Cell Bio Exam 2
To prevent cells with damaged DNA from passing through the G1 checkpoint, what are ATR or ATM doing on Chk1 or Chk2?, which then modulate protein phosphatase cdc35.
ATR and ATM are protein kinases that sense and initiate repairs when DNA is damaged. ATM and ATR also alert the cell to arrest the cell cycle until the DNA is repaired. ATM and ATR do this by phosphorylating Chk1 and Chk2 which then phosphorylate Cdc25. Cdc25 is an important protein involved in the activation of Cdk2/cyclin E so when phosphorylated Cdc25 is degraded, Cdk2/cyclin E is never activated and the cell cannot pass the G1/S checkpoint
List the different Ubiquitination events that occur over the cell cycle. Give their names and roles and regulatory mechanism(s).
-Cyclin Degradation: regulation of CDK-Cyclin complexes. CDKs activate cyclin ubiquitin ligases that destroy cyclin and inactivate CDK. -CKI Degradation: Reactivating the cell cycle. CKIs are degraded by SCF complexes and ubiquitin ligases, activating CDK-Cyclin complexes. -Metaphase/Anaphase Transition: regulation of sister chromatid separation. Anaphase Promoting Complex APC and cdc20 activator ubiquitinates securin. Separase is then freed from securin and cleaves cohesin at the centromere, allowing sister chromatids to separate.
The transport of proteins across the nuclear envelope has some distinct mechanisms compared to the transport of proteins across the mitochondrial membrane, although both are mediated by the presence of a signal sequence. What are the other differences?
-Mitochondria have an inner and outer membrane that proteins need to be transported through. -Nuclear transport has two signals: NLS (import) and NES (export). Mitochondrial transport has signals that depend on the destination. -Nuclear transport uses RanGTP, mitochondrial transport uses proton motive force as well as chaperones to move proteins around.
Which events does take place during the death receptor pathway of the tumor necrosis factor receptor (TNFR1) signaling?
1. Binding of death ligands of TNF family to death receptors 2. Death receptor ligation recruits adaptor proteins FADD via death domain DD. 3. DED-DED interaction recruits initiator procaspase 8. They dimerize and are activated by induced proximity. The death inducing signaling complex DISC 4. Activated caspase 8 leaves DISC and cleaves and activates executioner procaspases. 5. Executioner caspases cleave substrate. 6. Apoptosis
How would you design an experiment to demonstrate that the NLS of the yeast protein Swi5 is sufficient for the transport of this protein into the nucleus?
2 possible experiments: 1) Alter the sequence of Swi5 so that the NLS is deleted and demonstrate (via fluorescence) that the altered Swi5 protein cannot be detected in the nucleus. 2) Create a fusion between the NLS of Swi5 and a gene that is not normally found in the nucleus and demonstrate (probs w/ fluorescence) that this new fusion protein is now detected in the nucleus
How many kinetochores are there in a human cell at mitosis?
92 kinetochores in HUMAN during mitosis; TWO kinetochores on each of the 46 chromosomes
Which one of the following functions is NOT performed by a motor protein within the mitotic spindle?
>>> motor proteins within the mitotic spindle do NOT elongate microtubules at their end
What is the nucleolus?
A small dense spherical structure in the nucleus of a cell during interphase. It is best known as the site of ribosome biogenesis as well as rRNA transcription, processing & subunit assembly. in short, this organelle = where rRNAs are synthesized/processed AND where ribosomes subunits are assembled
CENP-E protein in an unattached kinetochore can slide along an adjacent, attached kinetochore fiber toward a centered chromosome to help the unattached chromosome get closer to the center. This may help shorten the distance for the microtubules searching for an unattached kinetochore. True or false?
CEMP-E protein on a mono-oriented chromosome will grab a neighboring bi-oriented chromosome and pull itself towards the mitotic plate, increasing the likelihood of the kinetochore being found and bound by other opposing spindle microtubule. It's like the mono-oriented chromosome is throwing a rope to the neighboring chromosome to pull itself back to safety.
Explain how the protein Bid links the death receptor pathway and the MOMP pathway to apoptosis.
Caspase-8 (activate as a result of the death receptor pathway) cleaves the BH3-only protein, Bid; thereby, activating it. Activated Bid in turn activates Bax and Bak to promote MOMP and cytochrome c release. As a result, caspase-9 is activated, leading to apoptosis.
How does cdc25 affect commitment to Mitosis?
Cdc25 is a phosphatase that removes the inhibitory P on CDK1/cyclin b(mitotic cdk), therefore activating M-CDK/cyclin b which promotes mitosis. >>> basically, if the Cdc25 enzyme is activated, then mitosis should be occurring -- as this phosphatase = AN ACTIVATOR !!!! >>> This breaks down the membrane surrounding the nucleus causing the nuclear envelope to break down and is the beginning of prometaphase stage of mitosis
What are the critical events for M phase to occur in animal cells?
Cdk1/cyclin B is activated, all the DNA is replicated correctly, and there are the proper nutrients and growth factors available to the cell.
What type of molecular sensors a cell possesses to respond to the late outcome of the absence of functional Retinoblastoma (Rb) protein?
Certain extracellular signals such as 'tumor necrosis factor-alpha' can detect the lack of the Rb protein and trigger apoptosis in the cell.
What are Chk1 or Chk2 doing on Cdc25?
Chk1 and Chk2 are kinases that phosphorylate cdc25. Phosphorylated cdc25 is degraded which leads the inactivity of Cdk2/cyclin E and arrest of the cell cycle while the DNA is repaired.
What is congression (that really important part of metaphase) during mitosis?
Congression is the lining up of chromosomes into the mitotic spindle equator. Congression is formed through pulling forces, leaving a zero net force on the chromosomes.
During mitochondrial outer membrane permeabilization (MOMP) what is allowed to escape from the mitochondria?
Cytochrome C is released.
Briefly explain how cytochrome c activates caspase-9.
Cytochrome c is released into the cytosol from the mitochondrial and binds to APAF-1. The APAF-1 complex (aka the apoptosome) then changes conformation and exposes its CARD domains which binds with CARD domains of initiator caspase-9
Initiation of DNA synthesis and replication of centrioles are tightly coordinated events. Briefly explain how this coordination is achieved.
DNA synthesis and centriole replication are coordinated because they are controlled by the same cdk/cyclin complexes; G1: Cdk2/cyclin E, and S Phase: Cdk2/cyclin A.
Explain how death receptors activate initiator caspases
Death receptor ligation recruits adapter proteins FADD vua the death domain. The DED-DED interaction with the FADD recruits the initiator procaspases where dimerization and cleavage causes activation by induced proximity.
The nucleus is completely filled with chromatin. True or False?
False Interchromosomal domain, nucleolus, etc.
The signal for the spindle assembly checkpoint comes from the spindle poles. True or false?
False The signal comes from the kinetochores (p. 667). Kinetochores not under enough tension release the "wait anaphase" signal (Mad2 and BubR1). This signal delays the activation of APC cdc20 which is responsible the degradation of the cohesins holding the sister chromatids together.
XIAP is an inhibitor of caspases. How does it work?
It binds to the loops on the caspase and prevent cleavage that would activate them. XIAP acts as a ubiquitin ligase and tags initiator caspase-9 and executioner caspases -3 and -7 for degradation. On executioner caspase-3, XIAP blocks the cleavage site the initiator caspase uses. With the cleavage site blocked, caspase-3 cannot be activated
What is one function of the Cdk1-cyclin B complex in mammalian cells?
It promotes mitosis.
What mechanism ensures that cargo-protein complex does not exit the nuclear pore complex (NPC) once it has passed through the nuclear pore and into the nucleus?
Keeping the RAN protein as GDP will keep the cargo from being exported. Also, only protein cargos with a NES tag will bind to exportins and be transported to the cytoplasm. Protein cargos that are meant to stay in the nucleus do not have NES tags, just NLS tags so they don't bind to exportin.
Leptomycin is a compound that inhibits the growth of Human Immunodeficieny Virus in human cells, yet it cannot be used to treat HIV patients because it is toxic to all of a patient's cells. Explain how leptomycin functions in human cells.
Leptomycin inhibits Crm 1, the main exportin protein, so molecules that rely on Crm 1 to exit the nucleus are also inhibited. This build up of proteins in the nucleus disrupts cell function and is toxic making leptomycin useless as a drug to treat HIV.
After the nuclear envelope breaks down, microtubules gain access to the chromosomes and, every so often, a randomly probing microtubule connects with a kinetochore and captures the chromosome. True or false?
Microtubule-Kinetochore connection is a "search and capture" process.
Is all protein trafficking through nuclear pores unidirectional?
No, proteins selectively enter and exit nucleus through nuclear pores (Both directions are used) NO THEY CAN IMPORT AND EXPORT ("proteins can selectively enter AND exit the nucleus through nuclear pores" -- thus protein transport thru nuclear pores = BIDIRECTIONAL)
What are nuclear lamin proteins making up?
Nuclear lamin proteins make up the nuclear lamina, which plays a role in the nuclear envelope assembly. the nuclear lamina (is a filamentous meshwork just beneath the inner membrane) = impt bc: 1) rebuilds the nuclear envelope after mitosis 2) provides structural support to cell
How the ORI (origin of replication) being licensed to fire?
Once the Pre-RC is assembled, cyclin a can activate the CDK so it can phosphorylate cdc6, which allows the origin to fire.
Explain why cells that lack functional p53 are more prone to accumulate mutations in their DNA?
P53 is a checkpoint gene that restrains the cell cycle when DNA is damaged. When damaged DNA is allowed to replicate, more and more replication errors occur and mutations can accumulate.
How are Apoptotic cells completely removed from the tissue they reside in?
Phagocytes engulf the fragments of the apoptotic cells. >>>> these apoptotic cells send "eat me" signals or "find me" signals
Explain how a flippase enzyme assists in the clearance of apoptotic cells in mammals.
Phosphatidylserine is normally localized to the inner leaflet of the plasma membrane, this asymmetric distribution is maintained by an ATP-dependent "flippase" that flips the phospholipid from the outer to inner leaflet of the membrane. During apoptosis the asymmetry is not maintained and phosphatidylserine is found in the outer leaflet, where is can bind to several soluble molecules that can act as bridges to phagocytic cells. These proteins recruit phagocytic cells which then engulf the apoptotic cell.
There are several mechanisms for regulating Cdk activity. List three distinct mechanisms of Cdk activation or inhibition.
Phosphorylation: Wee1 phosphorylates in the inhibitory site of CDK. CAK phosphorylates in the activation site. Cdc25 is a phosphatase that removes the inhibitory phosphate placed by the Wee1 enzyme, but leaves the activator phosphate placed by CAK>> therefore Cdc25 = an activator. Proteolysis: CDK/cyclin activates cyclin ubiquitination ligases that destroy cyclin, inactivating the CDK.
What is one advantage to a cell of having a nucleus?
Protects the DNA from damage.
What are the forces in Anaphase B that move sister chromatids and poles further apart?
Pushing forces from kinesins and pulling forces from dyneins "DYNEINS ATTACHED TO PLASMA MEMBRANE PULL, AND MOTOR PROTEINS THAT INTERACT WITH OVERLAPPING MT OF OPPOSITE POLARITY" The dynein motor proteins move towards the minus end of the astral microtubules and generate pulling forces on the centrosome. Kinesin motor proteins (such as Eg5) walk towards the plus end of the overlapping interpolar microtubules and generate pushing forces on the centrosome.
Explain how activation of tumor necrosis factor receptor (TNFR1) can induce anti-apoptotic signals?
The ligation of TNFR 1 by TNF is capable of producing signals that both block and promote apoptosis. If NF-kb is activated, TNF fails to trigger apoptosis but instead participates in the inflammatory response. Alternatively, if NF-kb is blocked or if RNA or protein synthesis is inhibited, TNF triggers apoptosis.
What is the pre-Replication Complex (RC)?
The pre-RC is assembled during G1 after the checkpoint, but never when CDK-cyclin is high. The ORC (origin recognition complex) is bound to the ORI (origin of replication), then the licensing proteins cdc6 and cdt1 bind to ORC, then MCM helicase binds, completing the Pre-RC.
What prevents an origin of replication from being used more than once per cell cycle?
The s-cdk phosphorylates cdc-6 so that it dissociates from the ORC. The MCMs can no longer bind with the absence of cdc 6. This results in the prevention of the pre replication site from refiring. Also, cyclin a needs to be present to activate the CDK so it can phosphorylate cdc6, which allows the origin to fire.
How is the position of the spindle used to determine the formation of the contractile ring?
The spindle determines where the contractile ring forms. Stem bodies, bundles of parallel MTs, form where the metaphase chromosomes aligned. As anaphase progresses, the stem bodies coalesce into one large bundle called the mid body. Stem bodies signal to the cortex to cause the formation of the contractile ring.
What is the role of the spindle assembly checkpoint (SAC)?
To verify correct attachment of microtubule and kinetochore.
Importin bound to NLS-containing cargo is transported into the nucleus where the complex binds Ran-GTP and gets dissociated. True or false?
True Slide 24, Once inside the nucleus, interaction with Ran-GTP causes a conformational change in the importin that causes it to dissociate from its cargo
Cohesins help the cell remember the gene expression pattern that existed before the cell divided by marking transcription factor binding, helping the transcription factors find their correct places. True or false?
True. direct quote from his slide: "COHESINS HELP THE CELL REMEMBER THE ORDER THAT EXISTED BEFORE THE CELL DIVIDED BY MARKING TRANSCRIPTION FACTOR BINDING, HELPING THE TRANSCRIPTION FACTORS FIND THEIR CORRECT PLACES."
Each kinetochore fiber exerts a poleward-pulling force that is proportional to its length (until the pull forces from each pole are equal). True or false?
True. Longer microtubules exert a stronger pulling force on the kinetochore. In this way, chromosomes are arranged in the mitotic plate where net tension on the kinetochores of the centromere is zero.
Chromosomes are pushed away from the poles (polar winds) by chromokinesins and spindle microtubule interactions. True or false?
True. Chromokinesins are located on the surface of the chromosome arms and interact with spindle microtubules. These are NOT the same microtubules interacting with the kinetochore. These chromokinesin/microtubule interactions push the chromosomal arms away from the pole. This force stops mono-oriented chromosomes from being pulled all the way to one pole
Stable, bipolar attachment of sister chromatids is assessed by kinetochore passenger proteins and the MCAK motor (that depolymerizes microtubules) so that the SAC is not passed too soon. True or false?
True. SAC activation inhibits the action of APC, the protein complex that degrades securin which inhibits separase which breaks the cohesins holding the sister chromatids together. SAC is activated when errors occur in the microtubule/kinetochore coupling process (p. 718-720, fig 15.38, 15.39).The MCAK destabilizes microtubules when the chromosome is under merolitic or syntelic conditions, freeing the kinetochore to bind with the correct microtubule.
Does cdc25 expression oscillate during cell cycle?
Yes it does. Because it is a phosphatase that activates CDK1/cyclin B, which is only expressed during certain cell cycle phases to promote mitosis. >>>> Yes. Cdc25 levels are higher during G1 and G2.
What is the phenotype of a cell that loses the function of Wee kinase, the inhibitory kinase of Cdk1?
You get a small cell. Because the cell is always replicating, there's no time for growth.
What are astral microtubule? What is their origin?
astral MTs = "RADIAL ARRAY OF SHORT, HIGHLY DYNAMIC MTs TO FORM AROUND EACH CENTROSOME" astral MTs originate at the centrosome and radiate out in all directions forming an aster! they orient the centrosomes within the cell which also orients the mitotic spindle they also move the centrosomes away from each other during anaphase B
What are the events of prophase?
basically, 1) accumulation in nucleus of Cdk1/cyclin B complex 2) Cdk1/cyclin B phosphorylates proteins 3) phosphorylated proteins cause chromosomes to condense AND the nuclear envelope to break apart 4) centrosomes then align at opposite poles 5) centrosomes then send out MTs and create mitotic spindle During prophase, Cdk1/cyclin B accumulates in the nucleus. Phosphorylation of numerous proteins by Cdk1/cyclin B causes the chromosomes to condense and the nuclear envelope to fall apart. Once the nuclear envelope is gone, and the centrosomes are oriented at opposite poles of the cell, the centrosomes send out microtubules and form the mitotic spindle
How mRNA processing is linked to mRNA export from the nucleus?
basically, mRNA processing MUST be 100% COMPLETE before mRNA can be exported out of the nucleus !!!!! mRNA is processed near interchromosomal domains where the mRNA can move easily to the nuclear envelope for export from the nucleus. processed mRNAs then get packaged by proteins (hnRNPs thought to package mRNAs), forming a RNA-protein complex -- mRNAs can then be exported once they are in their RNA-protein complex form hnRNPs are proteins that help define sites of pre-mRNA processing. They are also thought to package mRNAs for export.
How are executioner caspases activated?
executioner zymogens are cleaved by initiator caspases, granzyme B, or already-activated executioner caspases (i.e. a positive feedback loop) >>> this cleavage results in a conformational change and the formation of the active site of that executioner caspase !!!!! >>>>>> also, initiator caspases are what usually regulates this cleavage -- aka they are responsible for activating executioner caspases.
Which features appear in cells undergoing apoptosis but not in cells undergoing necrosis?
in necrosis, the cell appears to SWELL and then lyse -- exposed cell parts will then ELICIT AN INFLAMMATORY RESPONSE (bc chromatin does NOT get condensed in necrosis) VS. apoptosis = cell is FRAGMENTED into multiple, ROUND, apoptotic bodies >> these fragmentations then get ENGULFED BY PHAGOCYTES
How is procaspase 9 induced?
initiator procaspase-9 is induced VIA INDUCED PROXIMITY when the CARD domain of procaspase-9 binds to the CARD domain of APAF-1.
What advantage does possession of a nucleus offer a cell? because of all the different things that get transported in and out of a nucleus (ions, RNAs, a variety of proteins, etc.). True or false?
pretty sure that this is FALSE bc "the NUCLEAR ENVELOPE contains pores used for transporting RNAs, proteins, and small molecules btwn the nucleus and cytoplasm" (so as long as we're considering the nuclear envelope to be a separate entity of the nucleus itself, then this statement is false.) the advantage of having a nucleus is that cell's have a place to store almost all of their genetic material and act as a center for controlling cellular activities
What is apoptosis?
programmed cell death FRAGMENTATION OF THE CELL and nucleus, blebbing of cell membrane "death is quick and clean with NO CELL SPILLAGE to cause an inflammatory response." cell receives EXTERNAL signals that initiate a cascade of protease activation events in the cytosol; the activated proteases digest the cell
what is the function of the anaphase-promoting complex?
regulation of sister chromatid separation. (when u think about it, makes sense bc this is literally what happens during anaphase) Anaphase Promoting Complex APC and cdc20 activator ubiquitinates securin. Separase is then freed from securin and cleaves cohesin at the centromere, allowing sister chromatids to separate.
What advantage does possession of a nucleus offer a cell?
ultimately allows a place for cells to store genetic material and process it; ultimately having a nucleus allows for the cell to highly control/regulate gene expression !!!! The DNA is protected from the shear generating forces of the actin. A nucleus makes more sophisticated/regulated gene replication and transcription possible
What happens when cytochrome C binds APAF-1?
when cytochrome C binds APAF-1, it causes APAF-1 to oligomerize and form the apoptosome which recruits procaspase 9 which is then activated.
How does centrosome reproduce?
"ABOUT THE TIME THE DNA IS REPLICATED, COMPOSED OF 2 CENTRIOLES SURROUNDED BY THE PERICENTRIOLAR MATERIAL. NEW CENTROSOMES REQUIRES DUPLICATION OF THE CENTRIOLES, CONTROLLED BY THE CELL CYCLE AND IS COORDINATED WITH DNA REPLICATION." >>> Centrosomes reproduce during the S phase about the time DNA is replicated. Cdk2/cyclin E takes it out and cyclin A act as regulators of centrosome reproduction and are active during S phase when both DNA and centrosomes are replicated.
What is the mitotic spindle?
"INTERACTIONS BTWN THE ASTERS, FORMED BY THE TWO CENTROSOMES, INITIATE THE FORMATION OF THE MITOTIC SPINDLE." The mitotic spindle is composed of the two centrosomes, the microtubules that extend out towards the chromosomes, and the motor proteins that generate pushing/pulling forces
Active genes are found adjacent to interchromosomal domains. True or false?
*Slide 5, highly transcribed genes located at the periphery of chromosomal domains near the inter-chromosomal domains and near pores.
List the Cdks-Cyclins complexes present at each phase of the cell cycle? On the same diagram show the cell cycle checkpoints.
G1- CDK4/cyclin d CDK2/cyclin e G1 checkpoint S- CDK2/cyclin a G2/M- CDK1/cyclin b G2 checkpoint
What is the definition of heterochromatin?
Highly condensed DNA, typically less active and more common, while euchromatin is less condensed, less common and more active. basically, heterochromatin = TIGHTLY bounded DNA that is NOT actively transcribed; DNA is most commonly found in this form
Know the cargo with the protein(s) necessary for nuclear export or import.
Import Importin proteins include monomeric karyopherin and a heterodimer called alpha/beta importin. Importins bind to cargo tagged with NLS and move through nuclear pore complexes into the nucleus. Once in the nucleus, Ran-GTP binds to importin which causes the importin to release the cargo. Ran-GTP and a exportin protein named CAS export the alpha and beta importins back into the cytoplasm. Back in the cytoplasm, Ran-GAP hydrolyzes the Ran-GTP creating Ran-GDP and the importins are released from the Ran protein. Export Ran-GTP allows exportins to bind to NES tagged cargo destined for export (fig 9.40). Crm 1 is an important exportin receptor protein (fig 9.36). Once the Ran-GTP, exportin, and cargo are all bound, the complex exits the nucleus and the GTP on Ran is hydrolyzed by Ran-GAP to form Ran-GDP and release the cargo.
The nuclear localization signal (NLS) is recognized by and binds to which protein in the process of nuclear protein transport?
Importin In other words, JUST KNOW THAT IMPORTIN PROTEINS ARE WHAT RECOGNIZE AND BIND TO NLS TAGS -- the binding of importin proteins to NLS tags is how nuclear proteins get IMPORTED into the nucleus
What are the features of the transport of proteins across the nuclear envelope?
Importins bind to proteins tagged with NLS >> once the importin binds, it can then transport the cargo across the nuclear envelope through nuclear pore complexes. NES-tagged cargo, exportin, and Ran GTP form a complex that can exit the nucleus.
A cell in quiescence is?
In a state of non-division.
Your textbook describes a simple experiment illustrating that export of transfer RNA (tRNA) from the nucleus to the cytoplasm is under the control of the small GTP binding protein Ran. Summarize the results of this experiment and explain how Ran is functioning in this process.
In the study, Ran-GAP was injected into the nucleus of a cell. Ran GAP is usually in the cytoplasm and hydrolyzes Ran-GTP to Ran-GDP. With Ran-GAP in the nucleus, all the Ran-GTP was converted to Ran-GDP and could not function as part of the exportin complex so tRNA remained in the nucleus.
Define the term "induced proximity", and give one example.
Induced proximity is when procaspases (aka the precursors for initiator caspases) are close to each other and dimerize because of their proximity. One example is during the death receptor pathway.
How are initiator caspases activated?
Initiator caspases are activated by dimerization. This mechanism of activation by dimerization is called induced proximity. Initiator caspases are activated by adaptor molecules that contain domains called death folds -- initiator caspases also have these same death fold domains in their prodomain. The activation of initiator caspases occurs when the adaptor's and caspases' death folds bind to one another, resulting in two caspase monomers dimerizing. >>>>Once initiated, the dimerized initiator caspase cleaves itself off and goes on to activate executioner caspases by cleaving them
Hypothesize the early and late outcome of a cell without a functional Retinoblastoma protein. How oncologist classify this protein?
Retinoblastoma (Rb) is a tumor suppressor protein that binds and inhibits the transcription factor E2F. E2F stimulates the genes that transcribe cyclin E which binds with Cdk2 and initiates S phase. When Rb is active, mitosis cannot start. If a cell did not have a functional Rb protein then E2F would not be inhibited and cyclin E levels would remain high and there would be no restrictions on a cell to enter S phase and begin DNA replication. Certain extracellular factors such as 'tumor necrosis factor-a' can detect the lack of Rb and trigger apoptosis in the cell. (pg 700) A late outcome would possibly just be referring to developing cancerous cells from lack of regulation.
Both U snRNAs and microRNAs are transcribed by polymerase II and then further modified including cleavage of the RNAs. What are RNAs?
Ribonucleic acid (RNA) also carry genetic information, much like DNA, but RNA typically acts as a messenger that carries the instructions for creating proteins.
The isolation of conditional cell cycle mutations in yeast helped identify many of the key proteins involved in cell cycle control. Many of these mutations were given the name cdc (cell division cycle) and are temperature sensitive. Hypothesize about the outcomes of A cdc temperature sensitive mutant.
Temperature sensitive mutants will experience arrest of the cycle when transferred to different temperature environment.
What is the complex found in the mitochondria that drives apoptosis?
The Bax/Bak complex oligomerizes and then gets inserted into the mitochondria's outer membrane >> MOMP then occurs >> and then cytochrome C is released from the intermembrane space >>> cytochrome C then binds APAF-1, which then initiates the very start of the activation cascades for initiator caspases
What is the functional role of CARD domains?
The CARD domains of APAF-1 and procaspase 9 bind and activate the procaspase 9 that allows it to cleave and activate executioner caspases.
The protein IκB functions in two independent and complimentary roles to control the import and export of the transcription factor NFκB. Explain these two roles.
The IkBα protein inhibits NFkB by masking the NLS, keeping them from entering the nucleus. IkB kinase phosphorylates IkBα, resulting in IkBα dissociating from NFkB, allowing NFkB to freely enter the nucleus.
MOMP, mitochondrial outer membrane permeabilization is a key event in apoptosis. What role do the apoptotic Bcl-2 proteins Bax and Bak play in MOMP?
The activation and function of bax and bak are inhibited by the antiapoptotic member of the bcl-2 family. These bind to the activator proteins and prevent their function and also bind to active forms of bax and bak in this way antiapoptotic bcl-2 family members prevent MOMP and prevent apoptosis. (page 738) BAX and BAK are two multidomain proteins in the bcl-2 family these two proteins are responsible for MOMP and probably form the pores through which the proteins of the mitochondrial intermembrane space diffuse. BAX or BAK are induced to oligomerize in the mitochondrial membrane in the presence of a BH3-only protein such as BID.
What are centromeres? What are their structure and roles?
The centromere is a specialized region on the chromosome that contains the kinetochores. They are condensed heterochromatin composed of highly repetitive DNA sequences that bind to a unique set of proteins
What is a complex of multimers of APAF-1 doing?
The complex of APAF-1 is called the apoptosome (composed of 7 APAF-1's) which binds and activates the initiator procaspase-9 via CARD domains
List the features of executioner caspases that distinguish them from other caspase types.
The executioner caspases (predominantly caspases-3 and -7 in vertebrates) are responsible for cleaving many different proteins to effect apoptosis. In mammalian cells the estimate is ~500 substrates for cleavage. In most cases of apoptosis, the cleavage and activation of these executioner caspases is mediated by the action of another set of caspases, the 'initiator' caspases.
