Ch 15

Which protein complex directs DNA bending into loops that contact RNA polymerase and transcription factors bound at the core promoter or with protein complexes bound to proximal promoter elements?

For the following gene, which mutant likely has lost its enhancer? Mutant Deletion Region % Transcription WT None 100% 1 1-200 150% 2 250-400 100% 3 500-800 50% 4 950-1100 0%

Many types of cancer are known to overexpress the receptor protein tyrosine kinase. Which molecular technique can be used to reduce expression of an oncogene in vitro? Southern blotting western blotting RNAi DNAse sensitivity assay PCR

Which protein complex binds dsRNA fragments to generate ssRNAs for RNAi?

Demethylation and acetylation lead to open chromatin structure and are associated with ________ regions of genomes.

You have identified a mutation in a gene which also seems to decrease transcription of another gene 2000 bp away from the mutation site. What regulatory sequence, which may be found within another gene, has likely been mutated in this instance? homeodomain motif upstream activator sequence enhancer sequence core promoter proximal elements

In yeast, if you want to prevent the Gal4 regulatory protein from binding near each of the GAL genes, which sequence element would you target? proximal elements core promoter homeodomain motif enhancer sequence upstream activator sequence

In yeast in the presence of galactose, you unexpectedly find that transcription is still blocked. Assuming Gal80 is present and functional, which other protein may be mutated to prevent transcriptional activation? Gal10 Gal2 Gal4 Mig1 Gal3

Galactose is absent, but you find active transcription of the GAL genes. Which protein is likely mutated or absent to allow for constitutive transcription of the GAL genes? Mig1 Gal80 Gal4 Gal10 Gal2

Prader-Willi syndrome is a genetic disorder involving a partial deletion of chromosome 15q on the paternal chromosome. When both copies of a gene (or chromosome) are functional but only one is expressed, this is an example of X inactivation. histone acetylation. position effect variegation. chromatin modifications. genomic imprinting.

During RNAi, what do miRNAs target for destruction? heterochromatic regions of DNA mRNAs CpG islands ribosomes histones

Compare and contrast promoters and enhancers with respect to their location (upstream versus downstream), orientation, and distance (in base pairs) relative to a gene they regulate. promoter sequence is likely to be: Check all that apply. located within a few dozens nucleotides of the gene it controls orientation dependent orientation independent located upstream of the gene it controls located downstream of the gene it controls located either nearby or at great distance from the gene it controls

Compare and contrast promoters and enhancers with respect to their location (upstream versus downstream), orientation, and distance (in base pairs) relative to a gene they regulate. enhancer sequence is likely to be: Check all that apply. located downstream of the gene it controls orientation independent located whitin a few dozens nucleotides of the gene it controls located upstream of the gene it controls located either nearby or at great distance from the gene it controls orientation dependent

A .....is a DNA sequence where RNA polymerase binds and begins the process of transcription.

A .... is a DNA sequence that binds regulatory proteins that interact with promoter-bound proteins to activate transcription.

A...is a DNA sequence that binds regulatory proteins that inhibit transcription.

... is the protein complex that is part of the RNA interference (RNAi) mechanism. It denatures short double-stranded RNAs to single strands that carry out RNAi.

...is the enzyme complex that is active in RNAi, where it cuts double-stranded regulatory RNAs into 21-bp to 26-bp segments that are subsequently denatured by RISC.

In Drosophila development, the effects of BICOID protein vary along the axis of the embryo. This is because _______. Hints BICOID mRNA exists in the egg prior to fertilization there is an anterior to posterior gradient in BICOID protein concentration in the embryo BICOID both activates and represses gene activity BICOID is a transcription factor

A set of known mutations in the BX-C genes in Drosophila results in the development of wings on a body segment that would not ordinarily have wings. This is an example of mutation in which class of developmental genes? Segmentation genes Maternal effect genes Lethal genes Homeotic genes

Maternal effect genes are transcribed in the egg after fertilization. Hints True False

The regulation of gene expression in individual cells coordinates the development of multicellular organisms, ensuring that tissues and organs form in their characteristic places. Researchers can study how multicellular organisms develop their spatial organization by examining normal and mutant forms of organisms, such as the fruit fly, Drosophila. Which of the following statements about pattern formation are true? Select the four statements that are true. Hints Differential gene expression affects the developmental process in animals. Pattern formation begins in adult animals. Cells receive molecular signals that communicate their position in relation to other cells. Homeotic genes establish major body axes. Homeotic genes code for transcription factors that control the development of segment-specific body parts. Positional information controls pattern formation. The process of pattern formation involves segmentation of the body but does not involve the development of segment-specific body parts.

How is positional information provided along the anterior-posterior axis in Drosophila? What are the functions of bicoid and nanos?

How are the sharp boundaries of expression of eve stripe 2 formed? hunchback and bicoid are present and giant and Krüppel absent only in the region corresponding to stripe 2; therefore, eve expression is limited to this region. giant and Krüppel are present and hunchback and bicoid absent in all regions except for the region corresponding to stripe 2; therefore, eve expression is limited to this region. hunchback and bicoid are present and giant and Krüppel absent in all regions except for the region corresponding to stripe 2; therefore, eve expression is blocked in this region. giant and Krüppel are present and hunchback and bicoid absent only in the region corresponding to stripe 2; therefore, eve expression is blocked in this region.

Consider the binding sites for gap proteins and bicoid in the stripe 2 enhancer module. What sites are occupied in parasegments 2, 3, and 4, and how does this result in expression or no expression?

What do you expect to see happen to even-skipped stripe 2 if it is expressed in a Krüppel mutant background? In a Krüppel mutant, hunchback and bicoid are bound in parasegment 3 but Krüppel is not, resulting in expression of eve in both parasegments 3 and 4. In a Krüppel mutant, giant is bound in parasegment 3 but Krüppel and hunchback are not, resulting in expression of eve in both parasegments 3 and 4. In a Krüppel mutant, giant is bound in parasegment 4 but Krüppel and hunchback are not, resulting in expression of eve in both parasegments 3 and 4. In a Krüppel mutant, hunchback and bicoid are bound in parasegment 4 but Krüppel is not, resulting in expression of eve in both parasegments 3 and 4.

What do you expect to see happen to even-skipped stripe 2 if it is expressed in a giant mutant background? In a giant mutant background, hunchback and Krüppel are bound to their sites in parasegment 3 but giant is not, resulting in eve expression in both parasegments 2 and 3. In a giant mutant background, hunchback and bicoid are bound to their sites in parasegment 3 but giant is not, resulting in eve expression in both parasegments 2 and 3. In a giant mutant background, hunchback and Krüppel are bound to their sites in parasegment 2 but giant is not, resulting in eve expression in both parasegments 2 and 3. In a giant mutant background, hunchback and bicoid are bound to their sites in parasegment 2 but giant is not, resulting in eve expression in both parasegments 2 and 3.

What do you expect to see happen to even-skipped stripe 2 if it is expressed in a hunchback mutant background? In hunchback mutants, expression in parasegment 4 is diminished. In hunchback mutants, expression in parasegment 1 is diminished. In hunchback mutants, expression in parasegment 2 is diminished. In hunchback mutants, expression in parasegment 3 is diminished.

What do you expect to see happen to even-skipped stripe 2 if it is expressed in a bicoid mutant background? In bicoid mutants, expression in parasegment 1 is diminished. In bicoid mutants, expression in parasegment 3 is diminished. In bicoid mutants, expression in parasegment 4 is diminished. In bicoid mutants, expression in parasegment 2 is diminished.

What is meant by the term chromatin remodeling? Select the two correct answers. Chromatin remodeling includes altering chromatin structure by lncRNAs acting as scaffolds linking chromatin regulatory proteins. Chromatin remodeling includes altering chromatin structure by altering the positioning of nucleosomes with respect to specific DNA sequences. Chromatin remodeling includes altering chromatin structure by genomic imprinting. Chromatin remodeling includes altering chromatin structure by altering the composition of histones within nucleosomes. Chromatin remodeling includes altering chromatin structure by altering the extent of DNA supercoiling.

Describe the importance of this process to transcription. Select the four correct answers. These alterations can be used to repress transcription of a gene by hiding its enhancer sequences. These alterations can be used to activate transcription of a gene by opening its promoter sequences. These alterations can be used to repress transcription of a gene by hiding its promoter sequences. These alterations can be used to activate transcription of a gene by hiding its promoter sequences. These alterations can be used to repress transcription of a gene by opening its promoter sequences. These alterations can be used to activate transcription of a gene by hiding its enhancer sequences. These alterations can be used to repress transcription of a gene by opening its enhancer sequences. These alterations can be used to activate transcription of a gene by opening its enhancer sequences.

What general role does acetylation of histone protein amino acids play in the transcription of eukaryotic genes? Histone acetylation events are most often associated with transcription termination. Histone acetylation events are most often associated with transcription repression. Histone acetylation events are most often associated with transcription proofreading. Histone acetylation events are most often associated with transcription activation.

Most biologists argue that the regulation of gene expression is considerably more complex in eukaryotes than in bacteria. List the four factors that in your view make the largest contribution to this perception. Select the four correct statements. The extensive use of attenuation allows eukaryotes to fine-tune transcriptional activity of a cell according to environmental signals. The coordinated regulation of sets of genes devoted to a single function (e.g. metabolic pathways) allows minimal wasting of a cell's resources on unneeded proteins and metabolites. Posttranslational processing of proteins allows cells to regulate the function of proteins after their synthesis by covalent modification, transport, or both. The chromatin structure of eukaryotic genomes allows the differential packaging of genes to be used in regulation. Eukaryotic cells possess a nucleus, which separates transcription from translation and allows RNA processing and transport to be used in regulation. Multicellularity and cell-type specificity is frequent in eukaryotes and requires organisms to regulate large suites of genes in a cell-type-specific manner.

A gene expressed in long muscle of the mouse is identified, and the regulatory region upstream of the gene is isolated. Various segments of the upstream sequence are fused to the lacZ gene, and each fusion is assayed to determine how efficiently it transcribes the gene. In the accompanying diagram, the dark bars indicate the upstream segments that are present in each of six different fusion genes. The transcriptional efficiency of each fusion is measured against the control fusion, that is, the full-length upstream segment fused to the lacZ gene. Part A Identify the upstream region that contains the main enhancer. 1 2 3 4 5 6 7 8

Identify the upstream region containing the promoter. 1 2 3 4 5 6 7 8

A muscle enzyme called ME1 is produced by transcription and translation of the ME1 gene in several muscles during mouse development, including heart muscle, in a highly regulated manner. Production of ME1 appears to be turned on and turned off at different times during development. To test the possible role of enhancers and silencers in ME1 transcription, a biologist creates a recombinant genetic system that fuses the ME1 promoter, along with DNA that is upstream of the promoter, to the bacterial lacZ(?-galactosidase) gene. The lacZ gene is chosen for the ease and simplicity of assaying production of the encoded enzyme. The diagram shows the structure of the recombinant, as well as bars that indicate the extent of six deletions the biologist makes to the ME1 promoter and upstream sequences. The blue bar is the site of the promoter whereas the gray bars span potential enhancer/silencer modules. The table displays the percentage of ?-galactosidase activity in each deletion mutant in comparison to the recombinant gene system without any deletions. Part A Does this information indicate the presence of enhancer and/or silencer sequences in the ME1 upstream sequence? yes no

Where is the enhancer sequence located? 1 2 3 4 5 6 7 8 9

Where is the silencer sequence located? 1 2 3 4 5 6 7 8 9

Part D Why does deletion D effectively eliminate transcription of lacZ? the deletion removes the promoter the deletion removes a silencer the deletion removes an enhancer element the deletion removes a part of coding region

Part E Given the information available from deletion analysis, can you give a molecular explanation for the observation that ME1 expression appears to turn on and turn off at various times during normal mouse development? Expression of ME1 is regulated by an enhancer and a silencer, which can be regulated such that ME1 expression is activated at some developmental stages and then repressed at others. Expression of ME1 is regulated by a promoter and an inhibitor, which can be regulated such that ME1 expression is activated at some developmental stages and then repressed at others. Expression of ME1 is a non-regulated process which is turned on and turned off randomly. None of the above.

Which of the following best explains why many developmental genes encode either transcription factors or signaling molecules? Transcriptional factors and signal-transducing proteins usually contain many different domains and thus present suitable raw material for developmental evolution. Cell differentiation is a fundamental aspect of development and results from changes in gene expression. Thus, genes that control development do so by controlling gene expression through the regulation of transcription. The complexity of multicellular organisms requires that most of their genes are involved in regulatory processes such as regulation of expression. Therefore genes that control the expression of other genes would comprise a major fraction of any particular group of genes, including those associated with development. The functions of developmental genes are extremely complicated because they stand in the very center of the complex regulatory network that organizes all cellular processes. Transcription factors and signaling molecules are the most readily detectable functions though not necessarily the most frequent ones.

Actinomycin D is a drug that inhibits the activity of RNA polymerase II. In the presence of actinomycin D, early development in many vertebrate species, such as frogs, can proceed past the formation of a blastula, a hollow ball of cells that forms after early cleavage divisions; but development ceases before gastrulation. Part A What does this tell you about maternal versus zygotic gene activity in early frog development? Check all that apply. Development beyond the gastrulation must be largely under control of maternal effect genes. Zygotic gene expression is required for gastrulation and beyond. Development from fertilization until gastrulation must be largely under control of maternal effect genes. Zygotic gene expression must be largely under control of maternal effect genes.

bicoid is a coordinate, maternal effect gene. Part A A female Drosophila heterozygous for a loss-of-function bicoid allele is mated to a male that is heterozygous for the same allele. What are the phenotypes of their progeny? One-fourth of the progeny will be homozygous bicoid mutants, and the males with this genotype will be sterile. Half of the progeny will be homozygous bicoid mutants and the females with this genotype will be sterile. Half of the progeny will be homozygous bicoid mutants, and the males with this genotype will be sterile. One-fourth of the progeny will be homozygous bicoid mutants and the females with this genotype will be sterile.

Correct Part B A female that is homozygous for a loss-of-function bicoid allele is mated to a wild-type male. What are the phenotypes of their progeny? The homozygous bicoid female will produce all defective eggs, which develop into headless embryos that do not complete development. One-fourth of the progeny will be homozygous bicoid mutants, and the females with this genotype will be sterile. All progeny will be heterozygous bicoid mutants, and therefore their progeny will have normal phenotype. All progeny will be heterozygous bicoid mutants, and the females with this genotype will be sterile.

Correct Part C If loss of bicoid function in the egg leads to lethality during embryogenesis, how are females homozygous for bicoid produced? Homozygous bicoid mutant females (and males) can be produced by crossing male and female +/bicoid heterozygotes. Homozygous bicoid mutant females (and males) can be produced by crossing male and female +/bicoid heterozygotes. In order to develop normally, the eggs resulting from this cross should be supplied with normal bicoid protein. This can be achieved by introducing a plasmid, which provides for transient expression of bicoid protein. Homozygous bicoid mutant females (and males) can be produced only as the result of genetic engineering. Homozygous bicoid mutant females (and males) can be produced only as the result of extensive mutagenesis.

Correct Part D What is the phenotype of a male homozygous for bicoid loss-of-function alleles? Homozygous bicoid mutant males that were produced by a female heterozygous for bicoid will have progeny that will develop into headless embryos that do not complete development. Homozygous bicoid mutant males that were produced by a female heterozygous for bicoid will develop into headless embryos that do not complete development. Homozygous bicoid mutant males that were produced by a female heterozygous for bicoid will be phenotypically normal. Homozygous bicoid mutant males that were produced by a female heterozygous for bicoid will be sterile.