Flashcards in Dr Shackleton's Work Deck (40):
Why is the DNA contained in the nucleus?
Provides control of gene expression
Regulation of timing of DNA replication and mitosis
Organization of huge and complex eukaryotic genomes
What are the main features of the nucleus?
What does peripheral heterochromatin do?
Act as a silencing environment
What is the nucleolus responsible for?
Transcription and processing of non-coding RNAs
Ribosome subunit assembly
DNA loops rich in rRNA genes forma structura component
Outline the basic NPC structure
Nuclear basket - regulate what passesthrough
8 fold symmetry
Multiple copies of 30 different nucleoporins
Aqueous pore in centre
How do small molecules (less than 40kDa)get in/out of the nucleus?
Diffuse through the NPC
How do large proteins get into the nucleus?
1. Cargo binds to importin in the cytoplasm
2. Goes through NPC
3. In nucleus Ran GTP binds displacing cargo
4. Goes through NPC
5. GAP activates Rans GTPase function causing Ran GDP to dissociate
What does the classical NLS constitute?
1/2 short stretches of basic amino acids (Arg, Lys, His) which binds to importin alpha
What is the most well known NES?
Leucine rich which binds to CRM1
Outline the process of nuclear export for a protein greater than 40kDa
1. Cargo and Ran GTP binds to the exportin in the nucleus
2. Ran GAP activates GTPase activity in Ran causing cargo and Ran GDP to dissociate
3. Exportin goes back through NPC
What are the methods for preventing nuclear export/import?
Masking the NLS/NES by:
Inhibitory Protein Binding
Give an example of inhibitory protein binding to prevent nuclear import
IkB binding to NFkB
Give an example of how phosphorylation can prevent nuclear export
Cyclin B1 - phosphorylation prevents it being exported stimulating onset of mitosis
Outline the structure of microtubules
Alpha/beta tubulin dimers polymerise to form a protofilament, 13 of which form a microtubule
Outline microtubule treadmilling
Dimers are added at the positive end and removed at the negative end
In vivo what stabilises microtubule treadmilling normally?
GTP cap on the positive end
Gamma tubulin ring on the negative end
Outline catastrophe and rescue
Microtubules depolymerize 100x faster from an end with GDP-tubulin therefore if the GTP cap is lost at the positive end causes rapid depolymerisation. Eventually the cap is restored
Name two drugs that act as MAPs
Taxal and Colchicine
What are the 6 actin isoforms of mammals?
4 alpha - contractile structures
Beta - cell cortex and lamellipodium
Gamma - Stress fibres
Outline the structure of actin microfilaments
G-actin polymerises to form f actin, which consists of two parallel helical structures
Outline actin microfilament treadmilling
Allows rapid reorganization, normally between C+c and C-c therefore leads to treadmilling
Addition at positive end = loss at negative end
What does tropomodulin do?
Block the negative end of actin microfilaments
What toxins inhibit actin polymerisation?
Cytochalasin D - blocks positive end
Latruncilin - Prevents G-actin formation
How is actin nucleation controlled?
Formins nucleate long unbranched actin filaments. When activated by Rho-GTP the formin straightens out allowing G-actin to access it's FH1 domain
What does fimbrin do?
Moulds actin into microvilli, filopodia and focal adhesions
What does alpha actinin do?
moulds actin into Stress fibres, filopodia and muscle Z line
What does spectrin do?
Support the cell membrane
What does filamin do?
Moulds actin at the leading edge, stress fibres and filopodia
What does ezrin do?
Attach microvilli actin to the plasma membrane
What are the steps of cell migration?
Extension -> Adhesion -> Translocation ->De-adhesion and endo-cytotic pathway
How do proteins establish polarity in migration??
Rho is only active at the rear end
Cdc42 establishes polarity
Give the structure of intermediate filaments
Curved with an alpha helical doain
Dimerize via the alpha helix, 2 dimers form anti-parallel tetramers = Protfilaments, 4 protofilaments = 1 protofibril
What are the main characteristics of intermediate filaments?
More stable than actin microfilaments/microtubules
Provide tensile strength and resistance to mechanical stress
What genes are lamins encoded by?
LMNA - A
What happens to nuclear lamins upon entering mitosis?
Become hyperpolarised causing disassembly
What do nuclear lamins do?
Provide structural support and organize chromatin
What are laminopathies?
Mutations in LMNA
e.g. Emery-Dreifuss muscular dystrophy -> makes nuclei susceptible to rupture
e.g. Hutchinson Guilford progeria syndrome -> toxic effects of farnesylated lamin A, lamina loses elasticity and early senscence
What is the structure of keratin?
Heterodimers of acidic and basic/neutral chains with disulphide bonds providing strength
What are desomosomes?
Structures made of keratin which connect adjoining cells in the epithelia