoxic T-cell antigen 4 (CTLA-4) structure.

(a) The CTLA-4 protein consists of three domains and a leader peptide. The leader peptide corresponds to exon 1 in the CTLA-4 gene, the ligand binding domain to exon 2, the transmembrane region to exon 3 and the cytoplasmic tail to exon 4. Four different isoforms of CTLA-4 protein can be generated by differential splicing: full length CTLA-4 (exons 1, 2, 3 and 4), soluble CTLA-4 (exons 1, 2 and 4), ligand independent CTLA-4 (exons 1, 3 and 4, murine only) and an isoform consisting of exons 1 and 4 only. (b) The gene structure of CTLA-4 contains four exons, two untranslated regions preceding exon 1 and downstream of exon 4 and a promoter region up to −335 bp. Several gene polymorphisms described within the CTLA-4 gene have been connected with autoimmune diseases. Some of these are indicated on the figure [e.g. in exon 1 A49G and (AT)_n in the −3′ untranslated region (UTR)].

Figure 3. Model describing the regulation of cytotoxic T-cell antigen 4 (CTLA-4) surface expression.

This model summarizes the known mechanisms involved in CTLA-4 surface expression. Newly synthesized CTLA-4 binds to the transmembrane adaptor TRIM in the TGN promoting its release to the cell surface. CTLA-4 externalisation is also dependent on PLD and ARF-1 activity. Alternatively, binding to clathrin adaptor protein AP-1 mediates direct transport to lysosomal compartments. Surface residency of CTLA-4 is stabilized by phosphorylation of tyrosine 201 in the YVKM motif, which also allows for PI3K, SHP2 and PP2A binding and therefore CTLA-4 signalling. Clathrin adaptor AP-2 binds to dephosphorylated tyrosine 201 and mediates internalisation of CTLA-4 and its transport to endosomes and lysosomes. Upon T-cell activation, CTLA-4 enriched lysosomes are released to the cell surface. ARF-1, GTPase ADP ribolysation factor-1; PLD, phospholipase D; TRIM, T-cell receptor interacting molecule; TGN, trans Golgi network.