Calcium Regulation of Hemorrhagic Fever Virus Budding: Mechanistic Implications for Host-Oriented Therapeutic Intervention

Abstract

Hemorrhagic fever viruses, including the filoviruses (Ebola and Marburg) and arenaviruses (Lassa and Junín viruses), are serious human pathogens for which there are currently no FDA approved therapeutics or vaccines. Importantly, transmission of these viruses, and specifically late steps of budding, critically depend upon host cell machinery. Consequently, strategies which target these mechanisms represent potential targets for broad spectrum host oriented therapeutics. An important cellular signal implicated previously in EBOV budding is calcium. Indeed, host cell calcium signals are increasingly being recognized to play a role in steps of entry, replication, and transmission for a range of viruses, but if and how filoviruses and arenaviruses mobilize calcium and the precise stage of virus transmission regulated by calcium have not been defined. Here we demonstrate that expression of matrix proteins from both filoviruses and arenaviruses triggers an increase in host cytoplasmic Ca2+ concentration by a mechanism that requires host Orai1 channels. Furthermore, we demonstrate that Orai1 regulates both VLP and infectious filovirus and arenavirus production and spread. Notably, suppression of the protein that triggers Orai activation (Stromal Interaction Molecule 1, STIM1) and genetic inactivation or pharmacological blockade of Orai1 channels inhibits VLP and infectious virus egress. These findings are highly significant as they expand our understanding of host mechanisms that may broadly control enveloped RNA virus budding, and they establish Orai and STIM1 as novel targets for broad-spectrum host-oriented therapeutics to combat these emerging BSL-4 pathogens and potentially other enveloped RNA viruses that bud via similar mechanisms.

A cationic, C-terminal patch and structural rearrangements in Ebola virus matrix VP40 protein control its interactions with phosphatidylserine

“…PS selective binding modules include C2 domains (49) and some PH domains (50-52 (54). In fact, VP40 was sufficient to increase cellular levels of calcium in mammalian cells (55) and its role in promoting some degree of VP40 lipid binding cannot be ruled out at this time.…”

Screening of FDA-Approved Drugs for Inhibitors of Japanese Encephalitis Virus Infection

“…Among the five hit drugs, manidipine, cilnidipine, and benidipine hydrochloride were VGCC inhibitors. It has been well documented in the literature that Ca 2ϩ inhibitors serve to inhibit virus infection at the stage of either entry (15,22) or replication (18) and even at the stage of budding (23). To this end, we first reviewed all 21 calcium inhibitors included in the current library of FDA-approved drugs and found that, in addition to the four DHP VGCC inhibitors listed in Fig.…”

Host Calcium Channels and Pumps in Viral Infections

“…The research work done on four distinct hemorrhagic fever viruses (Ebolavirus (EBOV), Marburgvirus (MARV), Lassa Virus (LASV), and Junín Virus (JUNV)) demonstrated that EBOV, MARV VP40, and JUNV Z proteins trigger host cell Ca 2+ signals by activating the ER Ca 2+ sensing protein STIM1 and the plasma membrane ORAI1 channel. The STIM1/Orai1-mediated Ca 2+ signal is critical for EBOV and related viruses budding from host cells [35]. The suppression of STIM1 expression and genetic inactivation or the pharmacological blockade of ORAI1 inhibits infections of EBOV, MARV, and JUNV in cultured cells ( Figure 2B).…”

“…It is possible that the viral proteins trigger the Ca 2+ depletion in ER or prevent ER refilled with Ca 2+ to maintain resting ER Ca 2+ levels [40][41][42]. Alternately, these viral proteins may directly modify STIM1 and uncouple the activation of STIM1 from Ca 2+ levels [35,43]. Thus, STIM1/ORAI might represent a conserved target to regulating the budding of enveloped RNA viruses and possibly DNA viruses that rely on similar host cellular proteins for budding.…”