Abstract
BackgroundThe glycan-targeting C-type DC-SIGN lectin receptor is implicated in the transmission of the human immunodeficiency virus (HIV) by binding the virus and transferring the captured HIV-1 to CD4+ T lymphocytes. Carbohydrate binding agents (CBAs) have been reported to block HIV-1 infection. We have now investigated the potent mannose-specific anti-HIV CBA griffithsin (GRFT) on its ability to inhibit the capture of HIV-1 to DC-SIGN, its DC-SIGN-directed transmission to CD4+ T-lymphocytes and the role of the three carbohydrate-binding sites (CBS) of GRFT in these processes.FindingsGRFT inhibited HIV-1(IIIB) infection of CEM and HIV-1(NL4.3) infection of C8166 CD4+ T-lymphocytes at an EC50 of 0.059 and 0.444 nM, respectively. The single mutant CBS variants of GRFT (in which a key Asp in one of the CBS was mutated to Ala) were about ∼20 to 60-fold less potent to prevent HIV-1 infection and ∼20 to 90-fold less potent to inhibit syncytia formation in co-cultures of persistently HIV-1 infected HuT-78 and uninfected C8166 CD4+ T-lymphocytes. GRFT prevents DC-SIGN-mediated virus capture and HIV-1 transmission to CD4+ T-lymphocytes at an EC50 of 1.5 nM and 0.012 nM, respectively. Surface plasmon resonance (SPR) studies revealed that wild-type GRFT efficiently blocked the binding between DC-SIGN and immobilized gp120, whereas the point mutant CBS variants of GRFT were ∼10- to 15-fold less efficient. SPR-analysis also demonstrated that wild-type GRFT and its single mutant CBS variants have the capacity to expel bound gp120 from the gp120-DC-SIGN complex in a dose dependent manner, a property that was not observed for HHA, another mannose-specific potent anti-HIV-1 CBA.ConclusionGRFT is inhibitory against HIV gp120 binding to DC-SIGN, efficiently prevents DC-SIGN-mediated transfer of HIV-1 to CD4+ T-lymphocytes and is able to expel gp120 from the gp120-DC-SIGN complex. Functionally intact CBS of GRFT are important for the optimal action of GRFT.
Highlights
Several studies have shown that carbohydrate-binding agents (CBAs) act as efficient inhibitors of human immunodeficiency virus (HIV) entry [1]
GRFT is inhibitory against HIV gp120 binding to Dendritic cells (DCs)-SIGN, efficiently prevents DC-SIGN-mediated transfer of HIV-1 to CD4+ T-lymphocytes and is able to expel gp120 from the gp120-DC-SIGN complex
The triple mutant (D30A/D70A/D112A) GRFT variant exhibited an approximately 400- to .900-fold diminished antiviral potency compared to wild-type GRFT with an EC50 of 52 nM and .185 nM for HIV-1(IIIB) and HIV-1(N.L4.3) infection in CEM and C8166 cell cultures, respectively
Summary
Several studies have shown that carbohydrate-binding agents (CBAs) act as efficient inhibitors of HIV entry [1] They block virus entry by inhibiting the fusion of HIV virions with their target cells. GRFT is a 121-amino acid dimeric lectin isolated from the red algae Griffithsia sp [6] It has a very potent and broad spectrum anti-HIV-1 activity showing nano- to picomolar inhibitory activities against cell-free virus infection and cell-to-cell transfer of HIV [6,7]. We have investigated the potent mannose-specific anti-HIV CBA griffithsin (GRFT) on its ability to inhibit the capture of HIV-1 to DC-SIGN, its DC-SIGNdirected transmission to CD4+ T-lymphocytes and the role of the three carbohydrate-binding sites (CBS) of GRFT in these processes
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