The use of MBP as a preferred substrate suggested that the
p48 SIP kinase might be a MAP kinase. Another hallmark of
MAP kinases is their activation via dual phosphorylation of
tyrosine and threonine residues by MAP kinase kinases
(Seger and Krebs, 1995). To determine whether the SIP kinase
was similarly activated, extracts from SA-treated cells
were subjected to immunoblot analysis, using the phosphotyrosine-
specific monoclonal antibody 4G1 O (Figure 3A).
lncreases in the amount of a phosphotyrosine-containing
48-kD polypeptide were associated with the SA-mediated
activation of the p48 SIP kinase. Furthermore, the transient
activation of the p48 kinase after SA treatment (Figure IA)
correlated with a transient increase in the level of a phosphotyrosine-
containing 48-kD protein (Figure 3A). These results
suggest that the p48 SIP kinase is a MAP kinase.
