Unit 3 - Unit 3 Control of Gene Expression and Animal...

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Unformatted text preview: Unit 3 Control of Gene Expression and Animal Development 42 UNIT 3: CONTROL OF GENE EXPRESSION AND ANIMAL DEVELOPMENT Readings: Chapter 18 (pages 351-373) Chapter 16 (pages 320-323) Chapter 20 (pages 412-414) Chapter 21 "Morphogens and Homeotic Genes" at the end of this unit Chapter 47 To Do This Unit: 1. 2. 3. 4. 5. 6. Skim the key concepts and objectives below. Read the text assignment. Examine the demonstration materials, both online and in the Study Center. Study the slides on the course website, using the notes at the end of this unit to guide your study. Write out the answers to the objectives, using the text and demonstration materials. Take an examination. KEY CONCEPTS AND OBJECTIVES: After you have studied the material in this unit you should understand the following concepts and you should be able to carry out the objectives listed for each. It is very "expensive" for the cell to synthesize enzymes when they are not needed, so various control mechanisms act to turn on or turn off specific genes. 1. Using Fig. 18.2 (p. 352) explain the two ways in which bacteria can control their metabolic pathways. Of the two, which would produce the more rapid response? Why? First, cells can adjust by using feedback inhibition. When the environment provides what it needs, it can turn off its production process and conserve its m aterials. Second, they can adjust the expression of their own genes. in hibition of the first enzyme in the pathway is a more rapid response than the respression of gene expression (long term response). The gene inhibition occurs more later in the pathway where as the enzyme repression occurs earlier. 2. Using Fig. 18.3 (p. 353), explain what an operator is and describe the trp operon. In doing so, be able to answer the following questions: a) How many genes does the transcription unit code for? Is there one mRNA molecule produced by transcription of these genes or many smaller ones? How does the cell correctly translate the coding information for each polypeptide? I n terms of trp operon, it has 5 genes that code for one transcription unit and thus all 5 genes are coded into one mRNA with one promoter for all. The coding is correctly translated because the mRNA is "punctuated" with start and stop codons for each enzyme subunit gene. Unit 3 Control of Gene Expression and Animal Development 43 b) State the roles of the promoter and regulatory genes. Relative to the operator, where are each of these genes located? Are they near the operator or far from it? t he promoter is the site of mRNA polymerase binding. The regulatory gene produces trp repressor. t he regulatory gene is situated away from the operon it controls and has its own promoter. The regulatory gene does not have an operator. The operator is situated either in the promoter itself or between the promoter and the genes of operon. NOTE: operon: the entrie stretch of DNA required for enzyme production for trp pathway. it has operator that precedes it. The operator is controlled by the repressor that switches it off. The REPRESSOR is produced by the regualtory gene on a s eparate loci. c) What is a repressor protein and what does it do? Is the trp repressor active or inactive when synthesized? What is a corepressor and what does it do? repressor protein is produced by regualtory gene. it switches the operon off by binding to the operator that precedes it. The repressor protein is controlled by a corepressor. The repressor is an allosteric enzyme that when the corepressor is present (tryptophan) the enzyme becomes activated and binds to the operator. The relative amount of repressors to operators determines the duration of the operon state. Higher the number of repressors, less the operon will be expressed. d) How does a repressible operon control gene transcription? What happens when there is no tryptophan in the cell? What happens when there is an abundance of tryptophan in the cell? REPRESSIBLE OPERON: the transcription is usually on but can be inhibited in the presence of an regulatory protein such as tryptophan (corepressor). INDUCIBLE OPERON: the transcription is usually off but can be stimulated to express when a specific small molecule interacts with regulatory protein. ex: lac operon (for lactose). The difference is that the trp operon's repressor is inactive alone. it needs the corepressor to become active. The inducible operon also functions the same, but in this case, the r epressor is active by itself. then the inducer (allolactose) binds to the repressor and inactivates it when lactose is consumed. And thus with the consmption of lactose, the operon is induced to transcription. W hen there's no tryptophan in the cell, the operon is not completely turned off. the bidingin of repressors are reversible so the activity level of the operator is relative to the number of repressors around. in the abundance of tryptophan the operator activity will be greatly reduced so less tryptophan will be produced by the cell itself. 3. NOTE: inducible enzymes for c atabolic activity. repressible enzymes for anabolic activity. a) Using the information in Fig. 18.4 (p. 354), explain what an operon is and show how the expression of the lac operon of E. coli is regulated according to the operon model. This simplified diagram of the lac operon may help: Regulator Activator Promoter Operator Structural Genes Z Y A Identify and explain the roles of the inducer, the operator, the promoter, the repressor protein (is it active or inactive when synthesized?), the regulatory gene, and the structural genes in prokaryotic cells. Explain how the presence of lactose induces the operon. OPERON: promoter + Oprator + coding DNA NOTE: the operator for lac operon follows the promoter. the operator for trp operon is in the promoter. The lac operon expression is controlled by the oregulatory gene which codes for an active repressor which binds to the operator and inhibit the expression of the structural gene. But once lactose is consumed, containing an isomer allolactose, then the repressor becomes inactive and so the lac oepron is activated and the transcrition begins. Unit 3 Control of Gene Expression and Animal Development 44 b) How is a repressible operon (trp) different from an inducible operon (lac)? Focus on the activity of the repressor protein. Why are both considered examples of negative control? See demo. t he repressible operon has an repressor that starts inactive so the transcription is usually on but can be inhibited with the pressence of t he repressor activator (corepresor). the inducible operon starts active so the transcription is inhibited for the start. then with the presence of an inducer, the repressor becomes in active and the transcription is induced. Both are examples of negative control because the operons are switched off by the active form of the repressor protein. c) What is the advantage to the bacteria of having their DNA organized in operons? the advantage is that a single on, off switch can control the entire cluster of functionally related genes. 4. Using the information in the text (p. 355) and in the demo, differentiate between positive regulation and negative regulation of prokaryotic gene transcription. Can both types of control operate at the same time? in positive regualtion, the binding of RNA pol is facilitated and therefore the transcription rate is increased. and so the attachment of ie. CAP stimulates gene expression. IN POSITIVE REGULATION< REGULATORY PROTEIN INTERACTS DIRECTLY WITH THE GENOME TO SWITCH TRANSCRIPTION ON. 5. Give one example of positive control in bacteria and explain how it works in E. coli (Fig. 18.5, p. 355). in senses glucose through the use of positive control. it uses the cAMP which accumulates when glucose is scarce. then the regualtory protein CAP (activator) becomes activated and binds to the promoter region to increase the affi nity for RNA pol binding and so transcription is promoted. THE ATTACHMENT OF CAP DIRECTLY STIMULATES GENE EXPRESSION. a) In the drawing below of lac operon indicate where the RNA polymerase binds and where the active CAP binds. Why is positive control important in the lac operon (see demo)1? Structural Genes Regulator Activator Promoter Operator Z Y A cap binding site RNA pol binding site CAP binds to the unstream end of the promoter and the RNA polymerase binds to the promoter and the active CAP. It turns out that the promoter for the lac operon is weak (i.e., not close to the consensus sequence) so the RNA polymerase does not bind strongly to the promoter and transcription of the structural genes occurs only at low levels. When active CAP is present, however, it facilitates the binding of RNA polymerase and transcription is rapid. 1 Unit 3 Control of Gene Expression and Animal Development b) complete the chart below to understand positive vs. negative gene regulation: Operon Level of transcription of lac operon (none, low, high) none 45 Environmental Conditions glucose, no lactose glucose, lactose no glucose, no lactose no glucose, lactose CAP Bound no Repressor Bound active no inactive active low low yes yes inactive high Each gene codes for only one kind of messenger RNA; regulators determine if and when each gene will be transcribed into its particular mRNA. Environmental influences can act by helping to turn on or to turn off the transcriptional activity of the various genes or by influencing the synthesis and activity of enzymes. 6. a) Using Fig. 16.21 (pp. 320-321), explain the general organization of the eukaryotic chromosome and give the general roles of the nucleosomes and histone proteins. Differentiate between euchromatin and heterochromatin. the histones group by 8 and form a histone core. The histone groups by sticking the tail out and the heads binding t ogether. To this histone bead, the double helix DNA wraps twice around it. this forms the nucleosomes. Then these beaded s trands coil up tightly to form the 30-nm fi ber. the fi ber coils and forms the looped domains with the scaffold core. then the looped domains coil and fold to form the tightou wound chromosome. the nucleosomes help control the movement of DNA, position wise and windind, by chemically being altered. euchromatin is less tightly wound that it can be expressed. heterochromatin is more compact that it is inaccessible for t ranscritipn. b) How does the organization of the eukaryotic genome differ from that of prokaryotes? Complete the following chart to summarize the differences. Consider the number of chromosomes, whether or not they are linear or circular, presence and absence of exons and introns, and the presence and absence of operons. Prokaryotes number of chromosomes? generally haploid or diploid? linear or circular chromosomes? introns & exons present? operons present? 1 haploid circular exons present Eukaryotes multiple diploid linear exons and introns present c) What percentage of the eukaryotic genome is not translated? What does the rest of the DNA consist of? See p. 434 How much of the prokaryotic genome is transcribed and translated? ________ See demo. In prokaryotic genmes, most of the DNA codes for protein, t RNA, or rRNA; the small amount of noncoding DNA consists mainly of regulatory sequences, such as promoters. The c oding sequence of nucleotides along a prokaryotic gene proceeds from start to fi nish without interruption by noncoding s equences (introns). the rest of the DNA consists of introns, regulatory sequences, unique noncoding DNA, 98.5% _______ repetitive DNA, and t ransposable elements. most of it Unit 3 Control of Gene Expression and Animal Development 46 d) Explain what is meant by a multigene family, give an example, and explain how they might have arisen. How might transposition be involved? Normally all cells of a multicellular organism are genetically identical; control mechanisms determine which of the cell's inherited instructions will be acted upon and which will not. In all organisms the expression of certain genes is most commonly regulated at the level of transcription. In eukaryotes, most control is positive; transcription factors recognize and bind to the promoter sequence. In turn, they bind the activators and help the RNA polymerase to bind and position it at the right starting point. 7. Discuss the following aspects of control of gene expression in eukaryotes. a) State the role of histone acetylation in the regulation of gene transcription see pp. 357-358). t he histone acetylation neutralizes the histone tails from (+) charge and so the histone tails no longer bind to neighboring nucelosomes and allows the looser organization of chromosome. NOTE: methylated is usually permanent and c an be transferred to t he zygote METHYLATION: addition of CH3 to histone tails; promotes condensation of the chromatin so the gene is not expressed. related to HETEROchromatin. GENOMIC IMPRINTING: methylation permanently regulates expression of either the materal or paternal allele of particular genes at the start of development. b) Explain the role that DNA methylation plays in modifying chromatin. How is methylation passed on from cell to cell? How is DNA methylation related to euchromatin/heterochromatin? What is genomic imprinting? c) Using Figs. 18.8-18.9 (pp. 359-360), explain the role of activators and enhancers in controlling gene transcription in eukaryotes. Where are the regions located with respect to the gene to be transcribed? What role do transcription factors play? Would a eukaryotic gene be transcribed if no transcription factors were present? Is the control of gene transcription in eukaryotes primarily positive or negative control? ACTIVATOR binds to distal control elements (grouped as enhancer). then when the DNA is bent by the DNA-bending protein, the activators bind to mediator proteins and general transcription factors to from the active transcription initiation complex on the promoter. ENHANCER: distal control elements in a group about thousands of nucleotides upstream or downstream of a gene. a gene can have multimple enhancers and it's gene specifi c. activators bind to the individual distal control elements on the enhancer. TRANSCRIPTION FACTORS: asisst binding of RNA polymerase to the promoter to from the initiation complex. Transcription cannot occur without transcription factors: only when the initiation complex is complete, transcription begins. postive control (?) Unit 3 Control of Gene Expression and Animal Development 47 d) Summarize the differences between prokaryotic and eukaryotic gene control by completing the following chart: Prokaryotes presence of nucleosomes genes organized in operons control primarily positive or negative role of transcription factors role of enhancers levels of control (1 or 2 vs. 3 or more) need for mRNA processing 1 Eukaryotes yes no yes both yes (?) none ? help RNA pol bind to promoter helps bring general transcription f actor in to complete assembly a lot none 5' cap, poly-A-tail, splicing (introns) e) Explain what is meant by post-transcriptional control of gene expression and give three examples. post-transcriptional control fo gene expression is the determinatino of expression by how mRNA is modifi ed after the t ranscription occurs. 1. alternative RNA splicing: the treatment of introns depend on the cell type. alternate expression can arise from one primary transcript. 2. mRNA degradation: euk take a longer time than the prokaryotes to degrade the mRNA and stop expression by protein production. 3. initiation of translation: the mRNA translation can be held back; not suffi cient poly-A-tail, blocked by sequences in 5' UTR to prevent ribsome from binding. f) Use Fig. 18.6 (p. 357) to summarize the various steps in which gene expression can be controlled in eukaryotes. 1. modifi cation of histones. 2. post-transcriptional control 3. presence of enhancers and activators; depending on which combo of enhancers and activators are present, t he gene may be transcripted or not. All the cells in a multicellular organism except the germ cells have genomic equivalence; cells become different because of differences in gene expression. Unit 3 8. Control of Gene Expression and Animal Development 48 Define the terms differentiation and morphogenesis and explain how each process contributes to embryonic development. See pp. 366-367. differentiation: cells become specialized in structure and function morphogenesis: development of body shape and organization. differentiation occurs from difference in regulation of gene in each type of cell. As development of a multicellular organism proceeds, the individual cells become more and more committed to one particular course of differentiation. However, in some organisms, differentiation may be reversible. 9. Is adult cellular differentiation ever reversible? Can cells (nuclei) resume totipotency? See pp. 412-414 and the demo for examples. in plants, mature cells can "dedifferentiate" and give rise to all the specialized cell: TOTIPOTENT in animals it's a lot harder and requires advanced technology like nuclear transplantation. Differentiation is the process by which a cell undergoes a series of changes to become a specialized cell type. It is a matter of progressive determination; development is gradually restricted to one of the many initially possible pathways. 10. Distinguish between the processes of determination and differentiation. Using MyoD as an example and Fig. 18.16 (p. 369), explain the mechanisms of determination and differentiation in the development of muscle cells. differentiation is when the cell becomes specialized in structure and funcion where as in determination is when the cell is irreversibly committed to its fi nal fate. myoD gene codes for MyoD protein which is a transcription factor that binds to the control elements in the t arge genes and stimulate expression. all genes activated by MyoD have enhancer control elements recognized by MyoD. 11. The cytoplasm of the unfertilized egg is responsible for many of the initial characteristics of the cells in an early embryo. Describe how the cytoplasm of the egg impacts early development of the zygote. egg's cytoplasm contains both RNA and proteins encoded by the MOTHER's DNA. the cytoplasm also contains other s ubstances and organelles. the mternal substances infl uce the course of early development; CYTOPLASMIC DETERMINANTS. After the fertilization the zygote undergoes mitosis. Depending on what contents of the cytoplasm it received each cell develops accordingly. Unit 3 12. Control of Gene Expression and Animal Development 49 Using Fig. 18.19 (p. 372) in your text and the section “Morphogens and Homeotic Genes” at the end of this unit, describe the process of pattern formation in Drosophila. In your answer, a) Explain the role that maternal effect genes (e.g., bicoid) have on determining the body axis and pattern formation. maternal effect genes are genes that are active and code for proteins and mRNA that affect the egg and zygote. Bicoid: codes for morphogen that establishes the anterior end of the egg. nanos codes for a morphogen that establishes the posterior end of the egg. pattern formation: development of a spatial organization in which the tissues and organs of an organism are all in their characteristic places. body axis is set by morphogens b) Describe the role of morphogens and positional information in pattern formation in the embryo, using as an example segmentation in Drosophila. What are morphogens? How do they exert their effect on target cells? How does this relate to the process of determination? What is their significance in orchestrating development? morphogens are proteins secreted from a specifi c poin in the embryo where its presence induces the cells around it to develop in a particular way. Morphogens are trasncription factors or lead to activation fo transcription factor in the localized region and thus bring about certain expression of gene. ...
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