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BIOMG 1350 Notes Week 10

by: genehan

BIOMG 1350 Notes Week 10 BIOMG 1350


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Week 10 Notes
Introductory Biology: Cell and Developmental Biology
Garcia-Garcia, M; Huffaker, T
Class Notes
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This 14 page Class Notes was uploaded by genehan on Saturday September 3, 2016. The Class Notes belongs to BIOMG 1350 at Cornell University taught by Garcia-Garcia, M; Huffaker, T in Fall 2015. Since its upload, it has received 6 views. For similar materials see Introductory Biology: Cell and Developmental Biology in Molecular Biology and Genetics at Cornell University.

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Date Created: 09/03/16
BIOMG 1350 Professor Bretscher & Garcia-Garcia Spring 2016 Week 10: Lecture 1 of 2 Monday, April 4, 2016 Lecture Title: Same genome, different cells Lecture Keywords: induction, housekeeping genes, regulated genes, Central Dogma, RNA polymerase, sigma factor, TATA boxes, general transcription factors, enhancers, mediators, repressors, histone, chromatin remodeling complexes, histone modifying enzymes, histone modifications, euchromatin, heterochromatin, basal level of expression, loss of expression, ectopic expression, Waddington’s landscape, epigenetic landscape, master regulators, positive regulatory feedback, gene regulatory cascades, DNA methylation, CpG islands ** This is the last lecture that will be on Prelim 2** Prelim #2 - Monday April 11th 9.05-9.55 am Kennedy Hall Auditorim - last names starting with A to Q Morrison Hall room 146 - last names starting R to Z Lectures 9 to 16, plus active learning sections 4-8 1. Transplants uncovered a new concept – induction a. Frog embryos were used and they transplanted a portion from the donor tissue to the opposite side of a recipient embryo to observe what those cells were. A tadpole with two heads resulted. b. Cells can convince, or induce, other cells to do something different than what they originally would do. Those cells were not responsible for giving rise to the head. By induction, those cells were duplicated and induced by nearby cells to give rise to a second head. 2. Two yeast strains have different genomes and hence different functions, proteins, and morphology. On the other hand, multicellular organisms have multiple cell types but all cells have the same genome. a. Two cell types have different morphology, different functions and different protein, but have the same genome. (example: intestinal epithelial cells vs. neurons) b. The same genes are expressed in a different way to give rise to the different cell types. 3. Housekeeping genes are expressed in all cells and perform functions required for all cells to survive. 4. Regulated genes are only expressed in certain cells and/or situations and perform specific functions that allow cells to specialize and/or respond to environmental conditions. 5. Central Dogma - DNA  RNA synthesis (transcription)  RNA  protein synthesis (translation)  protein 6. RNA polymerase is an enzyme with high affinity for DNA and transcribes DNA into RNA. a. In prokaryotes - i. Promoters are located very close to transcription start sites and when bound to the sigma factor, RNA synthesis begins. ii. The sigma factor is displaced and unwinds the double helix and makes space for the growing RNA strand that is synthesized 5’ to 3’ processively. iii. The sigma factor rebinds while the completed RNA chain is released. b. In eukaryotes - c. There are 3 RNA polymerases responsible for reading different part of the genomes. i. RNA pol II – mRNA protein encoding genes ii. RNA pol I – rRNA iii. RNA pol III – tRNA d. In eukaryotes, promoters are sometimes called TATA boxes. The main difference is all the subunits RNA polymerase II has. i. General transcription factors aid RNA polymerase to aid the transcription by recognizing the promoter. These factors can be modified and regulate their function. They are present in all cells, control all genes, and don’t contribute to differential gene expression amongst cells. e. Enhancers are relatively short binding sites for an activator protein. These sequences are non-coding. Their role is to regulate transcription in nearby genes. i. They can be close or far from the gene to influence it. ii. If upstream, the enhancer is located 5’ relative to the gene it is influencing. If downstream, the enhancer is towards the 3’ end. iii. Enhancers can work with a promoter from another gene. iv. Enhancers are recognized and bound by transcription factors that can be activators and repressors. 1. Proteins that encode these transcription factors usually have either a homeodomain, Zinc finger, or leucine zipper. 2. These transcription factors are present in only some cells and only control genes that contain specific regulatory sequences. 7. Enhancers regulate transcription by influencing the recruitment of general transcription factors and introducing changes in chromatin such as chromatin packing and histone modifications. a. Transcription factors can enhance the rate transcription is initiated. b. A mediator protein mediates by forming 3D loops in the DNA to bring the enhancer to where the promoter is to facilitate the formation of a transcriptional complex. c. If a repressor is bound to the regulatory element, transcription is delayed or halted. The repressor protein blocks access to the promoter. 8. Chromatin comes in many different compacted structures. a. During mitosis, chromatin is densely packed. b. During interphase, DNA is bound to histones to form nucleosomes. c. Histone cores are formed by an octamer of histone subunits. DNA binds around this puck-like structure. The histones recognize the negative charge of DNA so they can bind any type of DNA. d. Each nucleosome is formed by an octamer of histone proteins, about 147 nucleotide base pairs of DNA, and a linker DNA not bound to histones. e. Chromatin remodeling complexes can remodel the histone positions. This can limit the accessibility of RNA polymerase II to DNA – can relax the chromatin packing by introducing more linker DNA to enhance transcription or make the chromatin more compact. f. Histone tails can be methylated, phosphorylated, or acylated. These modifications can either be enhancing or repressing for nearby genes. For example, methylation of histone 9 results in gene silencing. g. Histone modifying enzymes are recruited by activators or repressors and introduce heritable changes onto DNA. h. Histone modifications can be perpetuated after cell division i. The daughter cell will have the same marker as the parental cell if the parental cell had a certain histone modification at a specific area. 9. Chromatin states – euchromatin (transcriptionally repressed chromatin) and heterochromatin (transcriptionally active chromatin). a. DNA is marked with certain histone tail modifications and this can impact the compacting of chromosomes. 10. Enhancer elements and transcription factors control differential gene expression. a. Cells have the same defined set of genes controlled by regulatory sequences and differential gene expression is determined by the presence of absence of a transcription factor in a given cell. 11. Iclicker question – The gene Bigred is controlled by an enhancer element which is located 50 kb upstream of a conserved TATA box promoter. The enhancer element can be bound by an activator and a repressor. Which of the following statements below indicates a possible scenario? a. When only the repressor is present, the gene will not be expressed. (Possible, the repressor can impede the recruitment of RNA polymerase II) b. When both repressor and activator are present, the gene is not expressed. (Possible, the repressor likely binds the enhancer with higher affinity than the activator) c. In the absence of both repressor and activator the gene will be expressed. (Possible, enhancers are not required for expression. It is likely that the TATA box recruits general transcription factors and RNA polymerase II with high affinity) d. All of the above. e. Not enough information to know. 12. The basal level of expression is determined by the strength of its promoter. A strong promoter can efficiently activate transcription and ensure high level of expression, while a weak promoter may promote low level of expression or no expression. 13. During development, many genes are expressed in patterns. Some genes are only expressed in specific cells. a. Precise transcriptional control is important for development. b. The expression of even skipped gene results in a normal larva with 7 stripes. c. When there is loss of expression of this gene, there is abnormal larva morphology. d. If ectopic expression occurs in which the gene is expressed everywhere, there is also abnormal larva morphology. e. During development, genes need to be expressed in the right place and time. 14. Waddington’s landscape represents cell specification and how cells take a series of decisions that ultimately determines their fate. a. Think of a marble rolling down an epigenetic landscape in which it will eventually become something specific. b. The cell takes different paths until it is specified. Each decision results in the turning on or off by different genes. c. Differential gene expression leads to cell differentiation and cell specification. 15. The combination of transcription factors can be integrated through regulatory sequences. a. A single transcription factor can regulate simultaneously multiple genes. b. Master regulators are transcription factors that can regulate simultaneously multiple genes in such a way that a whole organ can be specified. These master regulators are necessary and sufficient to producing an eye, for example. (If the eyeless gene is expressed in leg, a fully functional eye grows on the leg) 16. Cells remember the developmental decisions they have made through positive feedback loops, gene regulatory cascades, inheritable modifications, histone modifications. a. In a positive regulatory feedback, during development, a transient signal turns on expression of protein A. The effect of this signal is remembered throughout the cell’s life. b. Gene regulatory cascades - a transient signal turns on expression of protein A and A turns on expression of protein B, then B turns on expression of protein C, and so on so forth. This succession of transient changes in gene expression produces long-lasting effects on gene expression. c. Histone modifications propagate themselves and are transmitted from one cell to its daughter cells. d. DNA methylation occurs at CpG islands, areas in the genome rich in C-G nucleotides and will silence complete regions of DNA. DNA methyl transferases can introduce the same modifications of newly synthesized strands. BIOMG 1350 Professor Bretscher & Garcia­Garcia Spring 2016 Week 10: Lecture 2 of 2 Wednesday, April 6, 2016 Lecture Title: Social Networking I Lecture Keywords: cell specification, cell signaling, signal transduction, effector proteins,  ligands, cell­surface receptors, intracellular receptors, GPCR, ion­channel­coupled receptors,  enzyme­coupled receptors, endocrine, paracrine, neuronal, and contact­dependent signaling,  second messengers, adenylyl cyclase, cAMP, pKA, phospholipase C, IP3, diacylglycerol, PKC,  effectors, acetylcholine, signal integration, ligand and receptor degradation, constitutive and  regulated inhibitor 1. Cell specification leads to regulation of gene expression (such as insulin release). 2. Cells find ways to communicate with each and regulate their decisions and behavior through cell signaling. a. Cell signals can be external to the cell, received and decoded, and it modifies the cell’s behavior as a response to the signal. This is also known as signal transduction. b. A receptor recognizes a signal molecule, then intracellular signaling molecules function in a cascade and ultimately modify activity of different effector proteins that lead to the target cell responses. 3. Signal input – some of the signals can be proteins, gases, peptides, light, and mechanosensory stimulus. When the signal is a physical entity, signals can be called ligands. a. Receptors that recognize these ligands can be cell-surface receptors (for hydrophilic signal molecules), or intracellular receptors (for small enough and hydrophobic signal molecules that can enter the cell, such as a steroid hormone). b. Different types of cell surface receptors include G-protein coupled receptors (GPCR) responsible for our vision and smell, ion-channel-coupled receptors embedded in the plasma membrane activated exclusively when bound to a signal molecule (neuron stimulation), and enzyme-coupled receptors that can activate their own catalytic activity. c. Each receptor activates specific intracellular proteins. 4. When the signal is light, specialized structures rich in GPCR transduce signals by secreting neurotransmitters to the brain to give us vision. 5. Not all cells can respond to a specific signal so only cells with the appropriate receptor will be able to respond. Each cell contains a limited set of receptors. 6. Different types of signaling - a. Endocrine signaling cells secrete hormones that travel through bloodstream to be distributed widely in body to influence cells far in distance. b. Paracrine signaling works locally since its molecules cannot travel in bloodstream. c. Neuronal signaling is limited to the proximity of a neuron and a target cell. d. Contact-dependent signaling involves signals embedded in a plasma membrane that can only influence a target cell if in contact. 7. Intracellular signaling proteins work as switches when cycling between On and Off states. a. Signals can both proteins on or off. A pointed arrow represents a positive relationship, such as activation whereas a blunt arrow is a negative relationship, such as deactivation. 8. Production of second messengers a. Adenylyl cyclase is active in generating cAMP from ATP. cAMP are small messenger molecules that amplify and relay signals to act on intracellular signaling proteins. b. A possible signaling pathway results in cAMP activating pKA (protein kinase A), which then phosphorylates effectors that ultimately change the cell behavior. c. Another intracellular signaling pathway for GPCR leads to activated phospholipase C. This releases IP3 and diacylglycerol that leads to opening calcium channels and activates PKC only if both calcium ions and diacylglycerol are bound. 9. Iclicker question – Indicate which one of the statements below about GPCR is false? a. Always activate a G protein b. Lead to activation of enzymes c. Function similar to GEFs d. Always lead to production of cAMP – FALSE – different cells can produce different second messengers e. Are transmembrane proteins 10. Effectors determine the ultimate cell response. They are the last proteins in the signaling cascade. a. Activation of intermediate proteins in the end will change cells in different ways. 11. Cells can respond to signals by altering protein function (fast) or altering protein synthesis (slow). The speed of the response depends on what effectors are activated. a. Signals requiring a lot of transducing proteins will take longer than signals that only require a few proteins. b. If transcription needs to be initiated, this will result in a slow response (needs ribosome, RNA machinery, etc.) c. Signals can be relayed, amplified, and distributed. 12. The same signal can elicit different responses because of different receptors. a. Acetylcholine is a neurotransmitter used by a variety of neurons. i. If it signals a salivary gland cell with a GPCR, this leads to secretion. ii. If it signals a skeletal muscle cell with a channel, the muscle will contract. b. If acetycholine signals a heart cell with a GPCR, this can decrease heartrate. This is an example of how the same receptor can also result in different responses. 13. The integration of signals - a. Two signals received by two receptors can be integrated when downstream in the cascade and the signal is relayed onward. b. Two signals can independently create two molecules that then form a dimer to further relay the signal. c. This allows the cells to correlate signals. Different combinations of signals can produce different outcomes. 14. Iclicker question – Indicate which one of the following scenarios will product the fastest response. a. Signaling by a GPCR that activates a second messenger. b. Signaling by an enzyme-coupled receptor that affects gene expression. c. Signaling by an enzyme coupled receptor that changes protein function d. Signaling by an ion-channel receptor – TRUE – this is based on the generation of a local alteration in the membrane potential, which then is propagated quickly to synaptic terminals of neurons. e. Can not be determined with the information provided 15. Switching off the signal and regulation a. Ligand degradation involves endocytosis and the lysosome. b. Receptor degradation is another way and the receptor and the ligand are both degraded. . c. Autonomous switch – enzyme can only maintain activity for so long, and it switches off itself after a period of time. d. Constitutive inhibitors – a protein is constantly inhibiting a signaling protein and switches it off when the inhibitor is activated. e. Regulated inhibitor is also known as negative feedback. f. The clearance of 2 nd messengers will also work to stop signaling activity. 16. Up to 7000 of our 23,000 genes in our genome are devoted to cell signaling. So, many challenges in drug design are due to target specificity and secondary effects. a. Many times, it is up to chance to see if a tested drug works. (“Happy Accidents”


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