Functional Analysis of the Putative Fusion Domain of the Baculovirus Envelope Fusion Protein F

Westenberg, Marcel; Veenman, Frank; Roode, Els C.; Goldbach, Rob; Vlak, Just M.; Zuidema, D.

doi:10.1128/jvi.78.13.6946-6954.2004

Cited by 32 publications

(30 citation statements)

References 44 publications

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“…Dm-cF, Ac23 and Op21 all lack the highly conserved region that contains a furin cleavage site and the fusion peptide. Thus, the absence of detectable membrane fusion activity in Dm-cF, Ac23, and Op21 is consistent with previous observations that an intact furin cleavage site and fusion peptide are both important for pH-induced membrane fusion activity in the SeMNPV and LdMNPV F proteins (42,48,49). We cannot however rule out the possibility that Dm-cF is an intracellular fusion protein with fusion activity regulated by mechanisms other than those tested in our assays.…”

Section: Discussionsupporting

confidence: 76%

“…Thus, these downstream ATG codons may serve as initiator codons by leaky scanning (24). Unlike the baculovirus F proteins from group I NPVs and GVs (those considered "fusion competent") (22,39,42,48,49), the F proteins from all three Drosophila species lack (i) a predicted furin cleavage site, (ii) a predicted fusion peptide, and (iii) a predicted coiled-coil domain. In contrast, the predicted Anopheles protein, Ag-cF, does have a potential furin cleavage site RTKR (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Mutational analyses of these two F proteins suggest that both the furin cleavage site and a fusion peptide are necessary for membrane fusion activity (40,42,48,49). However, F proteins from group I NPVs, AcMNPV and OpMNPV (viruses with both GP64 and F), do not contain a predicted furin cleavage site, and low pH-triggered membrane fusion activity was not observed when these F proteins were examined in standard syncytium formation assays (reference 41 and unpublished observations).…”

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confidence: 99%

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A Cellular Drosophila melanogaster Protein with Similarity to Baculovirus F Envelope Fusion Proteins

Lung

Blissard

2005

J Virol

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Baculovirus F (fusion) proteins are found in the envelopes of budded virions. Recently a Drosophila melanogaster gene (CG4715) that encodes a protein with sequence similarity to baculovirus F proteins was discovered. To examine similarities and differences with baculovirus F proteins, we cloned the D. melanogaster cellular F (Dm-cF) protein gene and analyzed Dm-cF expression and localization. The predicted Dm-cF protein sequence lacks a furin cleavage site, and transiently expressed Dm-cF showed no protein cleavage and no detectable membrane fusion activity. In cell localization studies, transiently expressed Dm-cF was localized to intracellular organelles in D. melanogaster S2 cells, unlike baculovirus F proteins, which localize to cellular plasma membranes. Using reverse transcriptase PCR and Western blot analysis to examine Dm-cF expression in animals, we detected Dm-cF expression in both larval and adult D. melanogaster cells. However, Dm-cF expression was detected only in third instar larvae and adults, suggesting that Dm-cF expression may be developmentally regulated. We also identified genes related to Dm-cF in the genomes of two other Drosophila species, Drosophila yakuba and Drosophila pseudoobscura, and the mosquito Anopheles gambiae. These observations suggest that f genes may be present in the genomes of many insects. Conservation within and between 22 baculovirus and 4 insect F proteins was examined in detail. These studies demonstrate that Dm-cF is expressed in D. melanogaster and suggest that if baculovirus f genes are derived from a host cellular f gene, the function appears to have changed substantially upon adaptation to the baculovirus infection cycle.

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Section: Discussionsupporting

confidence: 76%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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A Cellular Drosophila melanogaster Protein with Similarity to Baculovirus F Envelope Fusion Proteins

Lung

Blissard

2005

J Virol

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“…1b), or into the gp64-null AcMNPV bacmid (Lung et al, 2002), according to the Bac-to-Bac manual (Invitrogen). Transposition was confirmed by PCR as described previously (Westenberg et al, 2004).…”

Section: Methodsmentioning

confidence: 99%

GP64 of group I nucleopolyhedroviruses cannot readily rescue infectivity of group II f-null nucleopolyhedroviruses

Westenberg

Vlak

2008

Journal of General Virology

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The genus Nucleopolyhedrovirus (NPV) of the family Baculoviridae can be subdivided phylogenetically into two groups. The same division can be made on the basis of their budded virus (BV) envelope fusion protein. Group I NPVs are characterized by the presence of a GP64-like major envelope fusion protein, which is involved in viral attachment and the fusion of virus and cell membrane, and is required for budding of progeny nucleocapsids. Group II NPVs have an envelope fusion protein unrelated to GP64, named F. In contrast to GP64, F proteins are found in all baculoviruses, but they are not functional as envelope fusion proteins in group I NPVs. Autographa californica multiple NPV (AcMNPV) lacking GP64 can be pseudotyped by the F protein of Spodoptera exigua multiple NPV (SeMNPV), suggesting that F proteins are functionally analogous to GP64. GP64 homologues are thought to have been acquired by group I NPVs during evolution, thereby giving these viruses a selective advantage and obviating the need for a functional F protein. To address this supposition experimentally, attempts were made to pseudotype a group II NPV, SeMNPV, with GP64. Transfection of an f-null SeMNPV bacmid into Se301 cells did not result in the production of infectious BVs. This defect was rescued by insertion of SeMNPV f, but not by insertion of AcMNPV gp64. This suggests that the functional analogy between GP64 and F is not readily reciprocal and that F proteins from group II NPVs may provide additional functions in BV formation that are lacking in the GP64 type of fusion protein.

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“…Baculovirus F proteins are Nglycosylated and are found as homotrimers on the BV particle (28). A putative fusion peptide located on the N terminus of the F 1 subunit was reported to be critical to the biological function of Spodoptera exigua MNPV F (51). Furthermore, heptad repeat (HR) regions were predicted in baculovirus F proteins, with HR1 downstream of the fusion peptide region and HR2 upstream of the transmembrane domain ( Fig.…”

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confidence: 99%

Conserved Leucines in N-Terminal Heptad Repeat HR1 of Envelope Fusion Protein F of Group II Nucleopolyhedroviruses Are Important for Correct Processing and Essential for Fusogenicity

Long

Pan

Vlak

2008

J Virol

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The heptad repeat (HR), a conserved structural motif of class I viral fusion proteins, is responsible for the formation of a six-helix bundle structure during the envelope fusion process. The insect baculovirus F protein is a newly found budded virus envelope fusion protein which possesses common features to class I fusion proteins, such as proteolytic cleavage and the presence of an N-terminal open fusion peptide and multiple HR domains on the transmembrane subunit F 1 . Similar to many vertebrate viral fusion proteins, a conserved leucine zipper motif is predicted in this HR region proximal to the fusion peptide in baculovirus F proteins. To facilitate our understanding of the functional role of this leucine zipper-like HR1 domain in baculovirus F protein synthesis, processing, and viral infectivity, key leucine residues (Leu209, Leu216, and Leu223) were replaced by alanine (A) or arginine (R), respectively. By using Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) as a pseudotype expression system, we demonstrated that all mutant F proteins incorporated into budded virus, indicating that leucine substitutions did not affect intercellular trafficking of F. Furin-like protease cleavage was not affected by any of the leucine substitutions; however, the disulfide bridging and N-linked glycosylation patterns were partly altered. Single substitutions in HR1 showed that the three leucine residues were critical for F fusogenicity and the rescue of AcMNPV infectivity. Our results support the view that the leucine zipper-like HR1 domain is important to safeguard the proper folding, glycosylation, and fusogenicity of baculovirus F proteins.

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Functional Analysis of the Putative Fusion Domain of the Baculovirus Envelope Fusion Protein F

Cited by 32 publications

References 44 publications

A Cellular Drosophila melanogaster Protein with Similarity to Baculovirus F Envelope Fusion Proteins

A Cellular Drosophila melanogaster Protein with Similarity to Baculovirus F Envelope Fusion Proteins

GP64 of group I nucleopolyhedroviruses cannot readily rescue infectivity of group II f-null nucleopolyhedroviruses

Conserved Leucines in N-Terminal Heptad Repeat HR1 of Envelope Fusion Protein F of Group II Nucleopolyhedroviruses Are Important for Correct Processing and Essential for Fusogenicity

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