Fig. 8. Refinement of the cilia trafficking domain of PC2. (A) Deletion of amino acids 31-72 from PC2-L703X does not alter trafficking to cilia, whereas (B) deletion of amino acids 5-31 results in complete absence of (5-31)PC2-L703X from cilia. Replacement of the cytosolic N-terminus of hTFR with either (C) the first 31 amino acids or (D) the first 15 amino acids of PC2 is sufficient to traffic the chimeric reporter molecule to cilia. (E) Non-permeabilized living cells have epitopes available to anti-HA on cilia and (E^*) at the level of the apical plasma membrane. Anti-HA was applied on both the apical and basolateral aspects but there was no detectable immunoreactivity on the basolateral surfaces, indicating that the chimeric protein only trafficked to cilia and the apical membrane. Anti-acetylated -tubulin gave no signal in non-permeabilized, living cells (not shown) confirming that the observed signal is indeed extracellular. Anti-HA, green; anti-acetylated -tubulin, red. Bars, 10 µm.