Ms. Schwab et C. Dreyer, PROTEIN-PHOSPHORYLATION SITES REGULATE THE FUNCTION OF THE BIPARTITE NLS OF NUCLEOLIN, European journal of cell biology, 73(4), 1997, pp. 287-297
Nucleolin is a major component of the nucleolus, In Xenopus laevis, a
maternal store of nucleolin is accumulated in the multiple nucleoli ge
nerated during oogenesis, This maternal nucleolin is distributed throu
ghout the cytoplasm of the egg during oocyte maturation and after fert
ilization it is gradually reaccumulated in the nuclei of the embryo, C
ytoplasmic localization of nucleolin coincides with massive phosphoryl
ation by p34(cdc2) kinase, and nuclear translocation is accompanied by
net dephosphorylation. Multiple phosphorylation consensus sites for t
he cell cycle-dependent p34(cdc2) kinase and for protein kinase CK2 ar
e clustered in the N-terminal domain of nucleolin, To assess the effic
iency of the bipartite nuclear localization signal, we have constructe
d fusion proteins consisting of maltose binding protein (MBP) and the
nuclear localization signal of nucleolin, In addition, either an acidi
c domain of nucleolin without phosphorylation sites, or an acidic doma
in containing 4 CK2 sites, or a cluster of 5 cdc2 sites was fused to t
he MBP-nuclear localization signal (MBP-NLS), Nuclear translocation of
these constructs was tested in an in vitro system consisting of Xenop
us egg extract and sperm nuclei, Nuclear targetting of MBP by the bipa
rtite nuclear localization signal of nucleolin became significantly mo
re efficient after addition of either CK2 sites or cdc2 sites to the M
BP-NLS construct, Yet the cdc2 sites play a dual role, They enhance nu
clear translocation exclusively in their dephosphorylated state and pr
omote cytoplasmic localization when phosphorylated, thereby providing
a powerful cell cycle-dependent regulatory element of the nuclear loca
lization signal.