Periodic Activation of ERK2 and Partial Involvement of G Protein in ERK2 Activation by cAMP in Dictyostelium Cells

Intercellular signaling mediated by cAMP plays a pivotal role in coordinating cell movement into aggregates at the early stage of Dictyostelium development when the extracellular level of cAMP periodically changes at 6- to 7-min intervals. We have shown that MAP kinase ERK2 is activated via the cAMP receptor CAR1 in phase with this periodic change. This was revealed by assessing the level of ERK2 activation with immunoblots using two kinds of antibodies, commercially available anti-phospho-p44/p42 MAP-kinase antibody and anti-Dictyostelium ERK2 antibody. In this chapter, we describe the methods we have used to assess the level of activated ERK2 and partial involvement of G protein in cAMP-induced ERK2 activation.