Nucleus Flashcards
why do we need a nucleus?
genome organization, genome integrity, control of gene regulation, coordination of genome activities, make faithful copies of itself and genome
Major structures of nucleus?
nuclear envelope (outer and inner), heterochromatin/euchromatin, nuclear pore, nucleolus, contiguous with rough ER
T/F. Inner nuclear membrane is continuous with the ER
False, Outer membrane
What components of the nucleus interact with chromatin?
inner nuclear membrane and lamina
T/F. Nuclear envelope has two lipid bilayers, it is contigueous with the ER, it is connected with the cytoskeleton
All true
T/F. INM and ONM are contiguous and have the same resident proteins
False, they are contiguous but have very different protein compositions
Proteins larger than what size need to be actively transported?
40kDa
Features of FG repeat Nups
Line length of nuclear pore complex, believed to form intermolecular interactions, creating a semi-permeable mesh that restricts translocation.
Binding sites for transport receptors.
Sole determinant of directionality of nuclear transport
Ran-GTP gradient, Ran-GDP should be higher in cytoplasm and Ran-GTP in the nucleus
NLS
nuclear localization signal, Lys-Arg rich, binding site for importins. Not always sufficient for import
How do importin and imported cargo protein dissociate?
Through binding of Ran-GTP to importin in the nucleus. Ran-GDP then produced through dephosphorylation activity of Ran GAP, allowing for release of Ran-GDP and restarting of the cycle
How do you reinstate Ran-GTP from Ran-GDP?
Through action of Ran GEF in nucleus
Ran-GTP gradient cycle
Be able to draw with import/export
Example of disease caused by mutations in an NPC
ALADIN mutants, cause Triple A syndrome, autosomal recessive disorder which leads to all kinds of physical and nuero issues
T/F. Ratio of Ran-GTP : Ran-GDP in a normal cell is higher in the cytoplasm
False, it is higher in the nucleus
T/F. Ratio of Ran-GTP : Ran-GDP in a normal cell is dependent on RAN-GAP and RAN-GEF
True
Strategy to detect lamina or INM interactions with chromatin
DamID, where you exogenously label GATC in DNA stretches close to where the fused Dam/protein of interest is bound
Examples of different components throughout the nucleoplasm that contribute to gene regulation
Cajal bodies, nucleoli, nuclear speckles
Two types of lamins
Lamin A type - found in differentiated cells, Lamin B type found in virtually all cells
Lamin structure
Intermediate filaments that form coiled coils and have disordered region at C-terminus
Lamin A processing
Farnesylation, carboxymethyltransferase, etc, and dysregulation in this leads to disease - ex progeria
T/F. Nuclear lamins are inner nuclear membrane proteins, intermediate filament proteins, and very stable
False, the first statement is false – they are associated with the inner nuclear membrane but are not a part of it.
Proximity tagging methods
DamID for DNA, APEX-ID or BIO-ID to biotinylate/otherwise label adjacent proteins or DNA. Can do Mass spec
Nuclear envelope breakdown controlled by
Cyclin B1 - CDK1 complex, phosphorylation events
what is the largest structure in the nucleus
chromosomes
largest sub-nuclear structure
nucleolus
nucleolus assembly
at loci containing rRNA gene arrays - on short arms of 5 different somatic chromosomes
nucleoli purpose
factories to assemble rRNAs into ribosomes. Precursor RNA is processed, co-assembled with ribosomal proteins. Then small and large subunits are exported to cytoplasm. Assembly of other RNA-protein complexes also happen here
additional subnuclear bodies (6) and their putative purposes
speckles (recycle mRNA splicing machinery?), cajal bodies (snRNP assembly?), gems (snRNP assembly), PcG bodies (polycomb group proteins), cleavage bodies (pre-mRNA cleavage), and PML bodies (transcriptional regulation)
components of the nuclear envelope (5)
inner membrane, outer membrane, lumen, NPCs, peripheral lamina structure
T/F. Nuclear lamina has different characteristics depending on cell type
True, for example liver cell nucleus has unusually thick lamina
T/F. Lamin filaments are typically unstable
False, they are very stable
Name three nuclear activities that lamins support, either directly or indirectly
- Scaffolds for proteins involved in activating or inactivating chromatin
- Mechanical, adaptive protection of the genome
- Regulate signaling in specific tissues
Other answers:: import/export via NPCs, support replication and DNA repair, mechanically anchor cytoskeleton, rebuild nucleus post division
What are the components and functions of LINC complexes?
SUNs + Nesprins, mechanically link nucleoskeleton and cytoskeleton
Examples of laminopathies
Emery-dreifuss muscular dystrophy, progeria, lipodystrophy
what are some essential factors that NPCs mediate import of from cytoplasm?
signaling, regulatory, ribosomal and nucleolar proteins. histones, transcription factors
what are some essential factors that NPCs mediate EXPORT of from cytoplasm?
ribosome subunits, mRNA complexes, tRNA, snRNA
What two amino acids create the hydrophobic barrier within nuclear pores (nups)?
Phe-Gly, AKA FG repeats
How does the nuclear envelope structure help ensure chromosome segregation fidelity?
Through nuclear disassembly and reformation, mediated by phosphorylation and subsequent desphosphorylation of lamins
What protein is largely responsible for unifying chromosomes within a single nucleus?
BANF1 and its nuclear lamina partners
what are the 4 distinct types of nuclear filaments?
A, C, B1, and B2
What are some other nucleoskeletal proteins?
Titin, actin, myosins, spectrins
Three types of NLS signals
Classic PKKKRKV (60%), bipartite, and proline-tyrosine rich
What type of signal does nuclear export require?
A nuclear export signal (NES) on cargo protein.
What are the primary differences between NLS and NES?
NES are hydrophobic, leucine rich, and have no strict consensus, contrary to NLS
T/F. Protein unfolding required for nucleocytoplasmic transport
False, not required
Requirement for NLS/NES use in import/export
Signal must be accessible on cargo surface - layer of regulation in mods or partners that hide (or expose) the signal
Ran-GTP gradient responsible for (2)
importins taking cargo in, and exportins taking cargo out
What does Ran need in order to bind and hydrolyze GTP
Binds GTP easily, but needs GAP in order to hydrolyze
What does Ran need in order to kick out GDP?
GEF (GDP exchange factor)
T/F. Ran-GEF is chromatin associated
True
