PCB3134 test 4

Nucleolus

region within the nucleus made up of proteins and nucleic acids; its general function is to transcribe ribosome RNA (rRNA) and combine it with proteins to form incomplete ribosomes

At low salt concentration, chromatin..

resembles beads on a string, in this extended form, the string is composed of free DNA called "linker" DNA connecting beadlike structures - the nucleosomes.

Chromosome

"chromatic nuclear figure" - (DNA + proteins) that carries part (or all) of the hereditary information of an organism.

MPF

(Maturation/Mitosis Promoting Factor) When ~5 percent of the cytoplasm from an unfertilized Xenopus egg arrested in metaphase of meiosis II is injected into a G2 arrested oocytes, the oocyte enters meiosis 1, and proceeds to metaphase of meiosis II, generating a mature egg in the absence of progesterone. This process can be repeated multiple times without further addition of progesterone, showing that egg cytoplasm contains an oocyte maturation-promoting factor (MPF). Microinjection of G2 arrested oocytes provided the first assay for MPF activity.

Anaphase B

(spindle elongation): The second part of anaphase involves separation of the spindle poles in a process known as anaphase B. Anaphase B separates poles by the combined action of kinesins and dynein. A major contributor to this movement is the involvement of the bipolar kinesin-5 proteins. Anapahse B is driven in part by the bipolar kinesin-5 ( in a sliding filament mechanism) and the extent of the spindle elongation being driven by microtubule polymerization.

Cytokinesis

reformation of interphase microtubule array contractile ring forms cleavage furrow

Most destructive cancers exhibit six cellular properties:

-Sustaining proliferative signaling -Evading growth suppressors -Activating invasion and metastasis -Enabling replicative immortality -Inducing angiogenesis -Resisting cell death While less destructive cancers often display a subset of the six properties. These properties are inheritable by further generations of cells, this results in carcinogenesis.

Pluripotent stem cells:

-extensive but limited capability - can develop into most types of cells in an organism -the human ES cells of interest are pluripotent

Totipotent stem cells:

-unlimited capability to develop into all extraembryonic, embryonic, adult types of cells

Three classes of Microtubules:

- Astral - Kinetochore - Polar/overlap all their (-) ends are embedded. Note: All the microtubules in each half of the symmetrical spindle have the same orientation except for some polar microtubules, which extend beyond the midpoint and interdigitate with polar microtubules from the opposite pole. •Plus ends project away from the centrosomes • Motor proteins operate at or near the ends of microtubules • K-fibers - chromosome movement; astral - spindle positioning; anchoring for anaphase B; overlap - spindle shape; anaphase B elongation

Germ Cells and Fertilization: Meiosis I:

- Homologs pair up and synapsis occurs - The homologs separate - The sister chromatids stay together 1. In G2 and prophase of meiosis I, the two replicated chromatids of each chromosome are associated with each other by cohesin complexes along the full length of the chromosome arms, just as they are following DNA replication in a mitotic cell cycle. A major difference between meiosis and mitosis is that in prophase of meiosis I, homologous chromosomes (maternal, paternal) pair with each other a process called synapsis. (Synapsis is how homologs find one another: Synapsis takes place during prophase I of meiosis. When homologous chromosomes synapse, their ends are first attached to the nuclear envelope. These end-membrane complexes then migrate, assisted by the extranuclear cytoskeleton, until matching ends have been paired) 2. Another key difference between mitosis and meiosis is that in the metaphase of meiosis I, the kinetochores at the centromeres of sister chromatids attach to spindle fibers emanating from the same spindle pole, rather than from opposite spindle poles as in mitosis. However, the kinetochores of the maternal and paternal chromosomes of each tetrad attach to spindle microtubules from opposite spindle poles. 3. During anaphase of meiosis I, securin degradation releases separase, which then cleaves the cohesin rings holding the chromosome arms together as during mitosis. However, during meiosis I, cohesin rings at the centromere are not cleaved. This allows the recombined maternal and paternal chromosomes to separate, but each pair of chromatids remains associated at the centromere.

Meiosis II:

- Sister chromatids separate in Meiosis II - Meiosis does not cycle - There is no DNA synthesis between MI and MII - Genome size is reduced by exactly one half after MII 4. Cytokinesis yields two cells, which enter meiosis II without undergoing DNA replication . At metaphase of meiosis II the chromatids composing each replicated chromosome associate with spindle microtubules from opposite spindle poles, as they do in mitosis. 5 & 6: Segregation of chromatids to opposite spindle poles during the second meiotic anaphase followed by cytokinesis generates haploid germ cells containing one copy of each chromosome.

Mitosis vs Meiosis:

- Two rounds of cell division follow one round of genome duplication - Two step removal of cohesin during meiosis - Cohesin at the centromere is protected by Shugoshin - the guardian spirit - MI

Chromatin -

- nucleoprotein complex (the organization of nucleosomes) - the complex of histones, nonhistone proteins, and DNA constitutes chromatin - dispersed throughout much of the nucleus in interphase cells

Histone -

- the most abundant protein in chromatin - There are five major types of histone proteins - H1, H2A, H2B, H3, and H4 - and are rich in positively charged phosphate groups in DNA

Anaphase

APC/C activated and cohesions degraded Anaphase A: Chromosome movement to the poles Anaphase B: Spindle pole separation

Anaphase A

Anaphase A moves chromosomes to poles. This movement can be powered by microtubule shortening, utilizing the stored structural strain released by removing the GTP cap.

CDK1

Cyclin: A and B Function: Mitosis General Name: Mitotic CDKs

Prophase

Breakdown of interphase microtubule display and its replacement by mitotic asters Mitotic aster separation Chromosome condensation

CDK2

Cyclin: A and E Function: Entry into cell cycle S phase General Name: G1/S phase CDKs

CDKs

CDK's (cyclin dependent kinase) are proteins that regulate the cell cycle. As told by their name, CDK have little kinase activity without the presence of a cyclin ( a regulatory protein). CDK's bind to a cyclin, and only upon forming a cyclin-CDK complex is there active kinase activity. In the image below, you can see that when CDK is bound to an M-Cyclin (Mitosis) it activates mitosis machinery. The cyclin is then broken down and an S-Cyclin (synthesis) is attached at the beginning of the S-Phase of the Cell cycle, this cyclin triggers DNA replication machinery within the cell. - Phosphorylation of the CDK subunit regulates the kinase activity of MPF - Conformational changes induced by cyclin and binding and phosphorylation increase MPR activity.

Regulation of S-Phase cyclin CDK

CDKinases are required for initiation of DNA synthesis, they are called early S-Phase Cyclins. When a substrate called Cdh1 is inactivated, this allows the S-phase cyclin-CDK complexes to accumulate in late G1. As the S-phase cyclin-CDK heterodimers (the cyclin-CDK complexes) accumulate they are immediately inactivated by binding of an inhibitor, called Sic1, that is expressed late in mitosis and early in G1. Sic1 functions as an S-Phase inhibitor (it does not inhibit G1 cylcin-CDK complexes). Entry into the S phase is defined by the initiation of DNA replication. This occurs when the Sic1 inhibitor is degraded following polyubitiquination (see above for explanation of this process) by the ubiquitin-protein ligase called SCF. Once Sic1 is degraded, the S-phase cyclin-CDK complexes induce DNA replication by phosphorylating several proteins in pre-replication complexes bound to replication origins.

Centrosome Duplication in Interphase

Centrosomes separate before the nuclear envelope breakdown. Cyclin-E - CDK2 controls centrosome duplication. In order to separate the chromosomes at mitosis, cells duplicate their centrosomes, coordinately with the duplication of their chromosomes in S phase. Centrosomes must be closely monitored, many tumors contain more than two centrosomes. after the pair of parent centrioles(green) separate slightly, a daughter centriole(blue) buds from each and elongates. By G2 growth of the daughter centriole buds from each and elongates. By G2, growth of the daughter centrioles is complete, but the two pairs remain within a single centrosomal complex. Early in mitosis, the centrosome splits, and each centriole pair migrates to opposite sides of the nucleus. The amount of pericentriolar material and the activity to nucleate microtubule assembly increases greatly in mitosis. In mitosis, these centrosomes are called spindle poles.

CDK4 and CDK6

Cyclin: D Function: G1, entry into cell cycle General Name: G1 CDKs

Interphase

Chromosome duplication and cohesion Centrosome duplication

what is chromosome painting?

Chromosomes are located within the interphase nucleus in regions called domains. Using fluorescent tagging, each chromosome is "painted", a pair of homologous chromosomes can then be visualized as two discrete domains and their relative spatial location determined.

Nuclear Lamins

Class: V (five) Protein: Lamins Distribution: Nucleus Proposed Function: Nuclear Structure and Organization - Nuclear lamins, form a structural support network under the nuclear envelope - The nuclear envelope breaks down during mitosis when lamin is phosphorylated

Chromosome Structure:

Cohesin and condensin help organize chromosome structure. Although distinct in structure and function, cohesin and condensin both contain conserved chromosomal ATPases of the structural maintenance of chromosomes (SMC) protein family. Present models indicate that cohesin might encircle two sister chromatids to promote their cohesion, whereas condensin might stabilize supercoiled DNA loops on a single chromatid to promote its condensation. Whether the diversity of functions exhibited by each complex during interphase and mitosis reflects a common molecular mechanism remains unknown. An SMC protein complex consists of two monomers, SMC2 (blue) and SMC4 (red), whose hinge domains associate. The head domains, which have ATPase activity are linked by a kleisin protein, forming a ringlike structure. (b) The ringlike SMC complex topologically links two chromatin fibers (gray cylinders). The cylinder diameter represents the diameter of a nucleosome and is to scale relative to the dimensions of the SMC complex. (c.) Loops of transcriptionally active chromatin may be tethered at their base by several SMC complexes forming a topological knot.

Cohesin

Cohesin is a protein complex that regulates the separation of sister chromatids during cell division (mitosis and meiosis). During mitosis, sister chromatids generated by DNA replication in S phase are intially linked by cohesin complexes along the length of the chromatids. During chromosome separation the complexes become restricted to the region of the centromere at metaphase. Specific proteins recruit PP2A (an enzyme) to centromeres, where they antagonize other proteins, preventing the dissociation of cohesins from centromeric regions. Activation of separase leads to removal of cohesins at the centromere. Sister chromatids now separate, marking the onset of anaphase. Therefore the activation of separase triggers the metaphase to anaphase transition

Nucleus

Compartmentalization: •Nuclear envelope •Nucleolus •Chromatin Subnuclear Bodies: •Nucleolus - sub organelle inside the nucleus; produces ribosomes and is not bound by membrane ◦rRNA precursor → ribonucleoprotein particle → nucleolus •Cajal body - represented by coilin staining (see review at bottom) •Speckles - pre-mRNA splicing Areas of transcriptional silence are often associated with the nuclear envelope

Cyclin degradation

Cyclin degradation is equally important for progression through the cell cycle. Specific enzymes break down cyclins at defined times in the cell cycle. When cyclin levels decrease, the corresponding CDKs become inactive. In addition, cell cycle arrest can occur if cyclins fail to degrade.

Cytokinesis

During late anaphase and telophase in animal cells, the cell assembles a microfilament-based contractile ring attached to the plasma membrane that will eventually contract and pinch the cell into two, a process known as cytokinesis.

Structure of Cytoplasmic Dynein

Dyneins consist of six subunits, two large, two medium, two small. There are two "heavy chains" that contain the ATPase activity (in the doughnut shaped head), and so responsible for generating the energy for movement along the microtubule. The "motor" is minus end directed. The intermediate chains are believed to anchor dynein to its cargo. The two projections from the head are believed to bind the dynein to other cytoplasmic structures. One of the projections (the stalk) binds to microtubules and "walks" along their surface via bonding and unbonding.

Cell Cycle:

Four Stages of the Cell Cycle: G1 - S - G2 - M Gap1 - S Phase - G2 - Mitosis Cells grow in size and synthesize RNA's and proteins required for DNA synthesis. Once they have reached a certain point, they enter the cell cycle by traversing a point in G1 (known as start) Once this point has been crossed, cells are committed to division. The first step toward successful division is the S (synthesis) phase, the phase where cells actively replicate their chromosomes. After progressing through a second gap phase (G2) , the cells begin the mitotic process. Conservation of Cell Cycle Machinery is similar in all eukaryotes

Histones

Histone tails extend from the core particle - H2A; H2B; H3 and H4 Tails are from N-termini of the Histones; H2A and H2B have additional C-terminal tails Each of the histone proteins making up the nucleosome core contains a flexible N - terminus of varying amounts of residues extending from the globular structure of the nucleosome; the H2A and H2B proteins also contain a flexible C-Terminus extending from the globular histone octameric core. These termini, called histone tails, are required for chromatin to condense from the beads-on-a-string conformation into the 30-nm fiber. Histone tails are subject to multiple post-translational modification such as acetylation, methylation, phosphorylation and ubiquitination. A particular histone protein never has all of these transformations simultaneously, but the histones in a single nucleosome may collectively contain several different types of modifications. The combination of modifications found in different regions of chromatin have been suggested to constitute a histone code that influences chromatin function by creating or removing binding sites for chromatin associated proteins.

p53

In a normal cell p53 is inactivated by its negative regulator, mdm2. Upon DNA damage or other stresses, various pathways will lead to the dissociation of the p53 and mdm2 complex. Once activated, p53 will induce a cell cycle arrest to allow either repair and survival of the cell or apoptosis to discard the damaged cell. How p53 makes this choice is currently unknown. Activated by hyperproliferative signals, DNA damage, telomere shortening, hypoxia.

How do K-Fibers (kinetochore microtubules) attach? Step 3

In this orientation the unoccupied kinetochore on the opposite side is pointing toward the distal spindle pole, and eventually a microtubule from the distal pole will capture the free kinetochore (the chromosome pair is now said to be bi-oriented. With the two kinetochores attached to opposite poles, the duplicated chromosome is now under tension, being pulled in both directions. Simplified: The bi-oriented chromosomes then move to a central point between the spindle poles in a process known as chromosome congression.

Cancer cells - connection to cell cycle regulation; loss of heterozygosity and tumorigenesis

It is a common occurrence in cancer, where it indicates the absence of a functional tumor suppressor gene in the lost region. However, many people remain healthy with such a loss, because there still is one functional gene left on the other chromosome of the chromosome pair. However, the remaining copy of the tumor suppressor gene can be inactivated by a point mutation (mutation on a single gene), leaving no tumor suppressor gene to protect the body. Loss of heterozygosity does not imply a reversal to the homozygous state. The classical example of such a loss of protecting genes is hereditary retinoblastoma, in which one parent's contribution of the tumor suppressor Rb1 is flawed. Although most cells will have a functional second copy, chance loss of heterozygosity events in individual cells almost invariably lead to the development of this retinal cancer in the young child.

How do K-Fibers (kinetochore microtubules) attach? Step 1

Kinetochores attach to microtubules in prometaphase. Step 1a/1b : Microtubules nucleated from the spindle poles are very dynamic, and when they contact the kinetochore, either laterally or at their end, this can lead to chromosome attachment In the first stage of prometaphase, chromosomes become attached either to the end (1a) of a microtubule, or to the side of one (1b)

Loss of function

Loss of function mutations in tumor suppressors encode proteins with permanently shut off inhibitor activity, such as lack of a brake (which can cause retinoblastoma) or checkpoint control (lack of a p53).

An MPF is a mitosis promoting factor

MPF promotes the entrance into mitosis (the M phase) from the G2 phase by phosphorylating multiple proteins needed during mitosis. MPF is activated at the end of G2 by a phosphatase, which removes an inhibitory phosphate group added earlier. The MPF is also called the M phase kinase because of its ability to phosphorylate target proteins at a specific point in the cell cycle and thus control their ability to function. MPF must be activated in order for the cell to transition from G2 to M phase. There are three amino acid residues responsible for this G2 to M phase transition. The Threonine-161 (Thr-161) on CDK1 must be phosphorylated by a Cyclin Activating Kinase (CAK). CAK only phosphorylates Thr-161 when cyclin B is attached to CDK1

Nonhistone proteins / Chromatin loops -

Nonhistone proteins compose a chromosome scaffold, in general SARs (scaffold associated regions) are found between transcription units, and genes are located primarily within the chromatin loops. The loops are tethered at their bases by a mechanism that does not break the duplex DNA molecule, which extends the entire length of the chromosome. Evidence suggests that SARs may affect transcription of neighboring genes.

Prometaphase

Nuclear envelope breakdown Chromosomes captured, bioriented and brought to the spindle equator

Chromosome Organization:

Nucleosome - - the basic unit of DNA packaging in eukaryotes - the building block of chromatin - packs and organizes chromosomal DNA - nucleosomes consist of a protein core, with thread around its surface (like thread around a spool) - The core is an octamer containing two copies each of histones H2A, H2B, H3, and H4. - Nucleosomes from all eukaryotes contain 147 base pairs of DNA wrapped one and two thirds turns around the protein core. There are roughly 200 base pairs in total

Kinesin 7 - CENP - E

On the side with lengthening microtubules, a CENP-E/kinesin-7 protein holds on to the growing microtubule.

How do K-Fibers (kinetochore microtubules) attach? Step 2

Once attached, the motor protein dynein/dynactin (located at the kinetochore) moves the chromosome pair down the microtubule toward the spindle pole. Simplified: (2) The duplicated chromosome is then drawn toward the spindle pole by kinetochore-associated dynein/dynactin as this motor moves toward the ( - ) end of a microtubule.

Proteosome mediated Proteolysis

Proteolysis is a breakdown of proteins into smaller polypeptides or amino acids, this is generally achieved by hydrolysis that is triggered by enzymes called protease. The degradation of the cyclins, the proteins described in the previous section, is the key step that governs the exit from mitosis and progress into the next cell cycle.] Cyclins accumulate in the course the cell cycle, then abruptly disappear just before the anaphase of mitosis. These cyclins " disappear" due to an ubiquitin-mediated proteolytic pathway.

Protofilament Arrangement in Microtubules

Protofilaments are the building blocks of the microtubule structure. Thirteen protofilaments associate to create a single cytoplasmic microtubule, that can then extend with the addition of more protofilaments. Cilia/Flagella (doublet MT's) - 13 + 10 protofilament walls Primary function of Cilia and Flagella are environmental cell motility. Centriole/Basal Body (triplet MT's) 13+10 + 10 protofilament walls

DNA-damage checkpoint

Purpose: Detects damage to DNA throughout the cell cycle Sensor: ATM, ATR detect DNA damage Action: Inhibition of Cdc25A to prevent entry into S phase, P21 inhibition of all cyclin-CDK complexes to induce cell cycle arrest

Intra-S phase Checkpoint

Purpose: Ensure all DNA replication is complete before entering M-phase Sensor: ATR detects replication forks Action: Inhibition of Cdc25C to prevent activation of mitotic cyclin-CDKs, blocking early mitotic events

Spindle-assembly checkpoint

Purpose: Ensures all chromosome kinetochores are attached to spindle microtubules before anaphase Sensor: Mad2 detects kinetochores unattached to microtubules Action: Inhibition of Cdc20 to prevent activation of separase and onset of anaphase Mad2 and other proteins inhibit activation of the APC/C specificity factor Cdc20 required for polyubiquitination of securin, thereby preventing entry into anaphase. *Once entering anaphase, this is considered a point of no return.

Spindle-position checkpoint

Purpose: Ensures all chromosomes are properly segregated to daughter cells before telophaes and cytokinesis Sensor:Tem-1 detects proper position of spindle pole body. Action: prevention of Cdc14 activation and degradation of mitotic cylins, blocking late mitotic events.

Retinoblastoma

Retinoblastoma (Rb) is a rapidly developing cancer that develops from the immature cells of a retina, the light-detecting tissue of the eye and is the most common malignant tumor of the eye in children . The hereditary propensity for Retinoblastoma is a result of the inheritance of a defective Rb gene. The tumor can also be caused due to sporadic mutation, mutating both RB genes, causing an onset in adulthood.

Dynein Powerstroke

Rotation of the head leads to the generation of the power stroke, a process that is not completely understood. The protein dynactin (also consists of multiple subunits - 12) aids in intracellular transport by binding to dynein (and kinesin) and linking them to an object needing transport (organelle or vesicle).

The function of separase:

Separase also known as separin is a cysteine protease responsible for triggering anaphase by hydrolysing cohesin which is the protein responsible for binding sister chromatids during metaphase.

Pachytene

Stage 3 of prophase 1. the stage when chromosomal crossover (crossing over) occurs. Nonsister chromatids of homologous chromosomes may exchange segments over regions of homology. Sex chromosomes, however, are not wholly identical, and only exchange information over a small region of homology. At the sites where exchange happens, chiasmata form. The exchange of information between the non-sister chromatids results in a recombination of information; each chromosome has the complete set of information it had before, and there are no gaps formed as a result of the process

Diplotene

Stage 4 of prophase 1. homologous chromosomes separate from one another a little. The chromosomes themselves uncoil a bit, allowing some transcription of DNA. However, the homologous chromosomes of each bivalent remain tightly bound at chiasmata, the regions where crossing-over occurred. The chiasmata remain on the chromosomes until they are severed in anaphase I.

Diakinesis

Stage 5 of prophase 1. This is the first point in meiosis where the four parts of the tetrads are actually visible. Sites of crossing over entangle together, effectively overlapping, making chiasmata clearly visible. Other than this observation, the rest of the stage closely resembles prometaphase of mitosis; the nucleoli disappear, the nuclear membrane disintegrates into vesicles, and the meiotic spindle begins to form.

Leptotene

Stage one of prophase 1. In this stage of prophase I, individual chromosomes—each consisting of two sister chromatids—change from the diffuse state they exist in during the cell's period of growth and gene expression, and condense into visible strands within the nucleus. However the two sister chromatids are still so tightly bound that they are indistinguishable from one another.

Zygotene

Stage two of prophase 1. the chromosomes approximately line up with each other into homologous chromosome pairs. This is called the bouquet stage because of the way the telomeres cluster at one end of the nucleus. At this stage, the synapsis (pairing/coming together) of homologous chromosomes takes place. Pairing is brought about in a zipper-like fashion and may start at the centromere (procentric), at the chromosome ends (proterminal), or at any other portion (intermediate). Individuals of a pair are equal in length and in position of the centromere.

• What is APC? How is APC regulated during mitosis?

The APC/C's main function is to trigger the transition from metaphase to anaphase by tagging specific proteins for degradation. The two proteins of most importance that get degraded in this process as substrates of the APC/C are securin (inhibits separase activity) and S and M cyclins.

The wait signal

The Spindle Assembly checkpoint senses microtubule attachment and tension. Unattached kinetochores will generate a "wait" signal. This signal inhibits Cdc20 an activator of APC (anaphase promoting complex). Mad2 and Cdc20, two candidate proteins for components of a diffusible wait signal, have previously been shown to be recruited to and rapidly released from unattached kinetochores. Mad1, Bub1, and a portion of Mad2, all essential mitotic-checkpoint components, are stably bound elements of unattached kinetochores. After microtubule attachment, Mad1 and Mad2 are released from kinetochores and relocalize to spindle poles, whereas Bub1 remains at kinetochores. A long residence time at kinetochores identifies Bub1, Mad1, and a portion of Mad2 as part of a catalytic platform that recruits, activates, and releases a diffusible wait signal that is partly composed of the rapidly exchanging portion of Mad2.

The Cell Cycle Control System

The cell cycle control system is a protein kinase machine. Cyclin dependent kinase (CDK) concentration is constant, but it's kinase activity fluctuates. The three classes of cyclin-cdks occur in: G1, S, M.

Centromere:

The primary constriction on the chromosome

CDK

The stages in the cell cycle are controlled by cyclins. When these cyclins are bound with a CDK they can phosphorylate a protein. These cyclin-CDK combinations can turn on, or turn off a protein, at specific places in order to regulate the steps in the cell cycle. Therefore the cyclin-CDK combo makes sure the cell cycle runs properly.

Intermediate Filament

There are 5 classes of intermediate proteins. - the core region is structurally conserved - each type of intermediate protein has distinctive ends, not conserved across classes IF's have nonpolar structure Monomer •long alpha-helical central region Coiled-coil dimer Anti-parallel tetramer •no polarity in building block •associate end-to-end into protofilament Protofilaments form protofibrils. Protofibrils form filament - no intrinsic polarity

Kinetochore microtubules

To attach to microtubules, each chromosome has a specialized structure called a kinetochore. The kinetochore is located at the centromere (the constricted region of the condensed chromosome). Kinetochores contain many protein complexes to link the centromeric DNA eventually to microtubules. In animal cells, the kinetochore consists of an inner centromere and inner and outer kinetochore layers, with the positive (+) ends of the kinetochore microtubules terminating in the outer layer. Yeast kinetochores are attached by a singel microtubule to their pole, human kinetochores are attached by about 30 and plant chromosomes by hundreds. Kinetochores become attached to microtubules in prometaphase. When the microtubules nucleated from the spindle poles are very dynamic, and when they contact the kinetochore, either laterally or at their end, this can lead to chromosomal attachment. Microtubules "captured" by kinetochores are selectively stabilized by reducing the level of catastrophes, thereby promoting the chance that the attachment will persist.

Tumor Suppressor Proteins

Tumor Suppressor Proteins normally inhibit event cell cycle events in potentially tumorous cells until a feedback control allows the cell to continue through a checkpoint.

P87 Ubiquitin

Ubiquitin is a small regulatory protein that is found in almost all eukaryote tissue, while ubitiquination is a post translational modification (addition of a protein after it has been made) achieved via attaching ubiquitin to a substrate protein. Ubiquitination is carried out in three main steps; activation, conjugation, and ligation. These steps are tied to specific enzymes:. E1 - ubiquitin activating enzyme E2 - ubiquitin conjugating enzyme E3 - ubiquitin ligase (ex. APC - anaphase promoting complex)

Cancer:

a family of diseases characterized by uncontrolled cell proliferation (growth and division) •Cancer can result from transformation of a single cell in any tissue of the body Most cancers spawn from a single abnormal cell. Dividing cancer cells can generate benign tumors, growths that are confined. Cancers could also metastasize (movement to a different body part).

Chromatid

a single copy of a duplicated chromosome (generally bound to another copy via a centromere)

Chromosome

a single piece of coiled DNA containing several genes. A chromosome consists of organized protein, DNA, and RNA.

Cajal body

believed to modify RNA after it has been transcribed from DNA; located in the nucleolus and bound to it by a coilin protein. CBs participate in the storage and maturation of both snRNPs and small nucleolar RNAs, as well as other splicing factors necessary for mRNA and pre-rRNA processing, but they do not directly participate in transcription.

Conservation of cell cycle regulation: Cdc28=Cdc2=CdK1

cdc28 and cdc2 are genes that encode for CdK1. CdK1 is a kinase that plays an important role in cell cycle regulation. Phosphorylation of the kinase (only when bound to its cyclin) leads to cell cycle progression.

cdc mutant

cell division cycle mutant cdc's help to maintain the cell cycle, if one is malfunctioning the cell cycle may arrest. Cdc's often cannot function at high temperatures. At the higher temperature, the cells continue through the cell cycle until they reach the point where the function of the mutant gene is required for further progress, and at this point they halt

Metaphase

chromosomes aligned at metaphase plate

Polar/overlap

extend from each spindle pole body toward the opposite one and interact together in an antiparallel manner; these are called polar microtubules. These microtubules are responsible initially for pushing the duplicated centrosomes apart during prophase, then for maintaining the structure of the spindle, and then for pushing the spindle poles apart in anaphase B.

Astral

extend from the spindle poles to the cell cortex. By interacting, with the cortex, the astral microtubules perform the critical function of orienting the spindle with the axis of cell division

Kinetochore

link the spindle poles to the kinetochores on the chromosomes. This set of tubules first finds the chromosomes, then attaches them through the two kinetochores to both spindle poles and at anaphase A transports them to the poles.

Chromatin

made of histone Glycosylation of histone (H1) may promote chromatin condensation and compaction. Phosphorylation during interphase has been shown to decrease H1 affinity for chromatin and may promote chromatin decondensation and active transcription. However, during mitosis phosphorylation has been shown to increase the affinity of H1 for chromosomes and therefore promote mitotic chromosome condensation Heterochromatin is a tightly packed form of DNA, which comes in different varieties

Kinesin 13 (MCAK)

microtubule disassembly Congression involves bidirectional oscillations of chromosomes, with one set of kinetochore microtubules shortening on one side and the other set lenghtening on the other. On the shortening side, a kinesin-13 protein stimulates microtubule disassembly and a dynein/dynactin complex moves the chromosome toward the pole.

Kinesin 4 (chromokinesin)

moves toward the plus ends During the step where the chromosomes are bioriented, the chromosome arm points away from the closest spindle pole: this is due to chromokinesin/kinesin 4 motors on the chromosome arms moving toward the ( + ) ends of the polar microtubules.

Telophase

nuclear envelope reassembly assembly of the contractile ring

ATM

s activated and triggers three pathways that lead to an arrested G1 state: 1. Chk2 is phosphorylated and, in turn, phosphorylates Cdc25A, thereby marking it for degradation and blocking its role in CDK2 activation. 2. In a second pathway, phosphorylation of p53 stabilizes it, permitting p53 activated expression of genes encoding proteins that cause arrest in G1 and in some cases G2, promote apoptosis, or participate in DNA repair. 3. The third pathway is another way of controlling the pool of p53. The Mdm2 protein in its active form can form a complex with p53, causing p53 ubitiquination and subsequent degradation by the proteasome. ATM phosphorylates Mdm2 to inactivate it, causing increased stabilzation of p53. In addition Mdm2 levels are controlled by pArf14, which binds to Mdm2 and sequesters it in the nucleolus, where it cannot access p53. The human Mdm2 gene is frequently amplified in sarcomas, which presumably causes excessive inactivation of p53.

Cohesin: In Meiosis

•Cohesin - molecular glue between sister chromatids •Loaded onto chromosome at S-phase •Removed at M-phase

Somatic Cell Nuclear Transfer and Cloning:

•Inner mass cells can be removed from human blastocyst stage embryos and cultured in dishes •cells reproduce and maintain pluripotency (potential to develop into many different types of cells) •controversy: destroys "excess" human embryo (life?) that would have been discarded anyway One major difference between adult and embryonic stem cells is their different abilities in the number and type of differentiated cell types they can become. Embryonic stem cells can become all cell types of the body because they are pluripotent. Adult stem cells are thought to be limited to differentiating into different cell types of their tissue of origin.

Stem Cells

•Self renewal during cell division; can differentiate into specialized type of cells - heart, lung, skin, etc. •Development of pluripotent human stem cell lines (cultures) has brought biomedical research to the edge of a new frontier - Cell therapy •Origin of human embryonic stem cells has opened intense and passionate ethical debate about research and use of stem cells - highly politicized •Embryonic stem cells vs. adult stem cells

Normal Human Development:

•Sperm fertilizes egg to form first stem cell •First stem cell - Totipotent - can development into any type of cell in human body and placenta •In Vitro Fertilization (IVF) clinics fertilize human eggs with human sperm in culture dishes •blastocyst - cells specialize - pluripotent stem cells: -inner mass cells - potential to develop into any cell of fetus (human body) but not placental cells -outer layer cells - develop into placenta and supporting tissue but not fetus

Hierarchy of stem cells

•Totipotent cells divide - yield pluripotent stem cells •Pluripotent stem cells divide to yield other multipotent stem cells, e.g. blood stem cells, and specialized committed (unipotent) stem cells •Unipotent 'committed' stem cells divide and yield specialized 'differentiated' cells