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As seen in Table 1, expression of ADAM12 did not change either the total or the surface levels of the α4 integrin subunit. In contrast, lamellipodia protrusions and cell body translocation seemed less robust in ADAM12 transfected samples (Figure 4B, right, and Movie 1B). Confluent cell monolayers were then scratch wounded and observed by videomicroscopy, as described in the legend to Movie 1. After 24 h they were analyzed in transwell migration assays as described in the legend to Figure 2, except that the transwell inserts were coated with 2 μg/ml CS-1. Moreover, this pattern correlates with the selective adhesive activity of the ADAM12 disintegrin domain (Figure 1).
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Molt 3 cells, obtained from American Type Culture Collection (Manassas, VA), were maintained in RPMI 1640 medium containing 10% FBS, l-glutamine, antibiotics, sodium pyruvate, and 15 mM HEPES buffer. CHOB2-α5 and CHOK1 cells (provided by A. Rick Horwitz, University of Virginia) were maintained in DMEM containing 10% fetal bovine serum (FBS) (Invitrogen), l-glutamine, nonessential amino acids, and antibiotics. The α5β1 integrin engages the central cell binding domain (CCBD) of FN and forms focal adhesions, which function both as anchors and as signaling centers. In particular, the fibronectin (FN) binding integrins α4β1 and α5β1 are well characterized for their roles in cell motility.
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