The nuclear envelope
The nuclear envelope consists of two concentric membranes perforated by pore complexes that maintain different protein compositions even though they are continuous.
* Inner nuclear membrane - contains specific proteins that act as binding sites for chromatin and the nuclear lamina.
* Outer nuclear membrane - continuous with the rough ER membrane that has attached ribosomes.
Nuclear pore complexes:
* Composed of more than 50 different proteins called nucleoporins.
* The number increases as transcription activity increases (typical mammalian cell: 3000-4000 pore complexes).
* Each pore complex contains one or more aqueous channels through which small water-soluble molecules can passively diffuse (experimental data is consistent with a 9 nm in diameter and 15 nm long channel, which accounts for only a small fraction of the pore complex volume). Opening can dilate up to 26 nm in diameter during the transport process.
* Because the aqueous pore can be fairly large, proteins can be transported in a fully folded conformation (by contrast to transport in most organelles), however very large particles seem to “squeeze” through the pore indicating that some parts of them must undergo restructuring during transport.
* Freely permeable to molecules under 5000 daltons, almost impermeable to molecules larger than 60.000 daltons.
* Conducts traffic in both directions. It is not known how collisions and congestion are avoided.
Nuclear localization signals:
* They can be either signal sequences or signal patches (in many proteins one or two sequences consisting of positively charged amino acids lysine and arginine, the precise sequence varying for different proteins).
Nuclear import receptors:
* They are encoded by a family of related genes.
* They are soluble cytosolic proteins that bind to nuclear localization signals and some nucleoporins, some of which are the tentacle like fibrils that extend into the cytosol from the rim of the nuclear pore complex.
* Some nucleoporins (including fibrils) contain a large amount of amino acid repeats that contain phenylalanine and glycine (FG-repeats) and serve as binding sites for the import receptors. They are thought to line the path taken by the import receptors and their bound cargo proteins, which move along this path by successive binding, dissociating and rebinding.
Nuclear export receptors:
* Export from the nucleus works exactly like import, but in reverse. Nuclear export receptors are encoded by the same gene family (karyopherins = nuclear transport receptors) as nuclear import receptors.
* Macromolecules to be exported contain certain nuclear export signals. Nuclear export receptors bind both these export signals and nucleoporins to guide their transport into the cytosol.
The Ran GTPase drives directional transport:
* The monomeric GTPase Ran, found both in the cytoplasm and the nucleus,provides energy for concentrating specific substances inside or outside the nucleus.
* Like all GTPases, Ran functions like a molecular switch, existing in two conformations – one that has a bound GTP and one that has a bound GDP. Two Ran-specific regulatory proteins, the cytosolic GTPase-activating protein(GAP) that triggers GTP hydrolysis and the nuclear guanine exchange factor(GEF) that exchanges GDP for GTP.
* Ran-GTP promotes dissociation of imported proteins from the import receptors and binding of the export receptor to it’s cargo protein. Conversion of Ran-GTP into Ran-GDP is achieved when the protein reaches the cytosol by two enzymes: Ran Binding Protein(which displaces Ran-GTP from it’s receptor) and GAP.
Other methods of regulating transport:
* Some proteins (like those that bind newly made mRNA) contain both nuclear localization and nuclear export proteins. The steady-state of these shuttling proteins is determined by the relative rates of their import and export.
* Some shuttling proteins move continuously through the nuclear pores. In other cases, the transport is stringently controlled. One mechanism involves turning nuclear localization and export signals on and off by phosphorylating adjacent amino acids.
* Another mechanism involves binding a protein(like a gene regulatory protein) to inhibitory cytosolic proteins that either anchor them in the cytoplasm or mask their nuclear localization signals. Upon receiving a stimulus, the protein is released from it’s mask or anchor and is transported into the nucleus.
* Similarly, export of mRNA can only be done if some proteins containing nuclear export signals are added as transcription and splicing proceed. After arriving in the cytosol, the proteins coating the RNA are stripped off and rapidly returned to the nucleus.
Contributed by: Andrew.
Posted on June 21, 2006
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