What are the 2 main functions of the nucleus?
- Where chromosomes are localized and replicated
2. Where DNA is transcribed
What are 2 key features of the nuclear envelope? And What name each of it’s components.
- Double membraned system
- Outer membrane is continuous with the endoplasmic rectilium
Outer membrane, perinuclear space, inner membrane
What cell structure component is the nuclear matrix analogous to?
The cytoskeleton; helps maintain shape
What is type of microtubules is the nuclear lamina made up of?
Intermediate filaments
What is the function of a nuclear pore?
To provide a direct connection from the cellular cytosol to the nucleoplasm
What type of proteins are nuclear pore complexes made up of?
Nucleoporins
What is the granule in the centre of the porins called? What is its function?
Transporter; moves molecules across the nuclear envelope
What is the name of the protein that hold the NPC to the nuclear envelope?
Anchor proteins
On what side of the nuclear envelope do the projecting fibres create a basket shape?
On the nucleoplasmic side
What is usually a) imported b) exported c) mediated back and forth from the cytoplasm to the nucleus via nuclear pores?
a) enzymes and proteins
b) mRNA, ribosomes
c) small particles and molecules, ions.
What are the 2 types of transport possible across the nuclear membrane? Explain them.
1) simple diffusion (passive facilitated)
- small particles can pass through via tiny aqueous diffusion channels
2) active transport
- too large to transport through nuclear pores
- have NLS or NES (8-10 aa)
Explain the 5 step journey of a cytoplasmic protein with an NLS and importin.
- Cytoplasmic protein w/ NLS recognized by importin receptor protein, importin binds to the NLS, importin guides complex to nuclear pore
- Importin-protein complex transported into the nucleus via the transporter moving them through the nuclear pore
- Now in the nucleus: importin associates w/ Ran (GTP binded protein), releasing the cytosolic NLS-containing protein
- Ran-GTP importin complex is exported out of the nucleus through the NPC
- Now in the Cytoplasm: GTP attached to the Ran is hydrolyzed (promoted by GTPase activating protein; GAP), releasing importin. Nuclear transport factor 2 then shuttles Ran-GDP back into the nucleus
Export of mRNA outside of the cell does not require Ran. What are the 2 steps required then to transport mRNA outside of the nucleus, into the cytoplasm?
- Adapter protein binds to the mRNA
- adapter proteins have NES - NES sequences recognized by exportins which mediate transport of the mRNA-adapter protein complex into the cytoplasm via the transporter through the NPC
What is responsible for maintaining high levels of Ran-GTP inside the nucleus?
Guanine-nucleotide exchange factors
Chromatin have localized areas that the occupy; chromosome territory. How did we find this?
Through in situ hybridization using nucleic acid probes
One of the components of the nucleolus are fibrils. What takes place in the fibrils? What are granules?
DNA that is being transcribed into rRNa (ribosomal RNA). Granules are rRNA molecules that are being packed with proteins
What is the nucleoid?
The localized area of the bacterial cell where DNA is bound
What are the 5 steps of chromosome packing?
- DNA wound around a histone octamer to form a nucleosome
- Nucleosome then coils to create 30 nm chromatin fibre
- 30 nm chromatin fibre attach to scaffolds to create loops
- Scaffolded DNA loops coil to create heterchromatin
- During cell division: heterochromatin coils to create chromosomes
Explain the functions of H1, H2A, H2B, H3, H4.
H1: facilitates the formation of 30 nm chromatin but NOT part of the octamer
H2A,H2B,H3,H4: part of the octamer with lysine & arginine»_space; amino groups that make the histones overall positive and therefore attract and guide DNA winding
What stabilizes the DNA loops in mammals?
Cohesin proteins
DNA loops + nonhistone proteins (scaffold) = ?
Chromosomal Scaffold
How does the cell regulate what parts of the DNA are highly conserved and what parts are highly expressed?
By altering histones; protruding tail can be tagged by a methyl, acetyl, phosphate, or other groups
Methylation by histone methyltransferase : can activate or repress transcription
Acetylation by histone acetyltransferase:
Deacetylation by histone deacetyltransferase:
How long is the generation time in mammals?
18-24 hours
What are the 3 phases of interphase?
G1 phase: cell grows and synthesizes proteins and organelles (some are arrested in G0 phase)
S phase: DNA is synthesized and chromosomes duplicated
G2: cell continues growing, synthesizes proteins needed for cell division
Explain the 2 phases of Mphase
- Mitosis: sister chromatids are separated 7 partitioned into 2 nuclei
- Cytokinesis: division of cytoplasm
What are the 5 stages of mitosis?
- Prophase:
What are the 3 things that occur during prophase?
- Chromosomes condense
- MT begin to form mitotic spindles
- Nucleolus disappears
What are the 3 things that occur during prometaphase?
- Nuclear envelopes disappear
- Aster move to opposite poles
- Microtubules connect to the chromatid kinetochores
What are the 3 things that occur during metaphase?
The sister chromatids line up at the metaphase plate
What happens at Anaphase?
Sister chromatids are pulled to opposite poles of the cell
What 3 things that happen during telophase?
- The cell forms a cleavage furrow in humans
- Nuclear envelope and nucleolus reforms
- Chromosomes decondense
What is the name and function of the 3 main components of kinetochores?
- Inner kinetochore: has proteins that bind to centromeric DNA
- Outer kinetochore: has proteins that attach to the plus ends of microtubules
- Fibrous corona: function unknown
What are the 3 key things about the cell cycle that are highly controlled?
- Timing: all events of phase occur in order and ath the appropriate time
- Completion: all events in each phase have occured before next begins
- Response: respond and adjust according to external conditions
Describe the 3 routes that can occur during the G1 –> S checkpoint
- Cell passes restriction points
- Enter into G0
- Fail, and this try and reenter G1 and return to the checkpoint
Describe the 2 possible routes when the cell comes to the G2 –> M checkpoint
- Cell passes onto m phase
2. Arrested in G2 (just like G0)
What is the function of the metaphase —> anaphase checkpoint?
Ensures non-disjunction doesn’t occur
What are the 4 requirements for a cell to pass the G1 —> S checkpoint?
- Growth factor (kava)
- Nutrients (soil)
- Cell size (big pot)
- No DNA damage (eaten leaves)
What are the 3 requirements for a cell to move from G2 —> M phase?
- No DNA damage
- Good cell size
- DNA replication complete
What are the requirements for a cell to move from metaphase —> anaphase?
Chromosomes are all perfectly attached to the (+) end of the microtubules
What 2 protein structures do cells use to control the cell cycle? And how?
Protein kinases (phosphorylate)
Protein phosphatases (dephosphorylate)
When is the concentration of mitotic cyclin the greatest?
At the end of G2
What are the 2 ways that the cell controls the activity of mitotic Cdk-cyclin?
- by controlling the availability of mitotic cyclin
- low [mitotic cyclin] = low [mitotic Cdk-cyclin] - by phosphorylating mitotic cdk
- 2 inactivating phosphate groups added by inhibiting kinases & 1 activating phosphate group added by activating kinases = inactive mitotic cdk-cyclin
- phophatase removes 2 inhibiting phosphate groups = active mitotic Cdk-cyclin (activated Cdk stimulates more phosphatases creating positive feedback loop)
What are the 4 ways that mitotic Cdk-cyclin promote mitosis?
- Phosphorylates lamin on the nuclear membrane: breaks down nuclear membrane
- Phosphorylates condensin: triggers condensing of the chromosome
- Phosphorylates microtubule associated proteins: triggers assembly of mitotic spindle
- Phosphorylates anaphase promoting complex (ubiquitin ligase to mark for destruction): triggers degradation of mitotic cyclin = low [mitotic Cdk-cyclin] (negative feedback) & degradation of securin which usually inhibits separase which breaks down cohesins) = sisterchromatids can separate
How does G1 Cdk-cyclin regulate progression through the restriction point?
Does this by phosphorylating retino blastoma protein to intiate DNA replication
- RBP is phosphorylated 2. RBP lifts from E2F transcription factor 3. Allows E2F to activate transcription of genes that code for proteins that initiate DNA replication
What is a tumour represser gene?
- Loss or inactivation can lead to cancer
2. 2 mutations required to lose function and promote cancer
What is the a) normal and b) mutated function of retino blastoma protein?
Normal: prevents cells from passing from G1 –> S without the appropriate growth factor signals
Mutated: allows cell to pass from G1 –> S without appropriate growth factor signals
How does p53 act as a tumour suppressor?
If there’s DNA damage, p53 is phosphorylated by check point kinases which are phosphorylated by ATM protein kinases and attempts repair induces cell cycle arrest and cell death. But if there was a mutation in the p53 protein and the cell cycle and cell death were not triggered in a cell with damaged DNA, this creates cells with damaged DNA that may be cancerous
How is the cell cycle arrested when DNA replication is not complete even as it reaches the DNA replication checkpoint?
Cells that have incomplete DNA replication are prevented from progressing through to the m phase bc they fail to undergo the final dephophorylation step via a phosphatase
What are the 2 defining features of an oncogene?
- Presence of the gene can lead to cancer
2. Only 1 mutation is required to enhance function and promote cancer (gas pedal)
What are the 3 types of spontaneous mutation that can occur?
- Mispairing of bases due to tautomers
- Slippage during replication
- Damage to individual bases
What are the 2 types of environmental mutagens?
Chemicals and Radiation
Name and explain the 3 types of chemical environmental mutagens?
- Base analogue: resemble nitrogenous bases and are incorporated into DNA
- Base modifying agents: react with DNA bases to alter their structures; creating DNA adducts
- Intercalating agents: insert between adjacent bases distorting the DNA’s structure
How does is a) ultra violet radiation b) ionizing radiation an environmental mutagen?
A) Triggers formation of pyrimidine dimer formation
B) removes electrons from molecules creating reactive intermediates that damage DNA