Fig. 2. Temperature dependence of fusion by the F protein of SV5 strain W3A co-expressed with either SV5 HN (open squares) or influenza virus HA (closed squares). R18/CF double-labeled RBCs were bound at 4°C to CV-1 cells co-expressing the F proteins and either HN or HA. The samples were subsequently incubated at the reported temperatures for 5 min. Dye transfer was assayed by confocal microscopy. Fusion is expressed as contents mixing events per microscopic field, averaged over 4–6 fields. Lipid mixing (R18 transfer) and contents mixing (CF transfer, reported here) were coincident. (A) W3A F-mediated fusion is promoted by HN co-expression. (B) Fusion mediated by cleaved W3A strain FR3 (the cleavage site mutant) is also promoted by HN co-expression. Expression of HN, uncleaved HA or cleaved HA alone did not cause detectable fusion, and F+HN and F+HA fusion were inhibited by addition of the F-specific antibody F1a.