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Chapter 5
Nuclear structure and transport
By
Charles N. Cole & Pamela A. Silver
5.1 Introduction
• The nucleus contains most of the cell’s DNA, allowing for
sophisticated regulation of gene expression.
• The nuclear envelope is a double membrane that
surrounds the nucleus.
5.1 Introduction
• The nucleus contains subcompartments that
are not membrane-bounded.
• The nuclear envelope contains pores used
for:
– importing proteins into the nucleus
– exporting RNAs and proteins from the nucleus
5.2 Nuclei vary in appearance according to
cell type and organism
• Nuclei range in size from about one micron (1 μm) to
more than 10 μm in diameter.
• Most cells have a single nucleus, but some cells
contain multiple nuclei, and a few cell types lack
nuclei.
• The percentage of the genome that is
heterochromatin varies among cells and increases as
cells become more differentiated.
5.3 Chromosomes occupy distinct territories
• Although the nucleus lacks internal
membranes, nuclei are highly organized and
contain many subcompartments.
• Each chromosome occupies a distinct region
or territory.
– This prevents chromosomes from becoming
entangled with one another.
5.3 Chromosomes occupy distinct territories
• The nucleus contains both chromosome
domains and interchromosomal regions.
5.4 The nucleus contains subcompartments
that are not membrane-bounded
• Nuclear subcompartments are not membranebounded.
• rRNA is synthesized and ribosomal subunits are
assembled in the nucleolus.
• The nucleolus contains DNA that encodes rRNAs
and that is present on multiple chromosomes.
5.4 The nucleus contains subcompartments that are not membrane-bounded
• mRNA splicing factors:
– are stored in nuclear speckles
– move to sites of transcription where they function
• Other nuclear bodies can be identified with
antibodies, but the functions of most of these
are unknown.
5.5 Some processes occur at distinct
nuclear sites and may reflect an underlying
structure
• The nucleus contains replication sites where
DNA is synthesized.
• The nucleus may contain a nucleoskeleton
that could help to organize nuclear functions.
5.6 The nucleus is bounded by the nuclear
envelope
• The nucleus is surrounded by a nuclear envelope
consisting of two complete membranes.
• The outer nuclear membrane is continuous with the
membranes of the endoplasmic reticulum (ER).
• The lumen of the nuclear envelope is continuous
with the lumen of the ER.
• The nuclear envelope contains numerous NPCs.
– They are the only channels for transport of molecules and
macromolecules between the nucleus and the cytoplasm.
5.7 The nuclear lamina underlies the
nuclear envelope
• The nuclear lamina is constructed of intermediate
filament proteins called lamins.
• The nuclear lamina is located beneath the inner
nuclear membrane.
– They are physically connected by lamina-associated integral
membrane proteins.
• The nuclear lamina plays a role in nuclear envelope
assembly and may provide physical support for the
nuclear envelope.
5.7 The nuclear lamina underlies the nuclear envelope
• Proteins connect the nuclear lamina to chromatin;
– this may allow the nuclear lamina to organize DNA
replication and transcription.
• Yeast and some other unicellular eukaryotes lack a
nuclear lamina.
5.8 Large molecules are actively
transported between the nucleus and
cytoplasm
• Uncharged molecules smaller than 100 daltons
can pass through the membranes of the
nuclear envelope.
• Molecules and macromolecules larger than
100 daltons cross the nuclear envelope by
moving through NPCs.
5.8 Large molecules are actively transported between the nucleus and cytoplasm
• Particles up to 9 nm in diameter (corresponding to
globular proteins up to 40 kDa) can pass through
NPCs by passive diffusion.
• Larger macromolecules are actively transported
through NPCs and must contain specific information
in order to be transported.
5.9 Nuclear pore complexes are
symmetrical channels
• NPCs are symmetrical structures that are
found at sites where the inner and outer
nuclear membrane are fused.
• Each NPC in human cells has a mass of
~120 106 daltons (40 times that of a
ribosome).
• It is constructed from multiple copies of ~30
proteins.
5.9 Nuclear pore complexes are symmetrical channels
• NPCs contain:
– fibrils that extend into the cytoplasm
– a basket-like structure that extends into the
nucleus
5.10 Nuclear pore complexes are
constructed from nucleoporins
• The proteins of NPCs are called
nucleoporins.
• Many nucleoporins contain repeats of short
sequences, which are thought to interact with
transport factors during transport.
– Such as:
• Gly-Leu-Phe-Gly
• X-Phe-X-Phe-Gly
• X-X-Phe-Gly
5.10 Nuclear pore complexes are constructed from nucleoporins
• Some nucleoporins are transmembrane
proteins that are thought to anchor NPCs in
the nuclear envelope.
• All of the nucleoporins of yeast NPCs have
been identified.
• NPCs are disassembled and reassembled
during mitosis.
• Some nucleoporins are dynamic: they rapidly
associate with and dissociate from NPCs.
5.11 Proteins are selectively transported
into the nucleus through nuclear pores
• Mature nuclear proteins contain sequence information required
for their nuclear localization.
• Proteins selectively enter and exit the nucleus through nuclear
pores.
• Information for nuclear import lies in a small portion of the
transported protein.
5.12 Nuclear localization sequences target
proteins to the nucleus
• A nuclear localization sequence (NLS) is often a
short stretch of basic amino acids.
• NLSs are defined as both necessary and
sufficient for nuclear import.
5.13 Cytoplasmic NLS receptors mediate
nuclear protein import
• Receptors for nuclear import are cytoplasmic
proteins that bind to the NLS of cargo proteins.
• Nuclear import receptors are part of a large family
of proteins often called karyopherins.
5.14 Export of proteins from the nucleus is
also receptor-mediated
• Short stretches of amino acids rich in leucine
act as the most common nuclear export
sequences.
• A nuclear export receptor:
– binds proteins that contain nuclear export
sequences (NESs) in the nucleus
– transports them to the cytoplasm
5.15 The Ran GTPase controls the direction
of nuclear transport
• Ran is a small GTPase that is common to all
eukaryotes and is found in both the nucleus
and the cytoplasm.
• The Ran-GAP promotes hydrolysis of GTP by
Ran.
• The Ran-GEF promotes exchange of GDP for
GTP on Ran.
5.15 The Ran GTPase controls the direction of nuclear transport
• The Ran-GAP is cytoplasmic, whereas the RanGEF is located in the nucleus.
• Ran controls nuclear transport by binding
karyopherins and affecting their ability to bind
their cargoes.
5.16 Multiple models have been proposed
for the mechanism of nuclear transport
• Interactions between karyopherins and nucleoporins are
critical for translocation across the nuclear pore.
• Directionality may be conferred in part by distinct
interactions of karyopherins with certain nucleoporins.
5.17 Nuclear transport can be regulated
• Both protein import and export are regulated.
• Cells use nuclear transport to regulate many
functions, including:
– transit through the cell cycle
– response to external stimuli
• The movement of the transcription factor NF-κB
illustrates how nuclear transport is regulated.
5.18 Multiple classes of RNA are exported
from the nucleus
• mRNAs, tRNAs, and ribosomal subunits
produced in the nucleus are exported through
NPCs to function during translation in the
cytoplasm.
5.18 Multiple classes of RNA are exported from the nucleus
• The same NPCs used for protein transport
are also used for RNA export.
• Export of RNA is receptor-mediated and
energy-dependent.
• Different soluble transport factors are
required for transport of each class of RNA.
5.19 Ribosomal subunits are assembled in
the nucleolus and exported by exportin 1
• Ribosomal subunits are assembled in the
nucleolus where rRNA is made.
• Ribosomal proteins are imported from the
cytoplasm for assembly into the ribosomal
subunits.
• Export of the ribosomal subunits is carriermediated and requires Ran.
5.20 tRNAs are exported by a dedicated
exportin
• Exportin-t is the transport receptor for tRNAs.
• tRNA export requires Ran.
• tRNA export may be affected by modifications of
the tRNAs.
• tRNAs may be re-imported into the nucleus.
5.21 Messenger RNAs are exported from
the nucleus as RNA-protein complexes
• Proteins that associate with mRNAs during
transcription help to define sites of pre-mRNA
processing.
• They are also thought to package mRNAs for
export.
5.21 Messenger RNAs are exported from the nucleus as RNA-protein complexes
• Most proteins that associate with mRNA in
the nucleus are removed after export and
returned to the nucleus.
– A few are removed immediately prior to export.
• Signals for mRNA export may be present in
proteins bound to the mRNA.
• The export of mRNA can be regulated, but
the mechanism for this is unknown.
5.22 hnRNPs move from sites of processing
to NPCs
• mRNAs are released from chromosome
territories into interchromosomal domains
following completion of pre-mRNA
processing.
• mRNAs move to the nuclear periphery by
diffusion through interchromosomal spaces.
5.23 mRNA export requires several novel
factors
• Many factors required uniquely for mRNA export
have been identified.
• Factors able to bind to both the mRNP and nuclear
pore complex help to mediate mRNA export.
5.23 mRNA export requires several novel factors
• One factor, Dbp5, is an ATPase and may use
energy from ATP hydrolysis to remove mRNP
proteins during transport.
5.24 U snRNAs are exported, modified,
assembled into complexes, and imported
• U snRNAs produced in the nucleus are
–
–
–
–
exported
modified
packaged into U snRNP RNA-protein complexes
imported into the nucleus to function in RNA
processing
5.25 Precursors to microRNAs are exported
from the nucleus and processed in the
cytoplasm
• MicroRNAs are produced by:
–
–
–
–
transcription in the nucleus
partial processing to generate a hairpin precursor
export of the precursor by exportin-V
final processing in the cytoplasm