Crystal structures of human TIM members: Ebolavirus entry-enhancing receptors

Human T-cell immunoglobulin and mucin domain (hTIM) receptor was recently identified as the entry-enhancing receptor for various enveloped viruses, including ebolavirus. This hTIM-mediated virus entry is highly dependent on the immunoglobulin variable (IgV)-like domain of the receptor and on the specific interactions between hTIM IgV and the phosphotidylserine (PS) moieties on the viral envelope. There are three members in the hTIM family, including hTIM-1,-3, and-4. However, their IgV domain structures and the PS binding basis remain elusive thus far, though their murine homologues have been structurally well defined. In addition, it is suspected that the 2014-Zaire-ebolavirus, which has caused the largest pandemic ever recorded last year in West Africa, might have different pathogenic capacity from the earlier isolates. Nevertheless, it remains uninvestigated if the 2014-Zaire-ebolavirus can recognize hTIMs to enhance virus infection similarly to the previously tested 1976-Zaire-ebolavirus. In this study, we first showed that pseudoviruses incorporating either the 1976-or the 2014-Zaire-ebolavirus glycoprotein (GP) exhibit similarly enhanced cell entry via hTIM-1 and-4, but not with hTIM-3. Though 2014-GP mutations are suspected giving rise to a higher virulence, analogous entry rate was observed for 1976-and 2014-GP. We further showed that the hTIM receptors do not directly interact with either the mucin-deleted or the mucin-reserved intact ebolavirus GP, and that hTIM-1 and-4 engage envelope PS to facilitate the viral infection. We then solved all the three hTIM IgV-domain structures as well as the hTIM-4/phosphoserine complex structure. Several unexpected regional features, which have not been observed in any of the mouse homologues, were identified in the individual PS-binding clefts of the three hTIMs, including a shortened FG loop in hTIM-1, a novel metal binding site in hTIM-3, and an alternative conformation for the CC loop in hTIM-4. These characteristics astonishingly indicated variant PS-recognition basis established by individual hTIMs that is different from that of the mouse receptors. Taken together, our observations will merit our understanding of the binding basis between hTIMs and PS, thereby shedding light on the entry mechanisms of ebolavirus and other related enveloped viruses.