Cell Division Flashcards
(108 cards)
1
Q
Caner
A
Unregulated Cell division
***cancer = relates to cell division
2
Q
Cells + nucleus
A
Some cells do not have a nucleus
3
Q
Cells without a nuclues
A
- RBCs
- Human Skin
- Lens of the eye
4
Q
RBCs
A
Simple cell that can replicate – bag of hemoglobin
5
Q
Human RBCs vs. other animals RBCs
A
Humans = RBCs do not have a nucleus
Aviation + Amphibians (birds + frogs) = RBCs do have a nucleus
6
Q
Amatopoisis
A
Shed nucleus
7
Q
Human Skin Layers
A
Bottom layer = has nuclei
Strayum Corneum = No nucleus –> has cells but as the cells move up to the top they lose their nuclei –> allows the cells to compress = makes barrier properties
8
Q
Why does the Starum Corneus lose nuclei
A
So that as the cells move up they can compress = gives it barrier properties
9
Q
Lens of the eye Shape
A
- In lenticular shape
10
Q
Layers of lens
A
- Has epithelial Layer
- Has germal layer
11
Q
Lens of the eye division
A
Cells divide in the lens at the equator in the GERMAL LAYER –> as they divide = they differentiate into lens fiber cell with crystals
12
Q
Lens over lifetime
A
Lens throughout your whole life = keeps growing –> reason old people get glaucoma
13
Q
Lens fiber
A
No nucleus (Anuclease)
14
Q
Enucleated
A
Verb!!! – like in SCNT when you enucleate (remove the nucleus)
15
Q
Why shouldn’t lens have nuclei
A
Relates to phase microscopy –> because you can see differnt patrts of the cell in a phase microscope because of diffreent refractive indicies –> This would happen in the lens if it had a nucelus
- Having a nucelus would ruin your ability to see because it would interfere with refractive indicies
16
Q
Parts of the Nucleus
A
- Nuclear envelope
- Nuclear Pores
- Histones
- Non-Histones
17
Q
Nuclear envelope
A
Double membrane system around the nucleus
- Comes from the ER
***NE = connected to the ER because the ER generates the Nuclear Envelope
18
Q
Nuclear Pores
A
VERY COMPLEX
***Sites where mRNA + Proteins leave
Use - passivley diffuses proteins less than 62,500 da
19
Q
Diffusion of nuclear proteins
A
Most proteins in nucleus = can diffuse passively through nuclear pores BECAUSE most of the proteins in the nucleus are Histones
20
Q
Histones
A
Small 20,000 da proteins in the nucleus –> they can diffuse passively through nuclear pores
***Most proteins in nucleus = histones
21
Q
Use of Histones
A
Involved in Epigenetics – can be methylated + Phosphorylated
22
Q
Non-Histones
A
Includes Transcription factors + Other proteins
- Have many different molecular weights
Example – Lamins (Type of intermediate Filament)
23
Q
Example Active transport
A
Nucleoplasm (Nuclear protein) – exceed the passive diffusion weight
- Big protein = 165,000 da = can’t diffuse
- It is a Pentomer –> 5 SU each 33,000 da
- First molcular chaparone discovered
24
Q
Discovery of Nucleoplasm
A
Found originally in African frog toad – found in Xenous Leavits
10% of the protein in the X. Laeveis egg
25
Where do xenobots come from
X.levits
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Function of Nucleoplasm
Chaperone + nucleosome assumably + Genome stability + Transcriptional regulation
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Purpose of Nucleosome
Helps histones arrange
28
Synthesis of Nucleosome
Question -- is it synthesized in the cytoplasm and then shipped through a pore?
Experiment -- track synthesis:
***Use ultrastructural radiograhy
- Use gold labeled anti-nucleoplasm tag -- use Ultrasturctural radiography
- Idea = that it uses an ATP dependint synetshis to be trasnported through the pore SO you want to remove ATP BUT you can't remove all ATP or the cell will die -- SOLUTION = cool the cell to 4 degrees celcius
Result: Nucelosome gets transported through the nucleo pore
29
Lamins
Overall -- type of intermediate filament
Found -- in the inner envelope of nucleus
30
Types of Lamins
1. Lamin A
2. Lamin B
3. Lamin C
31
Function of Lamins
Functionally important because they have many functions
1. Provide structure for keroskeloton -- important for circularization of the nucleus + localizing it in the center of the cell
2. Connects chromatin to nucelar envelop -- at the nuclear pore
3. Phosphorylation of Lamin B triggers nuclear dissolution (MOST IMPORTANT FUNCTION)
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Nuclear Dissolution
Nuclear envelope disappears
33
Inducing nuclear dissolution experiment
Experiment = demonstrates nuclear dissolution occurs during mitosis
Exp:
Fuse a mitotic cell with a cell in G1
Results:
1. Get nucleus disappears = have nuclear dissolution
2. G1 chromatin condenses --> means that something in the mitotic cell that can cause premature formation of chromosomes
***Some entity (MPF) = causes nuclear dissolution via Lamin Phosphorylation
34
Lamin + Nucleus
Have lamin inside nucleus --> when nucleus goes through mitosis = have nuclear dissolution + lamin B is phosphorylated (triggers dissolution)
Late mitosis -- lamin B is dephosphorylated BUT doesnt mean that Lamin B comes back to make new nuclues --> the new nucleus comes from the ER
35
Defect in Lamin A
Associated with progeria
36
Progeria
Precoccious aging disease -- age faster
Issue = not elliptical nucelus --> mishappen nuclei -- not spherical
Normal -- the Fornesyl comes off after the lamin is insertes --> Normal = have Lamina around the rim
Affected -- The fornesyl stays on -- make the Lamin BUT the Fornesyl foes not come off in the end = cause smishapen nucleus
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Lamin Assmebley
Known process -- know that formesyl inserts lamin into memebrane
***Faciliates lamnin into nuclear envelope
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Lonafarnib
Farneyltrasnferace inhibitor used to treat Progeria
***Farneyltrasnferace inhibitor = researched a lot in cancer because Farnesyl links ocoproteins = get cancer but the inhibitors didn't help
39
Cell Cycle (overall)
G0 --> G1 --> Restiction/START point --> S (DNA synthesis) --> G2 --> M phase (mitosis) --> daughter cells
40
What is cell cycle related to
Relates to embryogenesis + Stem cells
41
Fidelity of cell division
Remarkable --> to not make mistake
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Cell division Occurance
Cell divison = occurs all of the time --> theoretically you should get bigger -- Need a balance between cell division and cell death
43
Hematopoeis
Type of cel division -- RBC division
44
G0
First part of cell cycle
***Technically NOT part of G1
- Quecet period --> part of the cell cycle chrachteristic of differenated cell
***Cells in G0 have no interest in dividing unless they are triggered to do so
45
What does G stand for?
Gap -- gap between mitosis
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G1
47
G1 length
For a cell that has a cell cycle of 24 hours --> 9 hours
G1 = 9/24 hours
48
Cancer cell cycle
Cancer cells can't go through the cell division any faster
Example -- some cancer cell cycle = 38 hours
49
Aurthur Pardee
Looked at G1 cycle -- looked at the cell cycle in general BUT he mostly looked at G1
Asked -- what does it take to get through G1 -- what do you need to get from G1 --> DNA replication
Results: Found that you need Growth factors
50
Cells authur used
Used 3T3 cells -- easy to convert from normal to cancer cells
***He introduced 3T3 cells -- made them famous
***Commonly used for cell cylce and oncogene studes because they are easy to convert from normal to cancer cells
***They are isolated from mouse embryo tissue
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What did authur show?
Showed that you need Growth factors to get from G1 --> S phase
Need PDGF --> EGF --> Insulin -- need the growth factors in THAT order
PDGF --> EGF --> Insulin (ALL before S)
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Genes in G1
Can identrify early and late response genes -- All activated by Growth factors
To Study -- look at mRNA over time
Results:
1. Get early response genes (see increase in mRNA early)
2. See late response genes (increase in mRNA later)
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Early + late response genes
If you prevent the degradation of early response genes = then you don't get the late response genes = SHOWS that they are connected
***Don't get late response without the early
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Studying Cell cycle
1. Most of the work done = done by fusing cells
2. Need to synchronize the cells so that they are all going through the cell cycle at the same time -- to study you need cell synchrony (doesn't occur naturally -- need to induce it)
55
Inducing Cell synchrony
It won't occur natural -- you need to induce it
***Most wats are based on G1
1. Amino Acid deprivation -- stall cells in G1 --> because no AA = the cells can't make proteins = stalls
- Can add back later
2. Serum deprivation -- stalls in GA (Prefered way)
- remove growth factors
3. Protein synthesis inhibtors -- stalls in G1 + can stall in G2 (since it can stall in both = this is not prefered method)
4. DNA synthesis inhibition -- stalls in S
5. Nucleosome inhibitor -- stalls in M because spindle us inhibited
56
Cell synchrony experiment
Take synchronized cells and watch them divide
Result: Get a graph with a very strange profile --> need to figure out what happened
- Found that the only way to get that data was if the cell cycle was 24 hours +/- 6 hours -- means that the cycle is varaible
57
What part of the cell cycle is variable
G1 phase -- most variable in time
Variability = explains why the cells fall out of synchrony over time
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Two points in the G1 cycle
1. G1 checkpoint
2. Resiction point
59
G1 checkpoint
Checking fidelity of process --> anylzye the fidelity (correctness) of G1 process
***Look at DNA
60
G1 restiction point
G1/S boarder -- Go or no go point
***At this point -- goes to S phase
IF it doesn't go to S = it goes backwards in G1 = spends more time there
***aka "pardee point"
61
Where is restiion point found
Found in mammal cells + non-mammal cells BUT not in yeast cells
***Start in yeast
62
Do cancer cells differ from normal cells in their repsective cell cycle conpartment time?
Answer -- they are not different
Experiment -- look at different compartment times (look at each separately)
Take G1 synchronized cells -- take synchrionized cells and watch
1. In G1 --> Add H3-thimadine --> Look at how much time it is before you see labeled cells -- because the labeled cells = tell you they are in S phase
- Once see labeled cells = know they are done in G1 -- now in s (Because they are doing DNA replication)
2. In S --> look at randomly dividing cells -- all go through the cell cycle at different times
- At H3-thiamdine --> look at the number of labeled cells -- if 3/10 are labeled = then do 3/10 X 24 hours = gives you the S phase time
3. G2 Time --> use randomly dividing cells again
- Use H3 thiamdine to pick only cells in S phase --> once they are in M = you will see lableed chromsomes
- can see m cells with radioactive probe = tells you length of G2
- can use the same system to find length of m Phase
Result: Normal cells + cancer cells = have the same compartment times
63
Adding H3-Thimadine
Add to cells to probe cells that are going through DNA replication (cells starting to divide)
64
What controls cell cycle transit
Cyclins + associated Cyclin dependent kinases
65
Rhudamin + hunt
Did classic experiment about cell cycle movement
Exp:
Took naturally occuring synchronized cells -- used sea urchin embryos (used ionmycin or sperm -- added at once to the eggs)
THEN they ran SDS gel to see proteins
Results: Discovered Cyclins
66
Types of cyclins
There are many BUT only two are important:
1. Cyclin B
2. Cyclin A
67
Mitotic Cyclins
Cyclin A and Cyclin B -- govern whether the cell goes through mitosis
68
What is associated with cyclin
Cyclins have partner cyclin dependent Kinases
***Cyclins won't do anything unless has partner CDK
NEither alone can do anything --> they are a pair
***They can be influenced by many things -- many things inhibit their function
69
CDKs
CDKs = increase and decrease over the cell cycle
70
Amounts of Cyclinc over cell cycle
Cyclin e -- G1 --> S
Cyclin A --> G1 --> S --> G2 (decrease at m)
Cyclin B -- M phase
Cyclin D --> G1
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Mitosis
72
What is required for G1 passage
Cyclin D
***Cyclin D = associated with G1
73
Cyclin D experiment
Questioon -- Us Cyclin D required for G1 cell cycle transit
T
ake G0 cell --> Add Growth factors --> Add BrdU --> Look at BrDU incorporation after time
Control cell -- leave alone
- Result: have BrdU positive cells -- means they are in S phase
Experimental cell -- add anti-cyclin D antibody -- blocking cyclin D
- Result: BrdU negative cells
***BrdU = like H3-Thiamine BUT safer
If add AB early in G1 = the control go to S BUT the experimental do not
If add AB late in G1 = the conrtol go to S and the experimental go to S (if add the antobody late in G1 = teh cells still go to S phase)
74
Use of BrdU
BrdU = like H3-thiamine BUT is safer -- floursecnt alternative to H3-Thimadine
75
S phase
DNA synthesis
76
How do we study S phase
We can use electron microscope to tell us about S phase
Exp -- Take cells in S phase --> Open cells + open the nucleus
Result: See replicons/replication bubbles -- bubbles tell you there may be multiple points of replication + DNA synthesis may be directional
77
Two ways to study S phase
1. Elecrton microscope
2. Use Fiber autoradiogrphy
78
Autoradiography in S phase
Use fiber Autoradiography -- Use autoradiography with chromosome inside --> Shows that electron microscope were right
79
Replication origins
Know there are many points where replication generates + H3 thiamine incorporates
Have Point ORC --> ORC makes replication bubble
80
What controls S phase
Cyclin A-CDK --> move cell cycle to mitosis
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Cyclin B
Main mitotic cyclin
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G2 phase
Some protein synthesis is required
***We know very little about G2 because non one cares about G2 = no one studies G2
83
What Happens in G2
1. Cell verifies that al of the DNA has been correctly duplicated and all DNA errors have been corrected
2. Chromosome condensation is initiated
3. Early organization of the cell cytoskeleton
4. Mitotic cyclin dependent kinases initiate activity
84
Mitosis (Overall)
Prophase -- Spindle poles
Metaphase -- sister chromatids
Anaphase
Telophase + Cytokinesis
85
M phase
***get new nuclear envelope
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Shortest Phase in Cell cycle
M phase -- know by looking at cell kinetic studies
87
Lamins in M phase
Lamin B = gets phosphorylated by MPF
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What triggers movement through M Phase
MPFs
E.Leavis -- Maturation phase factor
Mammalian cells (Cell fusion) -- Mitosis Phase factor
89
Discovering MPF
2 groups using 2 model organisms:
1. Using X.levais --> perfect organism to study because cycles go through sequence
- take cytoplasm out of cell --> put in new cell --> idnduces mitosis iommediatley = have factors to promote maturing in egg
Found MPF -- Matruartion Phase factor
2. Used cell fusion experiment --> tells you what tells cell to go through mitosis
Overall -- took a test tube + added cytoplasm from mitotic cell + nucleus + ATP --> Nucleus will duplicate
Used mammalian cells --> Did cell fusion --> found MPF
MPF -- Mitosis Phase factor
90
Use of MPF
MPF = phosphorylates Lamin B = causes nuclear dissolution
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When is MPF highest
MPF = highest at Metaphase
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MPF cycle
Metaphase = high MPF --> In anaohase the Cyclin B is uqiquinated = decrease cyclin B BUT still high CDK--> make more cycle B over the course of the cycke = have more at mitosis again
Low MPF at telophase
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What is MPF
MPF = Cyclin B + CDK
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MPF function
Phosphorylate Lamin B = have dissociation of nuclear envelope = reason why MPF is high during metaphase
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Cyclins in Cell cycle
Cyclin A = S phase + G1
Cyclin B = M phase
Cyclin D = specific to different cell cycle points
96
Ruth Sager
Discovered Cell cycle check points
***First to suggest that we have tumor supressor genes --> first to suggest that we have genes in system to prevent cancer
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P53 (chart)
98
Cell cycle Checkpoint
Sites of control -- we have many cell checkpoints
***Each cell cyle compartment = has checkpoint -- check fidelity
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Who discovered cell cycle check points
Ruth Sager
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Discovering Cell cycle checkpoints Experiment
Take normal cell + cancer cell --> Fuse cells --> Get heterokeryon ( have two nuclei) --> get hybridioma
Look several generation later -- Look at the phenotype --> Acts more like a normal cell NOT like a cancer cell
LOOK EVEN LATER -- look at phenotypes --> now you have cancer cell
Conclusion: Maybe we have genes in system that prevent camcer cells -- maybe we have tumor supressor genes
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Heterokeyron
Have cell with two nuclei --> during the fusion process --> have reorganzation of nuclei + chromosome exchange -- chromsomes are lost + genes are lost
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Where is p53 active
p53 = active in many checkpoints
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p53 name
Called "guardian of the genome"
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p53 (overall)
Tumor supressor gene
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p53 action
Triggered by DNA damage in G1
p53 = very unstable until it is phosphorylated by a ATMR --> activates p53
When p53 is active = it can do two things:
1. Fix probelm -- if few DNA errors = fixes the DNA errors --> cell goes to S phase
2. Kill cell --> if there are many DNA errors = cell goes through apoptosis
***p53 = checks for DNA errors
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Where were checkpoints first discovered
First discovered in Yeast --> budding yeast + Fission yeast
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Budding yeast Cycle
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Fission yeast Cycle
