FIG. 7.

Cells expressing ΔTrkA contain elevated levels of Ras-GTP and activated ERK MAP kinases and Akt. (A) The level of Ras activation was measured by the Ras glutathione S-transferase–Ras binding domain (GST-RBD) pull-down assay. Bound (active) Ras (Ras-GTP) and total cell lysates of NIH 3T3 cells expressing vector, TrkA, ΔTrkA, and H-Ras61L were analyzed by Western blotting with anti-pan (non-isoform-specific) Ras antibodies. H-Ras61L migrates faster than wild-type endogenous Ras. (B) NIH 3T3 cells were analyzed for activation of ERK (left, top panel) and Akt (right, top panel) by Western blotting total cell lysates with antibodies that recognize the activated, phosphorylated forms of these kinases. Blots were also probed with anti-ERK (left, bottom panel) and anti-Akt (right, bottom panel) antibodies as controls.