Cell Birth Differentiation Life

Front 1

What is flow cytometry?

Back 1

Technique used to determine the number of cells in G1, S, or G2-M. Add propidium iodine - they take up dye that goes to nucleus and intercalates into DNA. Amount of stain directly proportional to amount of DNA.

Front 2

What is the mammalian cell cycle basic regulatory apparatus?

Back 2

cyclins, cdks, and ckis (cyclin dependent kinase inhibitors)

Front 3

What controls progression between S and M phase?

Back 3

It is a conserved regulatory apparatus - links cell cycle with extracellular signals that control proliferation

Front 4

What regulates cyclin?

Back 4

Regulated by transcription, translation, and proteasomal degradation

Front 5

What regulates Cdk activity?

Back 5

phosphorylation/dephosphorylation

Front 6

What are CKIs?

Back 6

protein that binds to either cyclin/cdk complex or cdk and inactivates cdk activity

Front 7

What is the major G1 cdk?

Back 7

Cyclin D with Cdk4 and Cdk6 partners. Cyclin D binds to all G1 and G1/S Cdks. It is upregulated by growth factor stimulation of transcription

Front 8

What does Rb control?

Back 8

The restriction point. If Rb is non-functional, will have uncontrolled growth.

Front 9

How can growth arrest be induced?

Back 9

G1 growth arrest - increased destruction of cyclin D, increased phosphatase action to dephosphorylate kinase CDK4 or CDK6

Front 10

What are the 2 families of CKI proteins?

Back 10

CIP/KIP - Bind to cyclin/CDK complex

INK4 - find to the CDK

Both occur at G1/S checkpoint

Front 11

If CKI is bound to a complex, is it active or inactive?

Back 11

INACTIVE! blocks the ability of the complex to phosphorylate Rb.

Front 12

Why is there a cell cycle check point?

Back 12

TO insure that incomplete or damaged chromosomes are not replicated. Ensure genome is replicated only once per cell cycle.

Front 13

Restriction point

Back 13

Controlled by Rb. Only one copy is typically sufficient. Rb is compromised in many tumors.

Front 14

What are the 4 other checkpoints (not one with Rb)?

Back 14

-G1/S - DNA damage
-G2/M - Is cell big enough, DNA replicated, DNA damaged?
-Mitotic Spindle - Are all chromosomes aligned?
- S phase - is all DNA replicated

Front 15

Role of p53 with low level DNA damage

Back 15

Low level DNA damage - induces p53 and growth arrest. p53 induces p21, p21 binds to CDK and blocks G1/S progression.

Front 16

Role of p53 with high level of DNA damage

Back 16

Induces high levels of p53 and Bax. (Bax is a Bcl2 protein). Bax acts on mitochondria by blocking Bcl-2, making mitochondrial wall permeable - releases cytochrome C - promotes intrinsic pathway of cell death.

Front 17

What are two pro-apoptotic members of the Bcl2 family?

Back 17

Bax and Bak

Front 18

Intrinsic Pathway of Apoptosis

Back 18

Induced by p53 and other effector proteins

Front 19

Where is cytochrome C located?

Back 19

In the mitochondrial intermembraneous space

Front 20

What does cytochrome C active?

Back 20

Cytochrome C activated an Adaptor protein. It assembles and recruits procaspase 9 to form an apoptosome

Front 21

How is procaspase activated?

Back 21

Caspases exist as proenzymes. Proteases (cysteine/aspartate) are involved. They activate endonucleases that cleave DNA.

Front 22

What are two anti-apoptotic members of the Bcl2 family?

Back 22

Bcl2, BclXL. Pro-apoptotic are Bax, Bak, Bid

Front 23

Do cancer cells have to have a chromosomal translocation event to upregulate Bcl2 levels?

Back 23

No

Front 24

What are characteristics of an apoptotic cell?

Back 24

Cell membrane is intact until the end. (Trypan Blue dye can show this - it can only pass through a compromised membrane) Has shrinkage, blebbing, chromatin condensation, DNA degradation, nuclear fragmentation, apoptotic bodies

Front 25

What is an apoptotic body?

Back 25

cell membrane enveloped cellular organelles, degraded DNA, and other material. Engulfed by cells or macrophages

Front 26

phosphotidylserine in dying cell

Back 26

This does not set up an inflammatory reaction. PS is flipped to the outer leaflet in a dying cell, annexin V binds to dying cell.

Front 27

DNA degradation during apoptosis

Back 27

Chromosomal DNA is fragmented due to endonuclease cleavage - cleaves at linker regions. Flow cytometry shows a portion in subG1 area. Less than 2n

Front 28

Extrinisic Pathway of apoptosis

Back 28

Happens WITHOUT cytochrome C. Receptor/ligand interaction activates caspases 8 &/or 10 - activates executioner caspases 3, 6, 7 and endonucleases

Front 29

How are the intrinisic and extrinisic apoptosis pathways connected?

Back 29

They are connected through Bid. p53 can activate caspase-8, which activates Bid, t-Bid activates cytochrome C.

Front 30

What does the loss of p53 cause?

Back 30

leads to uncontrolled growth and genomic instability where genes may be amplified. Both alleles of p53 must be mutated or deleted

Front 31

What are some mechanisms for Multidrug resistant cancer cells?

Back 31

Loss of p53
Loss of Rb
cyclin D up-regulation
growth factor up-regulation
increased GF receptors
Bcl2 upregulation

Front 32

What regulates Rb activity?

Back 32

cyclinD/CDK

Front 33

What is the clinical application of flow cytometry?

Back 33

determine efficacy of cancer drugs. See growth arrested cells. Proliferating population

Front 34

What are common initiator/executioner caspases?

Back 34

Initiator - 8,9,10
Executioner - 3, 6, 7

Front 35

What does caspase 3 target?

Back 35

activates endonuclease

Front 36

What is the target for caspase 6?

Back 36

cleaves Lamin A

Front 37

What is the target for caspse 7?

Back 37

cleaves poly ADP ribose polymerase

Front 38

What is necrosis?

Back 38

When a cell membrane is compromised early. Genomic DNA and contents of cell spill out. Causes inflammatory response

Front 39

What is camptothecin?

Back 39

It inhibits topoisomerase I