Mg. Seidel et al., Activation of mitogen-activated protein kinase by the A(2A)-adenosine receptor via a rap1-dependent and via a p21(ras)-dependent pathway, J BIOL CHEM, 274(36), 1999, pp. 25833-25841
The A(2A)-adenosine receptor, a prototypical G(s)-coupled receptor, activat
es mitogen-activated protein (MAP) kinase in a manner independent of cAMP i
n primary human endothelial cells. In order to delineate signaling pathways
that link the receptor to the regulation of MAP kinase, the human A(2A) re
ceptor was heterologously expressed in Chinese hamster ovary (CHO) and HEK2
93 cells. In both cell lines, A(2A) agonist-mediated cAMP accumulation was
accompanied by activation of the small G protein rap1, However, rap1 mediat
es A(2A) receptor-dependent activation of MAP kinase only in CHO cells, the
signaling cascade being composed of G(s), adenylyl cyclase, rap1, and the
p68 isoform of B-raf, This isoform was absent in HEK293 cells. Contrary to
CHO cells, in HEK293 cells activation of MAP kinase by A(2A) agonists was n
ot mimicked by 8-bromo-cAMP, was independent of G alpha(s), and was associa
ted with activation of p21(ras), Accordingly, overexpression of the inactiv
e S17N mutant of p21(ras) and of a dominant negative version of mSos (the e
xchange factor of p21(ras)) blocked MAP kinase stimulation by the A(2A) rec
eptor in HEK 293 but not in CHO cells. In spite of the close homology betwe
en p21(ras) and rap1, the S17N\ mutant of rap1 was not dominant negative be
cause (i) overexpression of rap1(S17N) failed to inhibit A(2A) receptor-dep
endent MAP kinase activation, (ii) rap1(S17N) was recovered in the active f
orm with a GST fusion protein comprising the rap1-binding domain of ralGDS
after A(2A) receptor activation, and (iii) A(2A) agonists promoted the asso
ciation of rap1(S17N) with the 68-kDa isoform of B-raf in CHO cells. We con
clude that the A(2A) receptor has the capacity two activate MAP kinase via
at least two signaling pathways, which depend on two distinct small G prote
ins, namely p21(ras) and rap1, Our observations also show that the S17N ver
sion of rap1 cannot be assumed a priori to act as a dominant negative inter
fering mutant.