398
Cell Nucleus Biogenesis, Structure and Function
it is imported through the nuclear pores.
Assembly and maturation of the lamina
continues for an hour or two into the G1
phase. The fluid properties of the nuclear
membrane eventually lead to formation
of a complete double envelope. While
t
h
i
sp
r
o
c
e
s
si
so
n
g
o
i
n
g
,n
u
c
l
e
a
rp
o
r
e
s
are assembled into the envelope. Soon
after cell division, a fully operative nuclear
envelope is formed.
3.4
Nuclear Transport and Karyopherins
The fact that eukaryotes have evolved
to
conFne
the
genetic
material
within
a
specialized
organelle – the
nucleus – demands
that
mechanisms
must exist to regulate the passage of
information between the nucleus and
cytoplasm. This is self-evident as the
products of gene transcription must pass
to
the cytoplasm
for
translation
and
all the proteins that control and direct
gene function must pass in the opposite
direction, moving into the nucleus from
the cytoplasm where they are synthesized.
This nuclear-cytoplasmic trafFc takes
place through the nuclear pores. The de-
tailed structure of these pores has been
elucidated and, as we saw earlier, provides
a structural framework that can be used
to elaborate the mechanisms of nuclear
pore-mediated transport. The basic princi-
ples of transport are well known, though
some details remain to be conFrmed. The
Frst point to emphasize is that nuclear
pores do not provide an absolute barrier be-
tween the nucleus and cytoplasm. In fact,
small molecules and proteins that have
as
izebe
low
50 kDa are able to diffuse
through the pores. Any proteins that are
larger than this, and protein nucleic acid
complexes undergo either nuclear import
or export using speciFc transport mecha-
nisms. This active process involves nuclear
localization and export signals that are rec-
ognized by the import/export machinery.
The nuclear localization sequence of the
SV40 virus large T antigen was the Frst to
be analyzed in detail. In this protein, the
amino acid sequence: Pro-Pro-Lys-Lys-Lys-
Arg-Lys-Val directs nuclear accumulation
of the protein. In other proteins, it is
not uncommon to Fnd bipartite localiza-
tion signals where two short lysine and
arginine-rich regions are separated by
10
intervening amino acids.
The molecular details of the protein
transport process are by far the best
understood. The key components in this
process are the karyopherin proteins that
operate as shuttling transport receptors.
The
simple
eukaryotes
S. cerevisiae
is
known to have 14 karyopherin proteins
and at least 20 have been described in
human cells. Karyopherins can be further
subdivided into importins and exportins.
As the names imply, these are largely
dedicated to either the nuclear import
or export pathways; though examples are
known that can transport different cargoes
across the nuclear pores in both directions.
3.4.1
The RanGTP/GDP Cycle
Movement of a molecule across the nuclear
pore will generally take place against a con-
centration gradient and involves ATP hy-
drolysis. The basis of this process relies on
the way in which the karyopherin–cargo
complexes interact with the pore proteins
and subsequently pass through the pore to
be released on the other side. The behav-
ior of importins and exportins during this
process is regulated by the small GTPase
Ran. More speciFcally, substrate binding
and release by the karyopherins is regu-
lated by the asymmetrical distribution of
the GTP- and GDP-bound states of Ran.
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