Virion attachment and entry: HIV gp120 Env biotinylation, gp120 Env, or integrin ligand-binding assay

The HIV-1 entry receptors are CD4 and a chemokine receptor (CCR5 or CXCR4). In addition it has recently been demonstrated that HIV-1 gp120 binds to and signals through integrin α4β7, the gut-homing receptor (Arthos et al., Nat Immunol 9(3):301-309, 2008). Integrin α4β7 is not an entry receptor for HIV-1, although it can facilitate virion attachment to target cells (Arthos et al., Nat Immunol 9(3):301-309, 2008; Cicala et al., Proc Natl Acad Sci U S A 106:20877-20882, 2009). Recombinant HIV-1 gp120s bind to integrin α4β7 in a manner similar to its natural ligands (MAdCAM-1, V-CAM-1, fibronectin) (Andrew et al., J Immunol 153:3847-3861, 1994). gp120-α4β7 interactions are detected in a manner similar to assays developed for the natural ligands of α4β7. In this chapter we describe a method for the analysis of integrin-gp120 binding via a cell-based binding assay. In vitro ligand-integrin affinity can be modified by the presence of divalent cations (Mn(2+), Mg(2+), Ca(2+)) (Leitinger et al., Leitinger Biochim Biophys Acta 1498:91-98, 2000). Here we describe a protocol to detect biotinylated recombinant HIV-1 gp120 binding to integrin α4β7 in both primary cells and cell lines expressing the gut-homing receptor.