Chemistry Reference
In-Depth Information
with RanGTP in the nucleus and release both when they reach the cytoplasm. In both
cases, once in the cytoplasm, RanGTP is hydrolyzed to RanGDP to disassemble the
complex.
The messenger RNAs rely on the adaptor protein TAP for their transport in the
cytoplasm. The TAP-mediated export of the mRNAs appears to be unlinked from the
concomitant binding of RanGTP [48], and its marginal role in the process is due to
its involvement in nuclear import of TAP and other proteins rather than in the
mRNA transport itself. In that case the unidirectional movement seems to be
maintained by a highly conserved DEAD-box ATPase/RNA helicase essential for
mRNA export, Dbp5p [49]. This is a shuttling protein that associates with the RNA
early in transcription and translocates into the cytoplasm with the complex. On the
cytoplasmic side of the NPC multiple binding sites for Dbp5p anchor the helicase in
this region where Dbp5p is activated by the concomitant presence of Gle1 and
Inositol-P
6
[50, 51]. Remodeling of the mRNP causes the release from the NPC and
the recycling in the nucleus of the proteins involved in the transport, thus avoiding a
possible backward movement of the complex. Although we have considerable
information about different kinds of interactions and the mechanism of receptor-
mediated cargo transport, howmolecules actually translocate through the pore is still
unclear.
A key role is suggested for the FG repeats in the nucleoporins [47, 52, 53]. Since
these phenylalanine-rich domains are able to interact with each other and with the
transport receptors, several models have been developed to describe the possible
movements inside the NPC. A Brownian af
nity-gating model proposes the forma-
tion of an internal channel with binding sites at the tunnel entrance that facilitate the
access of the bound molecules but completely exclude those that are unbound [52].
Inside the channel, the particles move by Brownian motion. Macara [47] proposed
instead that the channel walls are actually covered with the FG repeats allowing the
molecules to jump from one repeat to another while inert molecules can diffuse in
the channel. Another possibility is the formation of a meshwork by interaction
among the FG domains creating a permeability barrier that restricts the passage for
inert molecules [53]. This selective phasemodel proposes that the nucleoporins form
this sieve-like structure within the pores, and transient interactions with the FG
repeats would allow the bound particle to dissolve into this structure. The carrier
would help the cargo to translocate by masking domains that enable them to interact
positively with the meshwork.
Fundamental insights into the translocation process require further investigation
and higher resolution structures of the intact NPC, a goal that can be achieved only
by single molecule approaches. These methods can provide unique information on
topographic properties and kinetic processes with excellent spatial and time
resolution. To this end, a single-molecule far-
eld
fluorescence microscopy
approach was applied to the NPC of permeabilized human cells [54], allowing the
measurement of dwell times of NTF2 and transportin with and without their
speci
c cargo molecules bound. These data highlight that binding at the NPC is not
the rate-limiting step and that particles can translocate simultaneously via multiple
parallel pathways.
