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Recently, we demonstrated that the Dictyostelium MEK1 is transiently
SUMOylated in response to stimulation by the chemoattractant cAMP
(Sobko et al., 2002), suggesting that SUMOylation may play a role in
regulating MEK1.
We examined the subcellular localization of ERK1 and MEK1 by using
epitope tags. In resting cells, MEK1 is concentrated in the nucleus, while
ERK1 localizes in the cytoplasm (Figure 17.4A). Upon cAMP stimulation,
MEK1 and ERK1 localize to the plasma membrane and are found at the
leading edge of chemotaxing cells, with kinetics similar to those observed for
PH domain-containing proteins and PI3K (Figure 17.4C). The kinetics of
MEK1 translocation are indistinguishable from those of MEK1 SUMOyla-
tion of MEK1. Mutational analysis identified MEK1
K105
as the site of
SUMOylation. MEK1
K105R
, which carries a K-to-R substitution at the site of
SUMOylation, remains in the nucleus. Constitutively active MEK1
(MEK1
S444E,T448E
) is constitutively SUMOylated, constitutively cytosolic
and partially membrane-associated. In contrast, MEK1
S444A,T448A
is not
SUMOylated and remains nuclear after stimulation. These data suggest that
MEK1 translocation requires SUMOylation and that MEK1 activation may
be the direct signal that leads to MEK1 SUMOylation (Figure 17.4B). MEK1
is rapidly dephosphorylated and de-SUMOylated, whereupon it relocalizes to
the nucleus and associates with MIP1, a MEK1-interacting protein identified
via a yeast two-hybrid screen. MIP1 functions in nuclear sequestration of
MEK1 in unstimulated cells. MEK1 is also ubiquitinated. MIP1 has a RING
domain and possesses ubiquitin E3 ligase activity. Mutational analysis
indicated that MIP1 mediates MEK1 ubiquitination. It is possible that
MEK1 ubiquitination, which is highest in time points after MEK1 relocalizes
to the nucleus, is a further mechanism of pathway down-regulation and may
mediate MEK1 degradation, although this model has not been proven
(Figure 17.4D). SUMOylation consensus sites are found in MEKs from a
wide variety of organisms, suggesting that SUMOylation of MAP kinase
kinases could be conserved and represent an ancient mechanism controlling
subcellular localization of the cascade.
The initial asymmetric signal and downstream asymmetric
signals
PH domain translocation to the leading edge, an initial event to establish cell
polarity, is regulated by spatial and temporal activation of its upstream
kinase, PI3K, and the phosphatase PTEN. PI3K jumps to the leading edge,
whereas PTEN delocalizes from the edge after stimulation. What makes
PI3K/PTEN localization polarized? Interestingly, the PH domain and PI3Ks
