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Chapter 28 Regulation of Gene Expression Multiple Choice Questions Principles of gene regulation Page: 1082 Difficulty: Ans: B “Housekeeping genes” in bacteria are commonly expressed constitutively, but not all of these genes are expressed at the same level (the same number of molecules per cell) The primary mechanism responsible for variations in the level of constitutive enzymes from different genes is that: A) B) C) D) E) all constitutive enzymes are synthesized at the same rate, but are not degraded equally their promoters have different affinities for RNA polymerase holoenzyme some constitutively expressed genes are more inducible than others some constitutively expressed genes are more repressible than others the same number of mRNA copies are made from each gene, but are translated at different rates Principles of gene regulation Pages: 1082-1083 Difficulty: Ans: D Which of the following statements correctly describes promoters in E coli? A) B) C) D) A promoter may be present on either side of a gene or in the middle of it All promoters have the same sequence that is recognized by RNA polymerase holoenzyme Every promoter has a different sequence, with little or no resemblance to other promoters Many promoters are similar and resemble a consensus sequence, which has the highest affinity for RNA polymerase holoenzyme E) Promoters are not essential for gene transcription, but can increase its rate by two- to three-fold Principles of gene regulation Pages: 1083-1084 Difficulty: Ans: D The operator region normally can be bound by: A) B) C) D) E) attenuator inducer mRNA repressor suppressor tRNA Principles of gene regulation Pages: 1083-1084 Difficulty: Ans: C Small signal molecules that regulate transcription are not known to: A) cause activator proteins to bind DNA sites B) cause repressor proteins to bind DNA sites C) directly bind to DNA sites D) prevent activator proteins from binding to DNA sites E) release repressor proteins from DNA sites Chapter 28 Regulation of Gene Expression 327 Principles of gene regulation Pages: 1083-1085 Difficulty: Ans: E The diagram below represents a hypothetical operon in the bacterium E coli The operon consists of two structural genes (A and B), which code for the enzymes A-ase and B-ase, respectively, and also includes P (promoter) and O (operator) regions as shown When a certain compound (X) is added to the growth medium of E coli, the separate enzymes A-ase and B-ase are both synthesized at a 50-fold higher rate than in the absence of X (X has a molecular weight of about 200.) Which of the following statements is true of the operon decribed above? A) All four genes (A, B, O, and P) will be transcribed into an mRNA that will then be translated into four different proteins B) The 3' end of the mRNA from the operon will correspond to the left end of the operon as drawn C) The 5' end of the messenger from this operon will correspond to the right end of the operon as drawn D) The repressor for this operon binds just to the right of A E) When RNA polymerase makes mRNA from this operon, it begins RNA synthesis just to the left of gene A Principles of gene regulation Pages: 1083-1085 Difficulty: Ans: A The diagram below represents a hypothetical operon in the bacterium E coli The operon consists of two structural genes (A and B) that code for the enzymes A-ase and B-ase, respectively, and also includes P (promoter) and O (operator) regions as shown When a certain compound (X) is added to the growth medium of E coli, the separate enzymes A-ase and B-ase are both synthesized at a 50-fold higher rate than in the absence of X (which has a molecular weight of about 200) Which one of the following statements is true of such an operon? A) B) C) D) Adding X to the growth medium causes a repressor protein to be released from the O region Adding X to the growth medium causes a repressor protein to bind tightly to the O region Synthesis of the mRNA from this operon is not changed by the addition of compound X The mRNA copied from this operon will be covalently linked to a short piece of DNA at the 5' end E) Two mRNA molecules are made from this operon, one from gene A the other from gene B Principles of gene regulation Pages: 1085-1087 Difficulty: Ans: B Transcription of the lactose operon in E coli is stimulated by: A) B) C) D) E) a mutation in the repressor gene that strengthens the affinity of the repressor for the operator a mutation in the repressor gene that weakens the affinity of the repressor for the operator a mutation in the repressor gene that weakens the affinity of the repressor for the inducer binding of the repressor to the operator the presence of glucose in the growth medium 328 Chapter 28 Regulation of Gene Expression Principles of gene regulation Page: 1088 Difficulty: Ans: D Protein amino acid side chains can hydrogen bond in the major groove of DNA, and discriminate between each of the four possible base pairs In which one of the following groups of amino acids can all three members potentially be used in such DNA-protein recognition? A) B) C) D) E) Ala, Asn, Glu Arg, Gln, Leu Asn, Gln, Trp Asn, Glu, Lys Glu, Lys, Pro Principles of gene regulation Pages: 1088-1089 Difficulty: Ans: A The DNA binding motif for many prokaryotic regulatory proteins, such as the lac repressor, is: A) B) C) D) E) helix-turn-helix homeobox homeodomain leucine zipper zinc finger 10 Principles of gene regulation Pages: 1088-1091 Difficulty: Ans: D Protein structural motifs often have general functions in common Which one of the following motifs is known to be involved in protein dimer formation, but not in direct protein-DNA interactions? A) B) C) D) E) β-barrel helix-turn-helix homeodomain leucine zipper zinc finger 11 Regulation of gene expression in prokaryotes Pages: 1093-1094 Difficulty: Ans: A Which of the following statements about regulation of the lac operon is true? A) B) C) D) E) Glucose in the growth medium decreases the inducibility by lactose Glucose in the growth medium does not affect the inducibility by lactose Glucose in the growth medium increases the inducibility by lactose Its expression is regulated mainly at the level of translation The lac operon is fully induced whenever lactose is present Chapter 28 Regulation of Gene Expression 329 12 Regulation of gene expression in prokaryotes Page: 1093 Difficulty: Ans: A The binding of CRP (cAMP receptor protein of E coli) to DNA in the lac operon: A) B) C) D) E) assists RNA polymerase binding to the lac promoter is inhibited by a high level of cAMP occurs in the lac repressor region occurs only when glucose is present in the growth medium prevents repressor from binding to the lac operator 13 Regulation of gene expression in prokaryotes Page: 1093 Difficulty: Ans: C Consider the lac operon of E coli When there is neither glucose nor lactose in the growth medium: A) B) C) D) E) CRP protein binds to the lac operator CRP protein displaces the Lac repressor from the lac promoter repressor is bound to the lac operator RNA polymerase binds lac promoter and transcribes the lac operon the operon is fully induced 14 Regulation of gene expression in prokaryotes Page: 1094 Difficulty: Ans: B A regulon is a(n): A) B) C) D) E) group of related triplet codons network of operons with a common regulator operon that is subject to regulation protein that regulates gene expression ribosomal protein that regulates translation 15 Regulation of gene expression in prokaryotes Pages: 1094-1095 Difficulty: Ans: C The tryptophan operon of E coli is repressed by tryptophan added to the growth medium The tryptophan repressor probably: A) B) C) D) E) binds to RNA polymerase when tryptophan is present binds to the trp operator in the absence of tryptophan binds to the trp operator in the presence of tryptophan is a DNA sequence is an attenuator 16 Regulation of gene expression in prokaryotes Pages: 1094-1097 Difficulty: Ans: A Which one of the following statements about the transcription attenuation mechanism is true? A) B) C) D) E) In some operons (e.g., the his operon), attenuation may be the only regulatory mechanism Sequences of the trp operon leader RNA resemble an operator The leader peptide acts by a mechanism that is similar to that of a repressor protein The leader peptide gene of the trp operon includes no Trp codons The leader peptide is an enzyme that catalyzes transcription attenuation 330 Chapter 28 Regulation of Gene Expression 17 Regulation of gene expression in prokaryotes Pages: 1094-1097 Difficulty: Ans: D Which of the following statements is true of the attenuation mechanism used to regulate the tryptophan biosynthetic operon in E coli? A) Attenuation is the only mechanism used to regulate the trp operon B) One of the enzymes in the Trp biosynthetic pathway binds to the mRNA and blocks translation when tryptophan levels are high C) The leader peptide plays a direct role in causing RNA polymerase to attenuate transcription D) Trp codons in the leader peptide gene allow the system to respond to tryptophan levels in the cell E) When tryptophan levels are low, the trp operon transcripts are attenuated (halted) before the operon’s structural genes are transcribed 18 Regulation of gene expression in prokaryotes Pages: 1094-1097 Difficulty: Ans: B Attenuation in the trp operon of E coli: A) B) C) D) E) can adjust transcription of the structural genes upwards when tryptophan is present can fine-tune the transcription of the operon in response to small changes in Trp availability is a mechanism for inhibiting translation of existing (complete) trp mRNAs results from the binding of the Trp repressor to the operator results from the presence of short leader peptides at the 5' end of each structural gene 19 Regulation of gene expression in prokaryotes Pages: 1097-1098 Difficulty: Ans: D RecA protein provides the functional link between DNA damage and the SOS response by displacing the LexA protein from its operator sites on the SOS genes RecA does so by: A) B) C) D) E) associating with polymerase holoenzyme to help it remove LexA from operator bending LexA operator DNA to force dissociation of LexA repressor binding to LexA protein to weaken directly its affinity for operator sites causing self-cleavage of LexA, thus inactivating its binding to operator competitively binding to LexA operators and serving as an activator 20 Regulation of gene expression in prokaryotes Pages: 1098-1099 Difficulty: Ans: D An example of coordinate control is the down-regulation of ribosomal RNA synthesis in response to amino acid starvation, which will cause synthesis of ribosomal proteins to be limited What is the correct order of the following events that participate in the signaling process? Binding of stringent factor to the ribosome Formation of the unusual nucleotide ppGpp Formation of the unusual nucleotide pppGpp Uncharged tRNA binds in the ribosomal A-site A) B) C) D) E) 1, 4, 2, 1, 4, 3, 4, 1, 2, 4, 1, 3, 4, 2, 1, Chapter 28 Regulation of Gene Expression 331 21 Regulation of gene expression in eukaryotes Page: 1103-1104 Difficulty: Ans: B Which one of the following statements about eukaryotic gene regulation is correct? A) B) C) D) E) Large polycistronic transcripts are common Most regulation is positive, involving activators rather than repressors Transcription and translation are mechanistically coupled Transcription does not involve promoters Transcription occurs without major changes in chromosomal organization 22 Regulation of gene expression in eukaryotes Page: 1104 Difficulty: Ans: C Which of the following is a DNA sequence? A) B) C) D) E) Coactivator Corepressor Enhancer Inducer Transactivator 23 Regulation of gene expression in eukaryotes Page: 1104 Difficulty: Ans: E Which one of the following types of eukaryotic regulatory proteins interact with enhancers? A) B) C) D) E) Basal transcription factors Coactivators Repressors TATA-binding proteins Transactivators 24 Regulation of gene expression in eukaryotes Page: 1108 Difficulty: Ans: A Which one of the following is not involved in steroid hormone action? A) B) C) D) E) Cell surface receptors Hormone-receptor complexes Specific DNA sequences Transcription activation and repression Zinc fingers 25 Regulation of gene expression in eukaryotes Page: 1112 Difficulty: Ans: C Which one of the following classes is expressed in the unfertilized egg and is involved in directing the spatial organization of the Drosophila embryo early in development? A) B) C) D) E) Gap genes Homeotic genes Maternal genes Segment polarity genes Segmentation genes 332 Chapter 28 Regulation of Gene Expression 26 Regulation of gene expression in eukaryotes Pages: 1112, 1116 Difficulty: Ans: B Which one of the following classes of genes is involved in specifying the localization of organs in the Drosophila embryo? A) B) C) D) E) Gap genes Homeotic genes Maternal genes Segment polarity genes Segmentation genes 27 Regulation of gene expression in eukaryotes Pages: 1112, 1116 Difficulty: Ans: C In the development of the fly Drosophila, homeotic genes: A) B) C) D) E) are transcribed during egg production; their mRNAs lie dormant in the egg until it is fertilized determine the number of body segments that will form are expressed late and determine the detailed structure of each body segment generally have no introns are not translated into proteins Short Answer Questions 28 Principles of gene regulation Page: 1083 Difficulty: Usually, a mutation in the promoter region of an operon causes reduced levels of synthesis of the proteins encoded by that operon Occasionally, a mutation in the promoter region actually causes increased levels of synthesis Can you suggest (briefly) a plausible explanation? Ans: It is possible for a mutation to result in a promoter with higher-than-normal affinity for RNA polymerase (i.e., a promoter that more closely matches the promoter consensus sequence), resulting in increased expression in the mutant of the genes in this operon 29 Principles of gene regulation Page: 1084 Difficulty: Describe and contrast positive regulation and negative regulation of gene expression Ans: Positive regulation involves an activator protein that, when bound in the vicinity of the regulated gene, facilitates the binding of RNA polymerase to its promoter The affinity of the activator for the DNA may be either increased or decreased by a signal molecule Negative regulation involves a repressor protein that, when bound near the regulated gene, hinders access of RNA polymerase to its promoter or inhibits its activity Repressor affinity for its binding site (the operator) may be increased or decreased by a signal molecule (See Fig 28-4, p 1084.) 30 Principles of gene regulation Pages: 1085 Difficulty: Define operon and polycistronic mRNA Ans: An operon is several contiguous structural genes that are coordinately regulated by promoter Chapter 28 Regulation of Gene Expression 333 and operator regions, as depicted in Fig 28-5, p 1085 All of the genes in an operon are transcribed in a polycistronic mRNA—a single mRNA molecule encoding more than one structural gene Note: operons are not found in eukaryotic organisms 31 Principles of gene regulation Pages: 1088-1091 Difficulty: Match the protein or structural feature on the left with one appropriate description on the right activator helix-turn-helix leucine zipper repressor zinc finger (a) a positive regulator (b) a negative regulator (c) facilitates transcription only when bound to a signal molecule (d) a DNA-binding structural motif found in many prokaryotic regulatory proteins (e) a structural feature involved in protein-protein interactions between some regulatory protein monomers (f) a protein that dissociates from DNA when bound to a signal molecule (g) a DNA-binding structural motif found in many eukaryotic regulatory proteins Ans: a; d; e; b; g 32 Regulation of gene expression in prokaryotes Pages: 1085-1086, 1093-1094 Difficulty: E coli cells are placed in a growth medium containing lactose Indicate how the following circumstances would affect the expression of the lactose operon (increase/decrease/no change) (a) Addition of high levels of glucose (b) A Lac repressor mutation that prevents dissociation of Lac repressor from the operator (c) A mutation that inactivates β-galactosidase (d) A mutation that inactivates galactoside permease (e) A mutation that prevents binding of CRP to its binding site near the lac promoter Ans: (a) decrease; (b) decrease; (c) decrease (this enzyme converts lactose into allolactose, the inducer); (d) decrease (because external lactose would not enter the cell, and allolactose would not be present to induce the operon); (e) decrease 33 Regulation of gene expression in prokaryotes Pages: 1086, 1093-1094 Difficulty: Draw a simple map of the lactose operon indicating the relative positions of promoter, operator, CRPbinding site, repressor gene (I), and the structural genes of the operon (A, Y, Z) Indicate where the CRP protein binds within this operon When it is bound to this site, does the CRP protein have a positive or negative effect on gene expression in this system? Ans: Bound CRP stimulates (has a positive effect on) transcription of this operon (See Fig 28-7, p 1086, and Fig 28-17, p 1093.) 34 Regulation of gene expression in prokaryotes Pages: 1093-1094 Difficulty: Briefly explain (a) why there is a lag in cell growth when bacteria are switched from a medium containing glucose to one containing lactose (b) When the growth medium contains both lactose and glucose, what proteins will be bound to the lac operon regulatory region? (c) If only lactose is in the 334 Chapter 28 Regulation of Gene Expression growth medium, what proteins will be bound to the lac operon regulatory region? Ans: (a) The lag represents the time required for lactose to enter the cell, be converted into the inducer allolactose, and turn on the synthesis of the genes of the lac operon that are essential for growth on lactose (b) When both glucose and lactose are present, neither Lac repressor nor CRP will be bound (c) If only lactose is present, CRP will be bound (See Fig 28-18, p.1094.) 35 Regulation of gene expression in prokaryotes Pages: 1094-1097 Difficutly: In prokaryotes such as E coli, many operons that encode enzymes involved in amino acid biosynthesis begin with a sequence coding for a leader peptide This peptide has no known enzymatic function and is rich in the amino acid that is synthesized by the enzymes coded for in the operon What is the function of this leader peptide? Ans: The leader peptide is integral to the regulatory mechanism called transcription attenuation In prokaryotes, transcription and translation are closely coupled The ability to translate the leader peptide signals that the relevant amino acid is readily available in the cell and that transcription of the operon is undesirable at this time Translation through the leader peptide (when the amino acid is abundant) allows the formation of a hairpin attenuator structure downstream in the nascent RNA chain This attenuator acts as a transcription terminator, causing dissociation of the RNA polymerase When the relevant amino acid is in short supply, the ribosome stalls trying to synthesize the leader peptide A different secondary structure forms in the nascent RNA, preventing formation of the terminator and allowing continued transcription of the operon (See Fig 28-21, p.1096.) 36 Regulation of gene expression in prokaryotes Page: 1097-1098 Difficulty: The SOS response in E coli is triggered by extensive damage to the cell’s DNA and increases the capacity for repairing such DNA What molecular events bring about expression of the SOS genes? Ans: The SOS genes are located in several operons, all under the regulation of the LexA repressor, which normally represses these operons When DNA is damaged extensively, RecA protein binds to single-stranded DNA at the damaged sites, and bound RecA activates specific proteolysis of the LexA protein, relieving the repression of the SOS operons (See Fig 28-22, p 1097.) 37 Regulation of gene expression in prokaryotes Pages: 1098-1099 Difficulty: Explain how synthesis of ribosomal proteins in E coli is regulated at the level of translation Ans: Ribosomal proteins are encoded in several operons One protein encoded in each operon, when not part of an assembled ribosome, binds tightly to that operon’s mRNA, preventing its translation Thus, when an excess of that protein is present, it slows its own translation (See Fig 28-23, p 1099.) 38 Regulation of gene expression in eukaryotes Pages: 1102-1103 Difficulty: Describe briefly the relationship between chromatin structure and transcription in eukaryotes Ans: Heterochromatin is highly condensed and transcriptionally inert because the chromatin proteins make promoters inaccessible The less condensed euchromatin has undergone alteration of its structure (“remodeling”), allowing some regions to be transcribed These alterations include covalent modification (such as acetylation) of histones and displacement of nucleosomes, creating exposed regions of DNA (hypersensitive sites) that are probably binding sites for regulatory proteins Chapter 28 Regulation of Gene Expression 335 39 Regulation of gene expression in eukaryotes Pages: 1102-1104 Difficulty: Define each in about two sentences: (a) hypersensitive sites in eukaryotic chromosomes; (b) enhancers (upstream activator sequences); (c) chromatin remodeling Ans: (a) A hypersensitive site is a region of eukaryotic DNA that is being associated with actively transcribed DNA and is unusually sensitive to degradation by nucleases (b) Enhancers are DNA sequences that regulate the transcription of certain genes by allowing binding of specific transactivator proteins These sequences may occur thousands of base pairs away from the gene whose transcription they regulate (c) Chromatin remodeling reflects structural changes resulting from enzymes that covalently modify the core histone complex in nucleosomes (such as histone acetyltransferases) and those that move or displace nucleosomes using the energy of ATP hdyrolysis (See Table 28-2, p 1103.) 40 Regulation of gene expression in eukaryotes Page: 1104 Difficulty: Describe in one or two sentences the role of each of the following types of proteins in the regulation of gene expression in eukaryotes: (a) basal transcription factors; (b) transactivators; (c) coactivators Ans: (a) Basal transcription factors interact with specific DNA sequences and/or each other and/or with RNA polymerase to form a complex at promoters (b) Transactivators interact with both target sites on DNA and with other regulatory proteins such as coactivators to activate transcription (c) Coactivators interact with transactivators and proteins at the promoters to activate transcription 41 Regulation of gene expression in eukaryotes Pages: 1106-1108 Difficulty: DNA-binding transactivating proteins often possess a domain separate from their DNA-binding domains that serves as a docking site for interactions with the transcription complex, coactivators, corepressors, or even chromatin remodeling proteins, to regulate gene transcription Describe three known kinds of such domains, and provide an example of each Ans: (1) Gal4p (in yeast) has an acidic activation domain rich in aspartic and glutamic acid, (2) Sp1 in higher eukaryotes has two activation domains comprised of about 25% glutamine, and (3) CTF1 has a proline-rich (>20%) activation domain 42 Regulation of gene expression in eukaryotes Pages: 1108-1109 Difficulty: Describe briefly the process by which steroid hormones affect gene expression Ans: Steroid hormones usually enter a cell by diffusion and then bind to a nuclear receptor protein The binding changes the conformation of the receptor protein such that the hormone-receptor complex can bind to specific DNA sequences (hormone response elements) This binding can either activate or repress the expression of adjacent genes 43 Regulation of gene expression in eukaryotes Page: 1110 Difficulty: What are three mechanisms of translational repression that are known to exist in eukaryotes? Ans: (a) inactivation of initiation factors usually by phosphorylation; (b) binding of repressor proteins 336 Chapter 28 Regulation of Gene Expression to the mRNA, thereby interfering with initiation factors or the ribosome; (c) interference with elongation by the binding of proteins to elongation factors 44 Regulation of gene expression in eukaryotes Pages: 1110-1111 Difficulty: Large numbers of micro-RNAs (miRNAs), also known as small temporal RNAs (stRNAs) have now been discovered in higher eukaryotes Describe their characteristics and general function Ans: Their primary transcripts are about 70 nucleotides, with self-complementary internal sequences that can form hairpin structures These are cleaved into 20-25 nt partial duplexes, one strand of which can bind to complementary stretches in cellular mRNAs This can then inhibit gene expression by blocking translation of the target mRNA or facilitating its degradation 45 Regulation of gene expression in eukaryotes Pages: 1112-1116 Difficulty: Describe briefly the general role of the protein products of each of the following types of genes in the embryonic development of the Drosophila: (a) maternal genes; (b) segmentation genes; (c) homeotic genes Ans: (a) A maternal gene in Drosophila is a gene that is expressed in the unfertilized egg (see Fig 28-36, p 1114) These genes are involved in directing the spatial organization of the Drosophila embryo early in development (b) Segmentation genes direct the formation of the correct number of body segments, and include the gap genes, pair-rule genes, and segment polarity genes as subclasses (c) Homeotic genes help specific localization of organs and appendages in the segments of the embryo They regulate their target genes by highly conserved DNA-binding homeodomains, depicted in Fig 28-13, p 1090