Regulation of cell death and neurite outgrowth by overexpressed focal adhesion kinase in rat pheochrome cytoma PC 12

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We established cells overexpressed focal adhesion kinase (FAK) or its mutated genes named Y397F, K454R or Y925F. After cells were differentiated with nerve growth factor (NGF), we estimated the sensitivity of each cell against cell death induced by oxidative stress or ER stress. We also determined the sensitivity of cells overexpressed antiapoptotic protein Bcl-2 or Bcl-xL.(1) In the cells overexpressed Bcl-2 or Bcl-xL, cell death was clearly suppressed induced by oxidative stress (rotenone, dopamine, H2O2) or ER stress (brefeldin A, thapsigargin). Activation of caspase-2 or 3 induced by rotenone was also inhibited.(2) Overexpression of FAK tended to resist against cell death by oxidative stress.(3) In the cells overexpressed Y397F mutation which is autophosphorylation site, cell death induced by oxidative stress was increased compared with vector cells. However, overexpression of K454R at catalytic site induced suppression of cell death. Moreover, rotenone-induced caspase-2 or -3 activity was also enhanced in Y397F cells, but clearly inhibited in K454R cells. In the cells overexpressed Y925F at C-terminus, sensitivity of cell death was not altered.(4) In the cells overexpressed Y397F, expression of Akt which is a part of survival signal, was decreased than vector cells. Antiapoptotic protein Bcl-xL expression was not altered. In the K454R cells, expression of both proteins were not altered. The above mutations, Y397F and K454R, are able to suppress FAK dependent signalling pathway, however, there is a difference against the sensitivity of cell death. It is necessary to clarify the varies of signalling after FAK activation.(5) In the cells overexpressed any genes, NGF-induced neurite outgrowth was similar to vector cells.