Entry of herpes simplex virus (HSV) 1 into cells requires the interaction of HSV gD with herpesvirus entry mediator or nectin1receptors, and fusion with cell membrane mediated by the fusion glycoproteins gB, gH, and gL. We report that the gD ectodomain in soluble form (amino acids 1-305) was sufficient to rescue the infectivity of a gD-null HSV mutant, indicating that gD does not need to be anchored to the virion envelope to mediate entry. Entry mediated by soluble gD required, in addition to the receptorbinding sites contained within residues 1-250, a discrete downstream portion (amino acids 261-305), located proximal to the transmembrane segment in full-length gD. We named it as profusion domain. The pro-fusion domain was required for entry mediated by virion-bound gD, because its substitution with the corresponding region of CD8 failed to complement the infectivity of gD ؊/؉ HSV. Furthermore, a receptor-negative gD (gD⌬6-259) inhibited virus infectivity when coexpressed with wild-type gD; i.e., it acted as a dominant-negative gD mutant. The pro-fusion domain is proline-rich, which is characteristic of regions involved in protein-protein interactions. P291L-P292A substitutions diminished the gD capacity to complement gD ؊/؉ HSV infectivity. We propose that gD forms a tripartite complex with its receptor and, by way of the proline-rich pro-fusion domain, with the fusion glycoproteins, or with one of them. The tripartite complex would serve to recruit͞activate the fusion glycoproteins and bring them from a fusion-inactive to a fusion-active state, such that they execute fusion of the virion envelope with cell membrane. H erpes simplex virus (HSV) enters cells through the coordinated action of four essential glycoproteins; gD, gB, gH, and gL, that act after the binding of gC and gB to the glycosaminoglycans of cell-surface proteoglycans (1, 2). Of the four glycoproteins required for entry, gD is the receptor-binding glycoprotein. It interacts with two alternative protein receptors, HVEM (herpesvirus entry mediator) and nectin1, that belong to the tumor necrosis factor receptor family (3), and to a growing family of intercellular adhesion molecules with Ig structure, respectively (4-8). The four essential glycoproteins required for HSV entry are required and are also sufficient to induce fusion of cells that express a gD receptor (9). The gD-binding site on HVEM maps mainly to the N-terminal cysteine-rich domain 1, with a hot spot at Y23 (10, 11). For nectin1, the N-terminal V domain, in particular its CCЈCЉ ridge (amino acids 64-104) is sufficient to mediate HSV entry (12-15). Critical residues were located in the 69-75 region and at positions 77 and 85 (16, 17). Insertion and deletion mutants in gD were the first mutants used to define functional regions (18). Subsequently, the x-ray crystal structure of the first 259 residues of gD [of the 315 that compose the ectodomain] was solved (19). The gD ectodomain is composed of an Ig-folded core (residues 56-184), with N-and C-terminal extensions. The latter folds bac...

Entry of herpes simplex virus into the cell requires the interaction of gD with one of its receptors, herpesvirus entry mediator or nectin 1, and the intervention of gB, gH, or gL, required to execute fusion of the virion envelope with cell membranes. The gD ectodomain is organized in two structurally and functionally differentiated regions. The N terminus (residues 1-260) carries the receptor binding sites, and the C terminus (residues 260 -310) functions as the pro-fusion domain (PFD), which is required for viral infectivity and fusion but not for receptor binding. The objective of our studies is to elucidate how gD links receptor recognition to the triggering of fusion. Here, we show that PFD is made of subdomains 1 and 2 (amino acids 260 -285 and 285-310). Each one partially contributed to herpes simplex virus infectivity. By means of glutathione S-transferase (GST) fusion proteins, we show that PFD bound soluble forms of gD, truncated at residue 260 (gD 260t) or downstream. Both PFD subdomains bound gD 260t, highlighting multiple contact sites between the N and C termini of gD. When gD 260t was in complex with either receptor, it failed to bind GST-PFD. In turn, the receptors did not bind GST-PFD, irrespective of whether they were in complex with gD. Thus, gD 260t interacted with the C terminus only if unbound to the receptor. We propose that (i) before receptor binding, gD adopts a ''closed'' conformation in which the N and C termini interact; and (ii) on encounter with a receptor, gD modifies its conformation and the N and C termini are released from reciprocal interactions (''opened'' conformation) and enabled to trigger fusion.

Objective-In the area of Malagrotta, a suburb of Rome (Italy), a large waste disposal site, a waste incinerator plant, and an oil refinery plant became operational in the early 1960s and have represented three major sources of air pollution. To evaluate the potential health risk due to airborne contamination around these point sources, a small area analysis of mortality was conducted. Cancer of the liver, larynx, lung, kidney, lymphatic, and haematopoietic systems were evaluated. Methods-Sex and age specific mortality (1987-93) and population denominators (1991) were available for the census tracts of the metropolitan area of Rome. Standardised mortality ratios (SMRs) were computed separately for males and females in bands of increasing distance from the plants, up to a radius of 10 km. Stone's test for the decline in risk with distance was performed with increments in radius of 1 km; SMRs were also computed after adjusting for a four level index of socioeconomic status. Results-No overall excess or decline in risk with distance was found for liver, lung, and lymphohaematopoietic cancers in either sex. For laryngeal cancer, an increased but not significant risk was found at 0-3 km and at 3-8 km. A significant decline with distance in mortality from laryngeal cancer was found among men (p=0.03); the trend remained after adjusting for the socioeconomic index (p=0.06). Conclusions-The study showed no association between proximity to the industrial sites and mortality for most of the several conditions considered. However, mortality from laryngeal cancer declined with distance from the sources of pollution. This result is interesting, as previous findings of an increased risk of laryngeal cancer near incinerators have been controversial. (Occup Environ Med 1998;55:611-615)

The identification of the AIP-RET complex represents a starting point to study key cellular processes involved in RET-induced apoptosis.