Membrane-associated RING-CH-type 8 (MARCH8) strongly blocks human immunodeficiency virus type 1 (HIV-1) envelope
glycoprotein (Env) incorporation into virions by downregulating its cell surface expression, but the mechanism is still unclear. We now report that MARCH8 also blocks the Ebola virus (EBOV)
glycoprotein (GP) incorporation via surface downregulation. To understand how these
viral fusion proteins are downregulated, we investigated the effects of MARCH8 on EBOV GP maturation and externalization via the conventional secretion pathway. MARCH8 interacted with EBOV GP and
furin when detected by immunoprecipitation and retained the GP/
furin complex in the Golgi when their location was tracked by a bimolecular fluorescence complementation (BiFC) assay. MARCH8 did not reduce the GP expression or affect the GP modification by high-
mannose N-
glycans in the endoplasmic reticulum (ER), but it inhibited the formation of complex N-
glycans on the GP in the Golgi. Additionally, the GP O-glycosylation and
furin-mediated proteolytic cleavage were also inhibited. Moreover, we identified a novel
furin cleavage site on EBOV GP and found that only those fully glycosylated GPs were processed by
furin and incorporated into virions. Furthermore, the GP shedding and secretion were all blocked by MARCH8. MARCH8 also blocked the
furin-mediated cleavage of HIV-1 Env (gp160) and the highly pathogenic
avian influenza virus H5N1
hemagglutinin (HA). We conclude that MARCH8 has a very broad
antiviral activity by prohibiting different
viral fusion proteins from glycosylation and proteolytic cleavage in the Golgi, which inhibits their transport from the Golgi to the plasma membrane and incorporation into virions.IMPORTANCE Enveloped viruses express three classes of fusion
proteins that are required for their entry into host cells via mediating virus and cell membrane fusion. Class I fusion
proteins are produced from influenza viruses, retroviruses, Ebola viruses, and coronaviruses. They are first synthesized as a type I transmembrane
polypeptide precursor that is subsequently glycosylated and oligomerized. Most of these precursors are cleaved en route to the plasma membrane by a cellular
protease furin in the late secretory pathway, generating the trimeric N-terminal receptor-binding and C-terminal fusion subunits. Here, we show that a cellular
protein, MARCH8, specifically inhibits the
furin-mediated cleavage of EBOV GP, HIV-1 Env, and H5N1 HA. Further analyses uncovered that MARCH8 blocked the EBOV GP glycosylation in the Golgi and inhibited its transport from the Golgi to the plasma membrane. Thus, MARCH8 has a very broad
antiviral activity by specifically inactivating different
viral fusion proteins.