To investigate cell–matrix adhesion, dissociated single GFP+ CNC cells were cultured for 45 min on fibronectin, and the percentage of the adherent cells was quantified (Fig. 6N). Approximately 90% of the Dse-MO-injected CNC cells lost adherence to fibronectin compared with the uninjected cells, whereas Dse-5MM-MO had no significant effect. As previously reported (Alfandari et al., 2003), CNC cells failed to attach to bovine serum albumin (BSA)-coated substrates. The observed defects were not a result of apoptosis because fluorescence-activated cell sorting (FACS) indicated that 2 h after the extraction, the CNC cells from the Dse-MO-injected embryos had >95% cell viability, which was comparable to that of the uninjected cells (Fig. S7A-C′). Moreover, the Dse-morphant CNC cells exhibited a normal cell cycle progression (Fig. S7D-F). At more than 4 h after the extraction, the Dse-MO-injected CNC cells did not have a lower survival rate compared with the uninjected controls (data not shown). Thus, our data on CNC cells indicate that IdoA in CS/DS is essential for cell adhesion and migration on fibronectin.
