G. Johnson and colleagues have analyzed the kinase cascade in which MEKK2, a MAP kinase kinase kinase, participates in mammalian cells. By a yeast two-hybrid screen (see Chapter 11), MEKK2 was found to bind MEK5, a MAP kinase kinase. To elucidate the signaling pathway transduced by MEKK2 in vivo, the following studies were performed in human embryonic kidney (HEK293) cells in culture.
a. HEK293 cells were transfected with a plasmid encoding recombinant, tagged MEKK2, along with a plasmid encoding MEK5 or a control vector that did not encode a protein (mock). Recombinant MEK5 was precipitated from the cell extract by absorption to a specific antibody. The immuno-precipitated material was then resolved by polyacrylamide gel electrophoresis, transferred to a membrane, and examined by Western blotting with an antibody that recognized tagged MEKK2. The results are shown in the figure below, part a. What information about the MEKK2 kinase cascade do we learn from this experiment? Do the data in part a of the figure prove that MEKK2 activates MEK5, or vice versa?
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