11 - Gene Regulation

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11 - Gene Regulation
2014-08-22 21:05:08
gene regulation
Gene Regulation
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  1. What are constitutive enzymes?
    Enzymes that are synthesized at a more or less constant rate at all times; house-keeping enzymes
  2. What are inducible enzymes?
    Enzymes which are synthesized depending on need and are synthesized in response to cellular or environmental cues; bacterial genes encoding inducible enzymes involved in the synthesis of a common end product are frequently organized into an operon
  3. What is positive regulation of a gene?
    The transcription of a gene is quantitatively increased by the presence of a specific activator protein (or by the inactivation of a specific repressor protein)
  4. What is negative regulation of a gene?
    Transcription of a gene is quantitatively decreased by the presence of a repressor (or by the inactivation of an activator)
  5. What does Cis-acting mean?
    Refers to a sequence of DNA that affects the activity of adjacent sequences; this DNA does not encode a protein
  6. What does trans-acting mean?
    Refers to a diffusible product (protein) that affects the activity of DNA
  7. When is the Lac operon activated?
    When no glucose is available; bacteria prefers to use glucose
  8. What is the lacI gene?
    It is the lac repressor protein that binds to the operator of the lac operon to constitutively repress the operon when glucose is plentiful in the medium
  9. Where is the operator for the Lac operon located?
    It is located between the promoter and the transcription initiation site for the lacZ gene
  10. What is the operon comprised of?

    Where I encodes for the repressor; Z codes for β-galactosidase; Y codes for permease; A codes for transacetylase
  11. How does the Lac operon turn on?
    When lactose enters the cell, β-galactosidase turns lactose into allolactose; allolactose binds the repressor protein on the operator DNA, thus allowing the transcription of genes in the operon; now the three enzymes needed for lactose metabolism are being produced
  12. How is the Lac operon regulated by a transcription activator?
    When glucose levels are low, cAMP levels increase, as these levels increase, it allows cAMP to bind to CAP, which changes the conformation of CAP allowing it to bind to the Lac promoter; once the CAP binds to the Lac promoter it increases the binding strength of the promoter to RNA polymerase
  13. What are the three functional domains of CAP?
    Ligand binding domain (allows it to bind to cAMP), DNA binding domain (allows it to bind to the Lac promoter), and protein-protein interaction domain (allows CAP to dimerize)
  14. What are the functional subdomains of the lac repressor protein?
    The A-terminus contains the binding domain; the C-terminus contains the protein-protein binding domain and the inducer-binding domain; the repressor binds as a dimer to a palindromic sequence of the operator DNA
  15. What is the tryptophan operon required for?
    The synthesis of tryptophan; it works through negative regulation by repressor binding and by attenuation
  16. What is attenuation?
    Amino acid levels activate or repress transcription of the corresponding amino acid synthesis operon by modifying the RNA structure of the leader RNA in the 5’ end of the polycistronic mRNA
  17. How does the repressor stop Trp production?
    When there are high levels of Trp, Trp binds to the repressor, and together they bind to the operator; at this point RNA pol cannot move past the repressor molecule and no transcription occurs; if Trp is lacking in the media, repressor does not bind to the operator and transcription begins
  18. How is the tryptophan operon attenuated?
    Attenuation occurs early in transcription of the leader region of the Trp operon; transcription stops if a terminating stem loop is formed in the RNA by complementary base pairing; a secondary structure can occur, the hairpin anti-terminator; if the anti-terminator forms the termination stem loop cannot form so transcription continues; which stem loop forms is determined by the speed through which the rsome goes through the region; if Trp is lacking the rsome stalls at Trp codons allowing formation of the anti-terminator
  19. What other operons are regulated by attenuation?
    Threonine, phenylalanine, and histidine; all contain multiple, respective codons in their leader sequences
  20. What are the two phases for a λ phage?
    Infective phase and dormant phase
  21. How does the λ repressor self-regulate?
    When the repressor binds to OR1, it prevents RNA Pol from binding to the promoter sequence for cro; this recruits another repressor to bind to OR2 which has lower affinity than OR1; this 2nd repressor recruits RNA Pol to stimulate transcription of the cI gene; as the level of λ repressor increases, a 3rd repressor binds to OR3, which inhibits further transcription of the cI gene; as the level of λ repressor drops, OR3 becomes unoccupied and the cI gene is activated to synthesize more repressor to maintain equilibrium in OR1,2,3
  22. What does an insulator do?
    It prevents the enhancer of a gene domain from activating the genes in the neighboring domains and also protects the resident gene from inadvertent transcriptional activation by the heterologous enhancers in the neighboring domains
  23. What modular, functional domains are found in eukaryotes?
    The ligand binding domain (binds inducers, hormones, etc.), the activating domain (binds to transcription factors and RNAP II to activate transcription), DNA binding domain (binds to specific DNA motifs in the promoters, enhancers and LCRs; helix-turn-helix, helix-loop-helix, leucine zipper, zinc finger, and more)
  24. What is helix-turn-helix?
    This is a conserved DNA-binding domain; it contains two α-helices, separated by a tight turn; it interacts with DNA in in its major groove
  25. What is helix-loop-helix?
    It is a dimeric transcription factor containing α-helices with hydrophobic residues; the residues are alternatively spaced so that the side chain of each residue is positioned toward one side of the helix to form a hydrophobic interacting surface; the α-helical monomers dimerize through looped helices
  26. What is the basic-leucine zipper?
    It is a dimeric transcription factor containing α-helices with leucine residues; the Leu residues are alternatively spaced so there is a Leu at every 7th residue; this forms a hydrophobic interacting surface; the α-helical monomers dimerize through strand interdigitation
  27. What is the zinc finger motif?
    It is ~30 AAs that are held in a ‘finger’ arrangement through Zn coordination; can serve as a hormone receptor, where the hormone binds to its cognate DNA sequence and stimulates transcription
  28. How can steroid hormones mix and match?
    You can have homo-dimers (class I) and hetero-dimers (class II); in hetero-dimers you can have the simultaneous turning off of one gene and turning on of another gene in response to the same hormone; dys-regulation of hormone receptors can cause many diseases
  29. What are coregulators?
    Do not bind DNA, recruited by transcription factors and interact with basal promoter complex assembled at TATA box; can either be co-activators or co-repressors
  30. How does Tamoxifen work?
    It competes with estrogen for binding the estrogen receptor; it prevents binding of coactivators and inhibits the ability of the estrogen receptor to activate expression of the responsive genes; breast cancers relying on estrogen-mediated pathways for growth and survival are suppressed
  31. What are the sites of epigenetic modification on histones?
    The amino terminal tails of histones sticking out from the nucleosome
  32. How does remodeling of chromatin structure to allow transcription factor binding?
    • 1. A transcription factor binds to its cognate site in the promoter or enhancer of a gene packaged in nucleosomes
    • 2. The bound transcription factor recruits a
    • co-activator with histone acetyltransferase (HAT) activity
    • 3. Co-activator with HAT activity acetylates lysine residues in the histone tails; acetylated histones recruit the chromatin
    • remodeling protein complex with ATPase activity
    • 4. ATP-dependent chromatin remodeling exposes DNA binding sites for other transcription factors including RNA pol II to bind and activate transcription of the gene
    • 5. Histone deacetylases (HDACs) associated with the co-repressors deacetylate the histones, close down the chromatin structure and turn off transcription
  33. What does acetylation and methylation of histones mean?
    Acetylated histones: active genes; deacetylated histones: inactive genes; methylated histones: mostly inactive genes
  34. How does methylation of DNA inactivate genes?
    Methylated DNA recruits histone deacetylases (HDACs) and other co-repressor proteins to close down the chromatin structure and repress transcription
  35. What are imprinted genes?
    Genes involved in fetal and placental growth exhibit parent-of origin dependent expression as a result of genomic imprinting; in the fetus, genes favoring fetal growth are expressed only when they are inherited from the father, and the allelic maternal genes are imprinted and inactivated; genes suppressing fetal growth are expressed only when they are inherited from the mother; the allelic paternal genes are imprinted and inactivated
  36. What is Beckwith-Wiedemann Syndrome and how does it occur?
    In normal, maternal csome 11, IGF2 gene is inactive and hyper-methylated; because the enhancer is blocked by the DNA/CTCF insulator protein complex between IGF2 and H19; in the father the insulator DNA is methylated and cannot bind CTCF (insulator protein) rendering the insulator non-functional, IGF2 is activated, while H19 is suppressed; in BWS, on the maternal csome the barrier-insulator DNA is mutated or deleted, so that IGF2 is activated by the 3’ enhancer, which leads to a double dosage of IGF2; this causes organ overgrowths, mental retardation, and susceptibility to cancer
  37. What do dysregulations of transcription factors cause?
  38. What is Wilms’ Tumor (WT)?
    It is a childhood malignant kidney cancer resulting from mutations in the WT1 gene; WT1 gene encodes a zinc finger transcription factor that regulates kidney and genitourinary formation during embryonic development; Mutations in the zinc fingers and some splice isoforms that affect the DNA binding strengths of the zinc fingers initiate the disease
  39. What is Burkitt’s lymphoma?
    In this C-myc is normally expressed at very low levels in B lymphocyctes, but the translocated c-myc gene is activated by a powerful enhancer (IgH) which activates the c-myc genes in B-cells; this causes abnormal B-cell proliferation and B-cell lymphoma
  40. How does HIV infection weaken immune cells?
    It hijacks host transcription factors and co-factors; the HIV RNA genome is copied into the DNA sequence of the host csome; the HIV contains strong binding sites for lymphoid transcription factors which deprives transcription of host genes
  41. What are the two major groups of non-coding RNAs in eukaryotes?
    Small non-coding RNAs (miRNAs and siRNAs), and long non-coding RNAs (lncRNAs)
  42. What are small non-coding RNAs?
    miRNAs and siRNAs that act post-transcriptionally to regulate mRNA stability and translation of mRNAs
  43. What are long non-coding RNAs?
    Regulate diverse cellular processes in development, differentiation, and malignancy
  44. What is RNA interference (RNAi)?
    It is post-transcriptional silencing of mRNA by siRNA and miRNA; siRNA and miRNA contain specific RNA sequences that base-pair with the target mRNAs to cause degradation or translational blockage; can be used to knockdown/silence specific mutated genes that cause human diseases; gene-specific drugs can avoid generating the undesirable side effects of many currently used, non-gene specific drugs