Endoproteolytic Processing of the Ebola Virus Envelope Glycoprotein: Cleavage Is Not Required for Function

Abstract

Proteolytic processing is required for the activation of numerous viral glycoproteins. Here we show that the envelope glycoprotein from the Zaire strain of Ebola virus (Ebo-GP) is proteolytically processed into two subunits, GP1 and GP2, that are likely covalently associated through a disulfide linkage. Murine leukemia virions pseudotyped with Ebo-GP contain almost exclusively processed glycoprotein, indicating that this is the mature form of Ebo-GP. Mutational analysis identified a dibasic motif, reminiscent of furin-like protease processing sites, as the Ebo-GP cleavage site. However, analysis of Ebo-GP processing in LoVo cells that lack the proprotein convertase furin demonstrated that furin is not required for processing of Ebo-GP. In sharp contrast to other viral systems, we found that an uncleaved mutant of Ebo-GP was able to mediate infection of various cell lines as efficiently as the wild-type, proteolytically cleaved glycoprotein, indicating that cleavage is not required for the activation of Ebo-GP despite the conservation of a dibasic cleavage site in all filoviral envelope glycoproteins.

Cathepsins B and L activate Ebola but not Marburg virus glycoproteins for efficient entry into cell lines and macrophages independent of TMPRSS2 expression

“…Our results show that TMPRSS2 and the related protease hepsin can cleave REBOV-and/or CIEBOV-GP, potentially at the same site as furin. ZEBOV-GP and SEBOV-GP are presumably also cleaved by TMPRSS2 but cleavage could not be demonstrated because of the efficient processing of these proteins by endogenous furin in 293T cells (Wool-Lewis and Bates, 1999). However, cleavage by TMPRSS2 and hepsin did not result in GP activation and cathepsin B/L-independent cellular entry, in agreement with previous findings indicating that the furin cleavage site is dispensable for viral spread and pathogenicity (Neumann et al, 2002(Neumann et al, , 2007.…”

“…Both EBOV-GP and HIV Env are cleaved by furin in the Golgi apparatus of infected cells (Hallenberger et al, 1992;Volchkov et al, 1998) and cleavage at the furin consensus site is essential for HIV infectivity (McCune et al, 1988). In contrast, GP cleavage by furin is dispensable for EBOV spread in cell culture and in animals (Neumann et al, 2002(Neumann et al, , 2007Wool-Lewis and Bates, 1999), indicating that furin does not activate GP. Work by Chandran et al (2005) and subsequent studies (Kaletsky et al, 2007;Schornberg et al, 2006) suggest that ZEBOV-GP is activated by the pH-dependent cysteine proteases cathepsin B and L (termed cathepsins B/L in the remainder of the manuscript) upon viral entry into host cell endosomes.…”

Furin cleavage of the SARS coronavirus spike glycoprotein enhances cell–cell fusion but does not affect virion entry

“…A similar paradigm has been proposed for entry by the Ebola filovirus (Chandran et al, 2005). As in MHV, furin cleavage of the Ebola virus envelope glycoprotein precursor occurs in nature yet appears unnecessary for viral entry (Neumann et al, 2002;Wool-Lewis and Bates, 1999).…”

Ebola Viral Glycoprotein Bound to Its Endosomal Receptor Niemann-Pick C1

“…Like all the other Class I viral fusion proteins, the EBOV GP is synthesized as a single polypeptide that is subsequently cleaved by furin-like proteases into GP1 and GP2 subunits, which remain together through an inter-subunit disulfide bond and non-covalent interactions. In contrast to other Class I viral fusion proteins, however, the simple furin cleavage event is not sufficient to prime EBOV GP (Neumann et al, 2007;Wool-Lewis and Bates, 1999). After entering into the cell, the virus is eventually trafficked to late endosomes, where GP is further primed to remove some ''cap'' components, thereby triggering the induction of the crucial membrane fusion event, which leads to viral penetration.…”