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BIO 201 Cell Biology with Todd Hennessey Week Eleven Notes

by: ChiWai Fan

BIO 201 Cell Biology with Todd Hennessey Week Eleven Notes BIO 201

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ChiWai Fan
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Class Notes
BIO201, Cell, Biology, BIO 201 Cell Biology Todd Hennessey
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This 25 page Class Notes was uploaded by ChiWai Fan on Saturday April 16, 2016. The Class Notes belongs to BIO 201 at University at Buffalo taught by TODD HENNESSEY in Spring2015. Since its upload, it has received 80 views. For similar materials see CELL BIOLOGY in Biology at University at Buffalo.


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Date Created: 04/16/16
Cell Bio on April 11, 13, 15, 2016 (All images taken from Professor Hennessey’s slide—edited by ChiWai Fan) In Prometaphase: Three types of cytoplasmic MTs in the mitotic spindle: 1. Astral MTs: Eminate from centrosome into cytoplasm to anchor and position the aster (mitotic spindle) 2. Chromosomal MTs: Connect from pericentriolar material of centrosome to kinetochores 3. Polar MTs: Extend from centrosome to equator but interact with other polar MTs instead of chromosomes What’s the point: Cytoplasmic MT is dynamic and always growing and shrinking (Dynamic instability). Mitotic spindle can disappear in low temperature if they keep shrinking. Two of the main forces for movements in prometaphase You have mitotic chromosome, we want them to connect and move around by dancing around until they find their position. 1. Polymerization (longer) and depolymerization (shorter) of tubulin to lengthen and shorten MTs. If you shorten the MT, it will pull towards pole of mitotic spindle. It heads away the pole by making MT longer. You do this to effect the movement. 2. “Cargo and rail” action of motor proteins on MTs to position mitotic chromosomes and poles. Dyneins and kinesins. 1. Tubulin polymerization and depolymerization and mitotic chromosome movements in Prometaphase You can only add to plus end to make longer and you can only make shorter by taking away from the minus end (Old thinking) now we know you can either subtract or add to each end. They’re growing and shrinking which provides force for movement (does not need ATP) it is not ATP dependent. We are using the dynamic instability (lengthen and shorten) for force of movement.  What does this do? Lengthen and shorten chromosomal spindle fibers  What would happen to the length if you stopped growth of these MTs? You can add inhibitor that inhibits MT polymerization, this will stop growth and cell will stop dividing. FYI: A depolymerase helps to shorten the chromosomal MTs at each end during prometaphase  Can chromosomal MTs shorten at both the plus and minus ends? YES Dam ring prevents things from falling off. Growth polymerization at either end!! Not just plus end anymore. 2. Motor proteins position the mitotic chromosomes and the poles Which phase is this? Prometaphase: made connection and in the process of lining up towards the middle of mitotic spindle. We lost nuclear membrane when we came to prometaphase. Prophase: no connection Remember, Kinesin is a plus-end directed motor. It moves the cargo towards the plus end 1. Polar spindle fiber MT sliding by Kinesin causes the poles to move apart; positioning the poles to make the mitotic spindle. 2. Dynein-based movement moves mitotic chromosomes toward the minus ends of chromosomal spindle fibers (going away from centrosome) 3. Kinesin-based movement moves mitotic chromosomes toward the plus ends of chromosomal spindle fibers.  Chromosomal spindle fiber want to make contact with kinetochores, polar spindle fibers want to make contact with other polar spindle fibers.  Chromosomal MT help position mitotic chromosomes; Polar spindle fiber help to position the poles  (Polar spindle fiber can’t actually connect to kinetochores but chromosomal spindle fiber can!)  Dynein goes towards centrosome. Kinesin goes away from centrosome  All MT in mitotic spindle connect to a mitotic chromosome: FALSE!!! There’s other stuff. FYI: Tubulin modifications a lot of different type of modification for tubulins Metaphase Mitotic chromosomes align in amphitelic orientation at the metaphase plate. Amphitelic means that each one of the two sister chromatids face opposite poles This is in the middle (equator) of the mitotic spindle. Colchicine and colcemid (inhibitors of MT polymerization) produces metaphase chromosomes because it stops them from being pulled apart in anaphase B.  How can you tell you’re in metaphase? You look at it...above  How many sister chromatids in metaphase of humans? 92!  Is this a eukaryotic cell? Eukaryotic=true nucleus. NO we have no nucleus (DNA surrounding by membrane). Not all eukaryotic cell have nucleus. Spindle Checkpoint assures that all mitotic chromosomes reach the spindle equator before anaphase begins o Tubulin (green) o Mitotic Chromosomes (blue) o Spindle checkpoint protein (pink) Note that the one mitotic chromosome that isn’t aligned has the spindle checkpoint protein (pink) bound to it. It starts cell cycle arrest. Remove checkpoint protein until pink has made it in. Have to wait for pink before we divide to prevent abnormal distribution of chromosomes. This won’t cause cancer but it may be a contributing factor, since this leads to genomic instability. “Inactivation of cell cycle checkpoints is a major cause of genomic instability and cancer in cells.” An improperly attached chromosome stalls the cell in metaphase a. No MT attaching to the right. Cell arrests at metaphase. (asymmetric tension triggers cell cycle arrest) b. Give tension applied with needle so the cell can proceed to anaphase  What would happen if there was no checkpoint arrest? You can get abnormal number of chromosomes at both daughter cells  Will this tricked cell produce normal daughters? NO Metaphase of Mitosis  Paired homologous (identical) sister chromatids form 46 mitotic chromosomes  Still diploid: 2n4x4c  46 pairs, all aligned at the middle of the cell  Still 92 pieces of double stranded DNA (chromatin) but now they are compacted as 92 chromatids  The paired 92 chromatids make 46 mitotic chromosomes. How do you know you’re in anaphase? Sister chromatids have come apart but they aren’t very close to the poles yet How many pieces of compacted chromatin in anaphase of humans? 92. How many cells in anaphase? One cell. Anaphase Homologous Sister chromatids separate Anaphase A. Chromatids move towards the poles by shortening the chromosomal MTs. Anaphase B. Poles move apart. Slide spindle fibers to do that, cell will extend polar spindle fiber to push apart the poles. What causes the poles to move apart in Anaphase? In A, shortening MT. In B, sliding polar spindle fiber Anaphase A Poles don’t move. Individual chromatids move toward poles These sister chromatids have to be in amphitelic orientation No change in the distance between the poles but MTs get shorter (the black lines are MT) What is the major force moving the sister chromatids in anaphase A? the shortening of MT due to dynamic instability. Does not require ATP. These are identical Anaphase B What is one of the major forces moving the poles apart in anaphase B? polar spindle fibers meet each other near the center with some kinesin in there. Now we have sliding pushing them apart. Telophase—still one cell! How do you know you’re in telophase? Chromatin becomes disperse and clustered near the poles. No mitotic chromosomes seen at the end of telophase. Starting to look like two cells but STILL ONE CELL that has two nuclei Cytokinesis 1. Contractile ring forms to pinch off cell at the center. This is microfilaments and myosin II 2. Two daughter cells produced, each in G1 3. Each cell has 46 strands of disperse chromatin. 4. Each is 2n2x2c. They are diploid all throughout  Are both daughter cells genetically identical? YES  Are they genetically identical to the mother cell? YES. Cytokinesis (the cell division)—obviously two cells Metaphase 2n4x4c (92 chromatids, paired to make 46 mitotic chromosomes) Two daughter cells, both diploid (2n2x2c) Each daughter cell has the same 46 pieces of disperse chromatin in G1 after cytokinesis identical daughters Cytokinesis Note: This is conventional myosin, myosin II What phase are the daughter cells in? G1 How many pieces of disperse chromatin in each daughter cell? After cytokinesis : 46 Identification of myosin II at the cleavage furrow with fluorescent antibodies 4/13/16 Meiosis The goal of mitosis is equational division (for identical daughter cells) 2n2x2c  2n4x4c  2n2x2c MITOSIS The goal of meiosis is reduction division (2 ROUNDS) 2n4x4c  1n2x2c  1n1x1c MEIOSIS The goal of sexual reproduction is a return to the diploid state Mitosis is nuclear division (equational division). There is an S-phase in between the next mitosis (MITOSIS  S-PHASE  MITOSIS) Meiosis is 2 nuclear divisions with no S-phase in between (reduction division) reducing amount of DNA into just the necessary amount for an egg or sperm Signals received in pre-meiotic cells trigger them (in G1) to start pre-meiotic S-phase. In other words, the decision about whether to go into mitosis or meiosis is made in G1. Body cell goes through only Mitosis. Sex cell goes through Mitosis and Meiosis. Primary meiocytes are cells that have just come out of pre-meiotic G2 and have entered prophase I of meiosis (first round) Meiotic prophase nuclei differ from mitotic prophase nuclei due to formation of synaptonemal complexes and bivalents (tetrads) in meiotic prophase I Only see bivalents tetrads in meiosis I. so you can use that to tell if you’re in meiosis 1 or meiosis 2. Bivalents (tetrads) in meiosis:  It is bivalent because it is two meiotic chromosomes  It is a tetrad because it has four chromatids  It is syntelic because both sister chromatids face the same pole In mitosis, copies of each chromatid align at the metaphase plate In meiosis, copies of each meiotic chromosome align at the metaphase plate (lining up mom dad) when split, they’re splitting differently and unevenly every time you shuffle the aligning Human karyotype o One mitotic chromosome. o Two pieces of condensed chromatin. o Two sister chromatids There are two identical double stranded chromatids above. In meiosis, you have two of these sets. o There are 23 bivalents in meiosis I o 23 pairs in metaphase of meiosis I First meiotic division (Meiosis I): we have twice as much DNA as in G1; Meiosis I produces two Homologous chromosomes pair as bivalents or tetrads (2n4x4c) Intact meiotic chromosomes (as paired chromatids) separate No S-phase in between Second meiotic division (Meiosis II): no more bivalents; Meiosis II produces 4 haploid gametes Homologous chromatids separate Each nucleus gets one set of chromatids Four haploid gametes, all 1n1x1c Mitotic Chromosome  Paired sister chromatids  Both chromatids are genetically identical  Same thing in meiosis but we’ll call it a meiotic chromosome for clarity  P2 paired together then separate when in meiosis 2 Bivalent (tetrad) in Meiosis I  Homologous Chromosomes (homologous meiotic chromosomes)  Paired meiotic chromosomes  Both meiotic chromosomes carry similar information but they are not genetically identical  What do you get when you pull them apart? Mitosis: you get chromatids. Meiosis 1: stay together. Meiosis 2: looks like mitosis, chromatids will come apart.  The goal of Meiosis is haploidization Each gamete contains 23 chromatids (22 autosomes and 1 sex chromosome). Each gamete is 1n1x1c Cohesin in mitosis and meiosis Removing cohesin causes separation of sister chromatids in both mitosis and in meiosis II  The sister chromatids are genetically identical  They are held together with cohesin  In metaphase, cohesin is only at the centromere  Cohesin is removed at the end of metaphase in response to APC (anaphase promoting complex)  This allows sister chromatids to separate in anaphase 1. During prophase, after DNA replication, the sister chromatids are held together by cohesin 2. In prometaphase, most of the cohesion is removed, except for some at the centromere 3. At the end of metaphase, a cyclin-Cdk complex activates the anaphase-promoting complex (APC), which activates separase, resulting in the removal of the remaining cohesin How does APC work? It acts by turning off existing signals o It is a multisubunit enzyme complex o It causes ubiquitination of many substrates.  A substrate is what an enzyme acts upon.  Ubiquitination attaches ubiquitin to a substrate. o Ubiquitination labels substrates for destruction. o A substrate with ubiquitin on it is targeted for degradation. (breakdown) o APC itself doesn’t destroy proteins themselves; it targets the proteins for destruction by other processes Ubiquitination targets proteins for destruction 1. A protein is targeted for degradation (breakdown) 2. An enzyme attaches ubiquitin to the protein and its gone 3. And the target protein is recognized by a proteasome (only to destroy proteins with ubiquitin on it) so you can turn over signals because you don’t want to have 30 signals to tell you to do things all at ones. You want one at a time and get rid of it afterwards. 4. Ubiquitin is released and recycled 5. The proteasome hydrolyzes the target protein What’s the point? The cell cycle can be regulated not only by the appearance of controlling factors, but also by their regulated disappearance Two ways to decrease protein amounts: decrease synthesis or increase degradation APC can specifically target proteins for degradation by ubiquitination Cohesin holds sister chromatids together: In mitosis (until anaphase) In meiosis I (until anaphase II of meiosis II) You should see APC in meiosis II only Two kinds of genetic variability in meiosis: these do not cause changes in DNA sequence and do not cause genetic mutations 1. Genetic recombination by crossovers 2. Independent assortment of maternal and paternal genes A mutation is a change in the DNA sequence of a gene A gene is a locus or region of DNA that encodes a functional RNA or protein product, and is the molecular unit of heredity Does all of DNA contain genes? NO 4/15/16 Cross-overs and genetic recombination: All of the next 7 slides are in prophase I of Meiosis I Synaptonemal complexes in prophase I of meiosis I They’re in Prophase I only Homologous chromosomes are held together by synaptonemal complexes Cohesin holds together sister chromatids in each meiotic chromosome Synaptonemal complexes hold together bivalents Tetrad Recombination This happens in prophase I of meiosis  Crossing-over and recombination occurs between (not within) non-sister chromatids (between red and grey; mom and dad)  Would this cause a mutation? NO  This is not recognized as a replication error and no checkpoint arrest is made  Would a cross-over between sister chromatids change anything? It wouldn’t do anything. Because they’re genetically identical Extended meiotic prophase I 1. In the beginning of meiotic prophase I, chromatin condenses (DNA compacts) lots of condensin 2. Then homologous chromosomes pair (to form bivalents or tetrads) and synaptonemal complexes form (mom and dad pair up) 3. Crossing over and genetic recombination can now happen while the synaptonemal complex Is there 4. Near the end, synaptonemal complexes disappear and recombination stops. Now cross-over points hold them together  How can you tell that this isn’t prophase of mitosis? Never see tetrads and bivalents, synaptonemal complexes, genetically mixing up in Prophase of mitosis. ONLY FOR PROPHASE I OF MEIOSIS Early prophase I Compacted meiotic chromosomes become visible Each is a pair of identical chromatids (from S phase duplication)  How can you tell that this is meiotic prophase I and not meiotic prophase II? Twice as much DNA in Prophase I than Prophase II. Middle of prophase I A process called synapsis starts to pair homologous chromosomes Synapsis makes synaptonemal complexes Homologous chromosomes are held together by synaptonemal complexes (do recombination) Paired homologous chromosomes are called bivalents or tetrads There are 23 bivalents When the synaptonemal complexes are gone, chiasmata continue hold them together Extended meiotic prophase I: Recap 1. In the beginning of meiotic prophase I, chromatin condenses 2. Then homologous chromosomes pair (to form bivalents or tetrads) and synaptonemal complexes form 3. Crossing over and genetic recombination can now happen 4. Near the end, synaptonemal complexes disappear and recombination stops The appearance of synaptonemal complexes is transient. They are gone by the end of meiotic prophase I. Now we leave prophase I of meiosis I Metaphase I of meiosis I This is still the primary meiocyte There are still 23 bivalents Is cohesin still there? YES Anaphase I of meiosis I Bivalents are gone. No more bivalents or tetrads in the rest of meiosis Paired sister chromatids begin to move towards the poles Telophase I of meiosis I This is still the primary meiocyte The first nuclear division is complete at the end of telophase I Cytokinesis follows to generate the two secondary meiocytes Meiosis II Prophase II: Since the chromatin was dispersed in telophase I, it must re-condense for prophase II The secondary meiocyte does not have bivalents. They are paired sister chromatids (like in mitosis) No genetic recombination in the secondary meiocyte. Why? We already did that in Meiosis I Metaphase II: Metaphase II has 23 pairs of sister chromatids at the metaphase plate. How many were there in metaphase I? 23 Bivalents. 46 pairs of sister chromatids. From 46 in Metaphase I to 23 in Metaphase II. Anaphase II: Sister chromatids separate. 23 pieces of compacted chromatin (chromatids) go towards each pole MEIOSIS I: When to hold and when to let go? 1. What holds the sister chromatids together throughout meoitic prophase I? o Cohesin When does all of the cohesin let go? Not until anaphase II of meiosis II 2. What holds the homologous meiotic chromosomes together in prophase I of meiosis I? o Synaptonemal complex in the middle of prophase I o Chiasmata in later prophase I When do these let go? In anaphase I 3. Why have synaptonemal complexes? In anaphase I o For Genetic Recombination (to add genetic diversity) good for evolution Which of the following are seen in both Prophase I and Prophase II of meiosis? A. Synaptonemal complexes—NO, only in Prophase I B. Tetrads (bivalents)—NO, only in Prophase I C. Chiasmata and genetic recombination—NO, only in Prophase I D. Copies of the entire maternal and paternal genomes—NO, only primary meiocyte does E. Compacted chromatin—YES Independent assortment of maternal and paternal genes Independent assortment after one complete meiosis: mixed up contribution from mom & dad  Four gametes Independent assortment after a different complete meiosis Each round of meiosis can produce different grouping in each gametes Analogy for Independent Assortment: You have two decks of cards: • A blue deck. This deck has 46 cards and they are all blue on the back but there are 46 kinds • A red deck. This deck also has 46 cards but they are all red. They have similar 46 kinds, just different printing Mix them together on the floor. There are now 92 cards. Pull out 46 with your eyes closed. What did you get? Now put them back, remix and take out 46 again. Will it be a different mix? Yes, it may Now add the effects of genetic recombination and independent assortment together The DNA sequences of each chromatid may be different. Each gamete can get different genetic information. Two sources of genetic variability in “normal” Meiosis: 1. Crossing over and recombination. This happens in prophase I It changes the distribution of maternal and paternal genes in some chromatids 2. Independent Assortment of genetic traits (maternal and paternal) This happens in Anaphase I of meiosis  What does all of this do? It changes the distribution of maternal and paternal genes in each gamete. All of this contributes to genetic diversity There are two general kinds of cells in the human body: 1. Somatic cells. “Body” cells. Mitosis but no meiosis. Genetic mutations in somatic cells are not passed along to the progeny 2. Sex cells. Cells that differentiate into either egg or sperm cells. Can do mitosis or meiosis Only sex cell mutations are inherited Sperm and egg both doing mitosis but end differently: Differentiation Sperm growth then differentiation at the end (Meiosis Imeiosis IIdifferentiation) Egg growth and differentiation happen first (DifferentationMeiosis IMeiosis II)


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