Human LAMP1 accelerates Lassa virus fusion and potently promotes fusion pore dilation upon forcing viral fusion with non-endosomal membrane
Authors: Zhang You , de la Torre Juan Carlos , Melikyan Gregory B. 
Published: June 2022

AbstractLassa virus (LASV) cell entry is mediated by the interaction of the virus glycoprotein complex (GPC) with alpha-dystroglycan at the cell surface followed by binding to LAMP1 in late endosomes. However, LAMP1 is not absolutely required for LASV fusion, as this virus can infect LAMP1-deficient cells. Here, we used LASV GPC pseudoviruses, LASV virus-like particles and recombinant lymphocytic choriomeningitis virus expressing LASV GPC to investigate the role of human LAMP1 (hLAMP1) in LASV fusion with human and avian cells expressing a LAMP1 ortholog that does not support LASV entry. We employed a combination of single virus imaging and virus population-based fusion and infectivity assays to dissect the hLAMP1 requirement for initiation and completion of LASV fusion that culminates in the release of viral ribonucleoprotein into the cytoplasm. Unexpectedly, ectopic expression of hLAMP1 accelerated the kinetics of small fusion pore formation, but only modestly increased productive LASV fusion and infection of human and avian cells. To assess the effects of hLAMP1 in the absence of requisite endosomal host factors, we forced LASV fusion with the plasma membrane by applying low pH. Unlike the conventional LASV entry pathway, ectopic hLAMP1 expression dramatically promoted the initial and full dilation of pores formed through forced fusion at the plasma membrane. We further show that, while the soluble hLAMP1 ectodomain accelerates the kinetics of nascent pore formation, it fails to promote efficient pore dilation, suggesting the hLAMP1 transmembrane domain is involved in this late stage of LASV fusion. These findings reveal a previously unappreciated role of hLAMP1 in promoting dilation of LASV fusion pores, which is difficult to ascertain for endosomal fusion where several co-factors, such as bis(monoacylglycero)phosphate, likely regulate LASV entry.Author SummaryLassa virus (LASV) enters cells via fusion with acidic endosomes mediated by the viral glycoprotein complex (GPC) interaction with the intracellular receptor LAMP1. However, the requirement for LAMP1 is not absolute, as LASV can infect avian cells expressing a LAMP1 ortholog that does not interact with GPC. To delineate the role of LAMP1 in LASV entry, we developed assays to monitor the formation of nascent fusion pores, as well as their initial and complete dilation to sizes that allow productive infection of avian cells by LASV GPC pseudoviruses. This novel approach provided unprecedented details regarding the dynamics of LASV fusion pores and revealed that ectopic expression of human LAMP1 in avian cells leads to a marked acceleration of fusion but modestly increases the likelihood of complete pore dilation and infection. In contrast, human LAMP1 expression dramatically enhanced the propensity of nascent pores to fully enlarge when LASV fusion with the plasma membrane was forced by exposure to low pH. We conclude that, whereas the role of LAMP1 in LASV fusion is confounded by an interplay between multiple endosomal factors, the plasma membrane is a suitable target for mechanistic dissection of the roles of endosomal factors in LASV entry.