FIG. 1.

Role for the N terminus of flagellin in NLRC4 activation. (A) Diagram of all retroviral constructs used in transduction experiments. Note that all constructs expressed GFP, either as a direct fusion to L. pneumophila flagellin (FlaA; as shown) or downstream of an internal ribosome entry site (IRES) (not shown). C65, C-terminal 65 amino acids of FlaA; C20, C-terminal 20 amino acids of FlaA; N65, N-terminal 65 amino acids of FlaA. FlaAΔN constructs lack the indicated number of amino acids from the N terminus. (B and C) Wild-type (C57BL/6) or isogenic Naip5^−/− or Nlrc4^−/− macrophages were transduced with the indicated constructs, and the percentage of GFP⁺ cells was enumerated by flow cytometry 3 to 4 days after transduction. (D and E) Wild-type, Naip5-deficient, or Naip5^−/− Nlrc4^−/− doubly deficient macrophages were transduced with the indicated constructs, and the percentage of GFP⁺ cells was enumerated by flow cytometry 3 to 4 days after transduction. More than 10,000 cells were analyzed for each experiment. For each panel, data shown are from a single representative experiment of at least three that produced similar results.