E27. A cloned gene fragment contains a regulatory element that is recog- nized by a regulatory transcription factor. Previous experiments have shown that the presence of a hormone results in transcriptional acti- vation by this transcription factor. To study this effect, you conduct a electrophoretic mobility shift assay and obtain the following results: Tube: 1 2 3 Transcription factor: Hormone: Explain the action of the hormone.
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- . An interesting mutation in lacI results in repressorswith 110-fold increased binding to both operator andnonoperator DNA. These repressors display a “reverse”induction curve, allowing β-galactosidase synthesis inthe absence of an inducer (IPTG) but partly repressingβ-galactosidase expression in the presence of IPTG. Howcan you explain this? (Note that, when IPTG binds a repressor, it does not completely destroy operator affinity,but rather it reduces affinity 110-fold. Additionally, ascells divide and new operators are generated by thesynthesis of daughter strands, the repressor must findthe new operators by searching along the DNA, rapidlybinding to nonoperator sequences and dissociating fromthem.). One mechanism by which antisense RNAs act as negative regulators of gene expression is by base pairingwith the ribosome binding site on the sense mRNA toblock translation. In a second, alternative mechanism,the act of transcribing an antisense RNA can somehow prevent RNA polymerase from recognizing thesense promoter for the same gene. Design an experimental approach that would enable you to distinguishbetween these two modes of action at a specific gene.(Hint: What would be the outcome in each case ifhigh levels of the antisense RNA were transcribedfrom a gene on a plasmid?)Enhancer RPA Gene A Use the diagram above, which depicts a chromosomal region less than 500,000 bp, to determine the expression of each gene under the scenarios specified in the table. Regulatory promoters are indicated by "RP". Genes A, B, and C have intermediate expression when no regulatory proteins are bound. Complete the table by choosing high, intermediate, or low to describe the expression level of each gene under each specified scenario (i and ii). No regulatory proteins bound (i) A repressor binds to RPA and an activator binds to RPC (ii) A regulatory protein binds at each of the activator, insulator, and silencer. Expression of A? Intermediate 1. [Select] 4. [Select] Insulator RPB Expression of B? 2. Gene B Intermediate 5. [Select] [Select] Silencer RPC Expression of C? 3. Gene C Intermediate 6. [Select] [Select]
- Discuss the following argument: “if the expression of every gene depends on a set of transcription regulators, then the expression of these regulators must also depend on the expression of other regulators, and their expression must depend on the expression of still other regulators, and so on. cells would therefore need an infinite number of genes, most of which would code for transcription regulators.” how does the cell get by without having to achieve the impossible?Explain in detail the role of super-enhancers. Give some examples. How the super-enhancers can be identified?Wilms tumor 1, or nephroblastoma, is caused by mutations in the WT1 gene, which encodes a transcription factor. You have identified a novel variant in WT1: Arg422Pro. You have control cells and cells that have been engineered to carry the homozygous WT1 p.Arg422Pro mutation. You want to assess effects of this mutation on a variety of endpoints. For each endpoint listed below, choose the one technique is best suited to answer the question. Choose from: array CGH, qRT-PCR, qPCR, RNA-seq, FISH, in situ hybridization, western blot, immunostaining, WT1 ChIP-seq, WT1 ChIP-PCR, ATAC-seq, 3C Endpoint Technique? WT1 protein amount (quantitative) Western blot WT1 protein binding to all enhancers, genome-wide Chip-seq WT1 mRNA amount (quantitative) WT1 protein subcellular localization Quantitative assessment of all mRNAs in these cells (genome-wide) RNAseq Chromatin interactions between a specific WT1 chromatin binding site (identified above)…
- Describe in detail the role of super-enhancers. Give some examples. How the super-enhancers can be identified?Many currently marketed drugs exert their pharmacological effects by binding to ligand-activated transcription factors and modulating gene expression. One example, are various drugs that target the estrogen receptor to treat breast cancer, osteoporosis and post-menopausal symptoms. Below is a ChIP experiment examining the effects of no drug treatment (C), the natural hormone estrogen (E) and the drugs tamoxifen (T) and raloxifene (R) on recruitment of coactivators (SRC-1 and CBP), Histone Deacetylase Complexes (HDACs) and acetylation of histones associated with the C-myc gene. Which of the following statements are correct based on this data (select all that apply)?The binding of a small effector molecule, protein-protein interactions, and covalent modifications are three common ways to modulate the activities of transcription factors. Which of these three mechanisms are used by steroid receptors and by the CREB protein?
- Describe the various post-translational modifications of HIF- 1alpha and how it affects the regulation of HIF-1al pha signaling. How might HIF- alpha alter the tumor microenvironment to promote tumor growth? Propose a strategy to prevent HIF-alpha signaling in the TME. What do you think would happen in a transgenic mouse with a total knockout of HIF-alpha?This is the full question.. "Both the cytokinin receptor encoded by CRE1 and the ethylene receptor encoded by ETR1 are examples of" Both the cytokinin receptor encoded by CRE1 and the ethylene receptor encoded by ETR1 are examples of O Leucine-rich repeat receptor kinase • The response regulator component of a two-component sensor histidine kinase (i.e., the component that directly activates transcription (type B ARRS, in the case of cytokinin) or directly mediates a response (type A ARRS)) O Regulatory molecules that bind calcium O The sensor histidine kinase component of a two-component sensor histidine kinase (i.e., the part that receives the signal and passes it to other components of the signal transduction cascade) O Proteins that shuttle from the cytoplasm to the nucleus to alter gene expressionDiscuss how the expression of a protein can be regulated post transcription in eukaryotic cells through, using the following key terms: Degradation of mRNA (two ways) Blocking translation Degradation of the protein