ZMBH J.Bericht 2000 - Zentrum für Molekulare Biologie der ...

switches between a GDP- and a GTP-bound form. Its
nucleotide-bound state is controlled by a number of
activities whose asymmetric nucleocytoplasmic distribution
is thought to result in a high nuclear RanGTP
concentration and very low RanGTP levels in the cytoplasm.
Importin-β like transport receptors are RanGTP
binding proteins that respond to this gradient by
loading and unloading their cargo in the appropriate
compartment. Importins bind their substrates in the
absence of RanGTP, i.e. in the cytoplasm and release
them upon encountering RanGTP in the nucleus. The
importins then return to the cytoplasm where RanGTP
is released (involving GTP hydrolysis), thereby allowing
the importin to bind and import the next cargo molecule.
Exportins respond to the RanGTP gradient in
exactly the opposite way; they bind their cargo preferentially
in the presence of RanGTP in the nucleus,
where they form a trimeric substrate-exportin-RanGTP
complex. The trimeric export complex is disassembled
after export to the cytoplasm and the „empty“ exportin
can re-enter the nucleus to participate in another round
of export.
Transport receptors bind their transport substrates in
many cases directly, an example being nuclear export
of tRNA by exportin-t (see below). In other cases, substrate
recognition is more complicated and involves an
adapter molecule. The best studied example for that is
the classical NLS import pathway, where the NLS is
recognised by the adapter importin α, which in turn
binds the actual transport receptor importin beta. This
complicates the corresponding transport cycle considerably
in that not only importin β, but also importin
α needs to be return to the cytoplasm. Interestingly,
importin α employs a specialised exportin, namely
CAS, for this purpose.
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Figure 1: Transport cycles of Importins (Imp) and exportins
(Exp). For details, please see main text.
Recycling of snurportin 1 back to the cytoplasm
F. Paraskeva, U. Kutay, in collaboration with R. Lührmann
(Univ. Marburg), F.R. Bischoff (DKFZ, Heidelberg),
E. Izaurralde (EMBL, Heidelberg)
Snurportin 1 functions as an import adapter for U
snRNPs. It recognises the m 3 G-cap import signal of
U snRNPs and also binds Importin β which in turn
is the actual mediator of import. Just as importin α,
also snurportin 1 needs to be returned to the cytoplasm
after each round of import in order to accomplish further
import cycles. We found that this recycling is
mediated by CRM1. CRM1 was originally identified
as the exportin specific for leucine-rich nuclear export
signals (NESs). However, the CRM1/ snurportin 1
interaction differs significantly from that with other
export substrates: Firstly, snurportin 1 binds CRM1
not through a short, leucine-rich signal, but instead
through a large domain that comprises more than 200
residues. Secondly, snurportin 1 binds CRM1 100
times more tightly than an NES from the HIV Rev
protein. In addition, snurportin 1 can bind either the
m 3 G-cap import signal or CRM1, but not both at the
same time. This property ensures that CRM1 exports
only those snurportin 1 molecules which have already
released their cargo and thereby allows snurportin 1 to
mediate productive import cycles.
tRNA export
U. Kutay, G. Liposwsky, P. Schwarzmaier, in collaboration
with E. Izaurralde (EMBL, Heidelberg) and
F.R. Bischoff (DKFZ, Heidelberg)
tRNAs are synthesised as precursor molecules (pretRNAs),
mature to functional tRNA and finally get
exported to the cytoplasm. There, they participate in
cycles of aminoacylation, binding to the elongation
factor eE1A and function in translation. We have identified
exportin-t as the tRNA-specific exportin and
showed that it functions according to the exportinparadigm
described above. The maturation of pretRNAs
to functional tRNAs involves trimming of the
3´and 5´ends, post-transcriptional addition of the 3´
CCA end to which the amino acid is later attached,
and the modification of a number of nucleosides. It
is quite remarkable that exportin-t preferentially binds
and exports mature tRNA which contain correctly processed
3´and 5´ends and the appropriate nucleoside
modifications. Exportin-t mediated export thus constitutes
a proof-reading or quality-control mechanism
that co-ordinates RNA processing with export and
thereby helps to ensure that only functional tRNA
arrives in the cytoplasm.
Nuclear import of ribosomal proteins and
histone H1
S. Jäckel
The biogenesis of ribosomes is a very complex process
that also involves nuclear import and export events:
Ribosomal proteins are first imported from the cytoplasm,
assemble with rRNA in the nucleolus to form
ribosomal subunits which are then finally re-exported
to the cytoplasm.
We have studied nuclear import of ribosomal proteins
in higher eukaryotes, focusing on rpL23a. We found
that at least 4 distinct transport receptors, namely
importin β, transportin, importin 5 and importin 7
can directly bind and import rpL23a. This not only
assigned the first functions to importin 5 and 7,
but showed an apparently quite common principle
in nuclear import, namely that some substrates can
„choose“ between several carriers. L23a binds through
the same domain or „import signal“ to each of the 4
transport receptors.
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