Differentiation and transforming growth factor-beta receptor down-regulation by collagen-alpha 2 beta 1 integrin interaction is mediated by focal adhesion kinase and its downstream signals in murine osteoblastic cells

Interaction of type I collagen (COL(I)) with alpha 2 beta 1 integrin causes differentiation and transforming growth factor (TGF)-beta receptor down-regulation in osteoblastic cells (Takeuchi, Y,, Nakayama, K,, and Matsumoto, T, (1996) J, Biol, Chem, 271, 3938-3644), The TGF-beta receptor down-regulation enables cells to escape from the inhibition of differentiation by TGF-beta. To clarify how the cell-matrix interaction regulates these phenotypic changes, signaling pathways were examined in murine MC3T3-E1 cells, Attachment of cells to COL(I) stimulated tyrosine phosphorylation of focal adhesion kinase (FAK) and extracellular signal-regulated kinase (ERK), a mitogen-activated protein kinase (MAPK), and enhanced MAPK activity, Inhibition of tyrosine kinase by herbimycin A, destruction of focal adhesion by cytochalasin D, or overexpression of antisense FAK mRNA prevented the activation of ERK/MAPK and the increase in alkaline phosphatase (ALP) activity. Transient expression of a MAPK-specific phosphatase, CL100, also sup pressed the elevation of ALP activity, In addition, introduction of a constitutively active MAPK kinase enhanced ALP activity in the absence of collagen production, TGF-beta receptor down-regulation was abrogated by treatments that inactivate FAK, whereas the expression of CL100 had no effect, These results demonstrate that COL(I)-alpha 2 beta 1 integrin interaction facilitates differentiation and down-regulates TGF-beta receptors via the activation of FAK and its diverse downstream signals, These signaling pathways may play an important role in the sequential differentiation of osteoblasts during bone formation.