HIV-tat peptide–mediated loading of peptides corresponding to β3-integrin cytoplasmic tail promotes PMN transmatrix migration.
In panel A, isolated human PMNs were pre-exposed to indicated concentrations of HIV-tat peptides motifs corresponding to β3-integrin cytoplasmic tyrosine 747 (closed circles), tyrosine 759 (closed squares), or scrambled peptide for 20 minutes at room temperature followed by examination of fMLP-stimulated (10−8 M) transmigration across matrix-coated substrates. The immunofluorescence insets for each peptide localize HIV-tat peptide within permeabilized (P) and nonpermeabilized (NP) PMNs following peptide loading (see “Materials and methods”). Arrowheads demonstrate predominant localization of peptides at the cytoplasmic face of PMNs. Transmigrated PMNs were quantified by determination of MPO. Data are derived from 9 monolayers in each condition from 4 separate experiments with results expressed as mean ± SEM number transmigrating PMNs. (B) The β3-integrin cytoplasmic sequence is a kinase substrate. Coincubation of HIV-tat peptides coupled to a scrambled peptide (top panel), the FTNITYRGT peptide (middle panel), or the ANNPLYKEATS peptide (bottom panel) with fMLP-activated PMNs were analyzed for phosphorylation by HPLC (representative tracings of 2 experiments are shown). Arrows indicate position of unphosphorylated or phosphorylated standards.
