The species D adenovirus E3/49K protein
Human adenoviruses (Ads) comprise more than 90 different types that are classified into 7 species, A-G. In immunocompetent individuals Ads typically induce self-limiting infections associated with acute disease, but can also establish persistent infections. In immunocompromised patients, Ad infections tend to be severe or even fatal. The disease pattern differs between Ad species. Species A, F and G are associated with gastroenteritis while species B, C, and E are mainly associated with respiratory diseases and species D with eye diseases. The majority of the known Ads belong to species D. However, only a few Ad types of species D, Ad8, Ad19a (recently renamed Ad64), Ad37, Ad53, Ad54 and Ad56 cause epidemic keratoconjunctivitis (EKC), a highly contagious and severe eye disease. The reasons for this association remain obscure. However, specific fiber knob sequences have been implicated in the association with EKC. Alternatively, the disease may be due to selective events occurring post-attachment, e.g. differential triggering of innate and adaptive immune responses or their differential manipulation by specific immunomodulatory functions encoded by EKC-causing Ads.
Such immunomodulatory functions are encoded in the early transcription unit 3 (E3). The E3 region is not required for Ad replication in vitro, yet is preserved in all human Ads, indicating an important role for virus-host interaction in vivo. Supporting this view, previous studies of species C Ad E3 proteins discovered multiple immune evasion mechanisms that seem to facilitate persistent infections. E3 is one of the most divergent regions of the Ad genome, differing considerably in size, gene composition and sequence between and within an Ad species. Species D Ads have the largest E3 region encoding eight open reading frames (ORFs). Of these, the E3/10.4K, 14.5K and 14.7K ORFs are present in all species and down-regulate various apoptosis receptors from the cell surface or affect their signaling, while E3/19K is only present in Ads of species B-E that do not cause gastroenteritis. E3/19K retains MHC class I molecules (MHC-I) and MHC-I related chain A and B in the endoplasmic reticulum (ER), thereby suppressing recognition by cytotoxic T-lymphocytes and natural killer (NK) cells. A few E3 genes are unique to a particular species and hence may allow for species-specific immunomodulation and differential disease outcome. One of these genes, designated E3/49K, is only present in species D Ads. The E3/49K ORF was initially identified in the E3 region of the EKC-causing Ad19a (recently renamed Ad64). Interestingly, E3/49K (also called CR1-beta) is the protein with the highest amino acid substitutions suggesting that it is under significant host immune pressure. E3/49K is a highly glycosylated type I transmembrane protein that migrates with an apparent molecular weight (mr) of 80-100 kDa and as such is the largest E3 protein identified so far. Ad19a E3/49K is abundantly synthesized in the early phase of infection but continues to be produced in the late phase albeit only with immature carbohydrates. The sequence of the extracellular domain revealed three internal repeats designated conserved region 1-3 (R1-3) that are predicted to form immunoglobulin-like domains.
E3/49K exhibits a novel processing pathway for E3 proteins. Approximately 1 h after synthesis it is cleaved by a matrix metalloprotease N-terminal to the transmembrane domain generating a small membrane-integrated 12-14 kDa C-terminal fragment and a large ectodomain (Sec49K) that is secreted or shed. Sec49K is the first secreted E3 protein and the first secreted Ad protein known to date. Unlike the other E3 proteins that act directly on infected cells, Sec49K can affect host immune functions over a distance by targeting leukocytes via binding to the cell surface tyrosine phosphatase CD45. This impairs the activation of CD4 T cells and NK cells, inhibiting cytokine production and cytotoxicity, respectively. Interestingly, the extracellular domain of CD45 is subject of a strong natural selection, which possibly is pathogen-driven. The intracellular parts of murine and human CD45 are 85.7% identical, whereas the identity for the extracellular domain is only 39.4%. Thus, the CD45 ECD and E3/49K hypervariability is most likely the outcome of a host-parasite co-evolution.