Collagen I induced rapid and sustained activation of Erk1/2 signaling pathway. MKs were kept in suspension in BSA-coated dishes (A-C lane 1), or plated on fibrillar collagen I (A lanes 2-6; B lanes 2-4; C lanes 5-7) or PLL substrates (C lanes 2-4) for the indicated times. MKs pretreated by PD98059 or UO126 (B lanes 3,4; 20 μM for 30 minutes) were seeded on collagen I–coated dishes for 2 hours. Cells were lysed as described in “Materials and methods.” Proteins were analyzed by Western blot analysis using mAbs against phosphorylated Erk or total Erk (A-C top and bottom panels, respectively). The graphs illustrate levels of phosphorylated Erk relative to the levels of total Erk present in each sample.