Business as usual: the p35 isoform of human CD74 retains function in antigen presentation.

CD74 (invariant chain, li) is most renowned for its role in preparing major histocompatibility complex (MHC) class II molecules for peptide presentation to CD4+ T helper cells.1, 2 Although we have known for some time that various CD74 isoforms exist, so far attempts to attribute distinct functions to each isoform have failed.3 The human-specific CD74 isoform called p35 contains a retention motif for the endoplasmatic reticulum (ER), suggesting that p35 may be limited in terms of trafficking within the cell. However, on page 896 in this issue of Immunology and Cell Biology Genève et al.4 show that p35 substitutes efficiently for CD74 in supporting antigen presentation to T cells in mice, indicating that p35 retains the antigen presenting functions of other CD74 isoforms despite its ER retention motif. In antigen presenting cells, CD74 facilitates export of MHC II/CD74 complexes from the ER while blocking premature loading of peptides on to the MHC II molecules (Figure 1, far left). Two di-leucine sorting motifs in the CD74 cytoplasmic tail direct MHC II/CD74 complexes towards endosomes, in which CD74 is partially degraded to leave only the CLIP (Class II associated invariant chain peptide) fragment in the peptide binding groove of MHC II. HLA-DM (H2-DM in mice) then exchanges CLIP for new peptides to be presented by MHC II on the cell surface.1 The absence of CD74 in mice causes low MHC II expression levels and floppy MHC II molecules5 concomitant with reduced thymic and peripheral CD4 T-cell numbers.6 CD74-deficient mice therefore provide an ideal environment in which CD74 variants can be systematically ‘added back’ to probe for distinct functions. Of all CD74 isoforms, the human-specific p35 molecule is the most enigmatic. Although in humans 80% of the CD74 protein pool is composed of the p33 isoform, use of an alternative translation initiation site in p35 adds an R-X-R motif,7 reported to cause ER retention of this CD74 isoform in the absence of MHC II.8 Although it is thought this motif is masked when MHC II binds CD74, allowing MHC II/CD74 complexes to egress from the ER,9 it was unclear whether p35 alone would substitute effectively should the other CD74 isoforms be absent. Genève et al.4 addressed this question by introducing human p35 under the control of the CD74 promoter into mice that lack endogenous CD74. As expected, p35 was expressed in antigen presenting cells and inducible by interferon gamma. The p35 transgene partially reinstated MHC class II expression by thymic epithelial cells, but fully restored the numbers and surface TCR levels of thymic and peripheral CD4+ T cells in mice lacking endogenous CD74, and the peripheral CD4+ T cells were found to express a diverse array of Vβ TCR chains. The p35 transgene also fully restored iNKT-cell formation in the thymus. Bone marrow-derived macrophages that only expressed p35 presented a CD74-dependent ovalbumin-derived epitope to antigen-specific T-cell hybridomas and stimulated IL-2 production. However, macrophages derived from p35 expressing mice only achieved intermediate stimulation of the T-cell line and these mice failed to make a detectable antibody response to OVA protein. The p35 transgene also failed to restore the deletion of superantigen-reactive Vβ5-expressing CD4+ T cells. Taken together the results suggest that p35 fulfils the functions of other CD74 isoforms in T-cell-positive selection and peripheral maintenance, however, certain functions may be lost. The authors attribute these functional shortcomings to fewer antigen presenting cells expressing p35 in this system. Given the evidence that p35’s ER retention motif is concealed by binding to MHC class II molecules,9 it is perhaps not surprising that p35 behaves similarly to isoforms lacking this motif once it is bound to MHC II. It might be highly informative, therefore, to explore p35’s capacity to carry out MHC II-independent roles proposed for CD74 (Figure 1); such as acting as a receptor for macrophage migration inhibitory factor, induction of vesicle biogenesis, regulation of dendritic cell migration and activation of the nuclear factor κB pathway.10 The invariant nature of CD74, as opposed to the polymorphic MHC II, and its central contribution to antigen presentation make CD74 an attractive target for therapies. Although distinct capacities have been proposed for its isoforms,8, 11, 12 in vivo functions of the various human and mouse CD74 isoforms have been difficult to distinguish. Genève et al.4 have shown that despite its ER retention motif human p35 fulfills the functions of murine CD74 in several MHC II-dependent processes. The niggling question that remains is: can the CD74 isoforms fully substitute for each other or are our current experimental approaches just unable to differentiate the essential role of each molecule?