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Functional Interplay Between Murine Leukemia Virus Glycogag, Serinc5, and Surface Glycoprotein Governs Virus Entry, with Opposite Effects on Gammaretroviral and Ebolavirus Glycoproteins

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ABSTRACT

Gammaretroviruses, such as murine leukemia viruses (MLVs), encode, in addition to the canonical Gag, Pol, and Env proteins that will form progeny virus particles, a protein called “glycogag” (glycosylated Gag). MLV glycogag contains the entire Gag sequence plus an 88-residue N-terminal extension. It has recently been reported that glycogag, like the Nef protein of HIV-1, counteracts the antiviral effects of the cellular protein Serinc5. We have found, in agreement with prior work, that glycogag strongly enhances the infectivity of MLVs with some Env proteins but not those with others. In contrast, however, glycogag was detrimental to MLVs carrying Ebolavirus glycoprotein. Glycogag could be replaced, with respect to viral infectivity, by the unrelated S2 protein of equine infectious anemia virus. We devised an assay for viral entry in which virus particles deliver the Cre recombinase into cells, leading to the expression of a reporter. Data from this assay showed that both the positive and the negative effects of glycogag and S2 upon MLV infectivity are exerted at the level of virus entry. Moreover, transfection of the virus-producing cells with a Serinc5 expression plasmid reduced the infectivity and entry capability of MLV carrying xenotropic MLV Env, particularly in the absence of glycogag. Conversely, Serinc5 expression abrogated the negative effects of glycogag upon the infectivity and entry capability of MLV carrying Ebolavirus glycoprotein. As Serinc5 may influence cellular phospholipid metabolism, it seems possible that all of these effects on virus entry derive from changes in the lipid composition of viral membranes.

No MeSH data available.


Y36A mutant of gGag is partially active in enhancing MLV(Xeno) infectivity. Specific infectivity of MLV(Xeno) with wild-type Gag-Pol (blue bar) or with mutant Gag-Pol lacking gGag (red bar), and MLV(Xeno) with mutant Gag-Pol cotransfected with increasing amounts of wild-type (green bars) or Y36A mutant gGag (purple bars) pCMV(glycogag). The wild-type or Y36A mutant gGag/Gag-Pol plasmid ratios used were increased by threefold increments from 1:243 to 1:3. The target cell line used in these experiments was HT1080/mCAT1. *, P < 0.0001.
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fig6: Y36A mutant of gGag is partially active in enhancing MLV(Xeno) infectivity. Specific infectivity of MLV(Xeno) with wild-type Gag-Pol (blue bar) or with mutant Gag-Pol lacking gGag (red bar), and MLV(Xeno) with mutant Gag-Pol cotransfected with increasing amounts of wild-type (green bars) or Y36A mutant gGag (purple bars) pCMV(glycogag). The wild-type or Y36A mutant gGag/Gag-Pol plasmid ratios used were increased by threefold increments from 1:243 to 1:3. The target cell line used in these experiments was HT1080/mCAT1. *, P < 0.0001.

Mentions: As mentioned above, glycogag has the ability to complement the Nef defect in Nef− HIV-1. Usami et al. reported that this activity is reduced or lost if tyrosine 36 (in the glycogag-specific region of glycogag) is replaced with alanine (Y36A mutation) (24). We introduced the Y36A mutation into pCMV(glycogag) and tested the ability of the mutant to enhance MLV(Xeno) infectivity. The results in Fig 6 show the effects of dilution series of wild-type and Y36A glycogag plasmids upon the specific infectivity of MLV(Xeno) lacking glycogag. While it is difficult to compare the two titrations precisely, the data show clearly that the mutant glycogag retains partial activity in this assay. As a 1:9 dilution of the Y36A plasmid (Fig 6, fourth purple bar) had an effect similar to the 1:81 dilution of the wild-type plasmid (Fig 6, second green bar), perhaps the mutant is ~1/10 as active as wild-type glycogag.


Functional Interplay Between Murine Leukemia Virus Glycogag, Serinc5, and Surface Glycoprotein Governs Virus Entry, with Opposite Effects on Gammaretroviral and Ebolavirus Glycoproteins
Y36A mutant of gGag is partially active in enhancing MLV(Xeno) infectivity. Specific infectivity of MLV(Xeno) with wild-type Gag-Pol (blue bar) or with mutant Gag-Pol lacking gGag (red bar), and MLV(Xeno) with mutant Gag-Pol cotransfected with increasing amounts of wild-type (green bars) or Y36A mutant gGag (purple bars) pCMV(glycogag). The wild-type or Y36A mutant gGag/Gag-Pol plasmid ratios used were increased by threefold increments from 1:243 to 1:3. The target cell line used in these experiments was HT1080/mCAT1. *, P < 0.0001.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC5120145&req=5

fig6: Y36A mutant of gGag is partially active in enhancing MLV(Xeno) infectivity. Specific infectivity of MLV(Xeno) with wild-type Gag-Pol (blue bar) or with mutant Gag-Pol lacking gGag (red bar), and MLV(Xeno) with mutant Gag-Pol cotransfected with increasing amounts of wild-type (green bars) or Y36A mutant gGag (purple bars) pCMV(glycogag). The wild-type or Y36A mutant gGag/Gag-Pol plasmid ratios used were increased by threefold increments from 1:243 to 1:3. The target cell line used in these experiments was HT1080/mCAT1. *, P < 0.0001.
Mentions: As mentioned above, glycogag has the ability to complement the Nef defect in Nef− HIV-1. Usami et al. reported that this activity is reduced or lost if tyrosine 36 (in the glycogag-specific region of glycogag) is replaced with alanine (Y36A mutation) (24). We introduced the Y36A mutation into pCMV(glycogag) and tested the ability of the mutant to enhance MLV(Xeno) infectivity. The results in Fig 6 show the effects of dilution series of wild-type and Y36A glycogag plasmids upon the specific infectivity of MLV(Xeno) lacking glycogag. While it is difficult to compare the two titrations precisely, the data show clearly that the mutant glycogag retains partial activity in this assay. As a 1:9 dilution of the Y36A plasmid (Fig 6, fourth purple bar) had an effect similar to the 1:81 dilution of the wild-type plasmid (Fig 6, second green bar), perhaps the mutant is ~1/10 as active as wild-type glycogag.

View Article: PubMed Central - PubMed

ABSTRACT

Gammaretroviruses, such as murine leukemia viruses (MLVs), encode, in addition to the canonical Gag, Pol, and Env proteins that will form progeny virus particles, a protein called &ldquo;glycogag&rdquo; (glycosylated Gag). MLV glycogag contains the entire Gag sequence plus an 88-residue N-terminal extension. It has recently been reported that glycogag, like the Nef protein of HIV-1, counteracts the antiviral effects of the cellular protein Serinc5. We have found, in agreement with prior work, that glycogag strongly enhances the infectivity of MLVs with some Env proteins but not those with others. In contrast, however, glycogag was detrimental to MLVs carrying Ebolavirus glycoprotein. Glycogag could be replaced, with respect to viral infectivity, by the unrelated S2 protein of equine infectious anemia virus. We devised an assay for viral entry in which virus particles deliver the Cre recombinase into cells, leading to the expression of a reporter. Data from this assay showed that both the positive and the negative effects of glycogag and S2 upon MLV infectivity are exerted at the level of virus entry. Moreover, transfection of the virus-producing cells with a Serinc5 expression plasmid reduced the infectivity and entry capability of MLV carrying xenotropic MLV Env, particularly in the absence of glycogag. Conversely, Serinc5 expression abrogated the negative effects of glycogag upon the infectivity and entry capability of MLV carrying Ebolavirus glycoprotein. As Serinc5 may influence cellular phospholipid metabolism, it seems possible that all of these effects on virus entry derive from changes in the lipid composition of viral membranes.

No MeSH data available.