Lecture 24: Regulation of Eukaryotic Gene Expression

Question 1

Eukaryotic vs. prokaryotic genes

Answer 1

• eukaryotic genes have more regulatory binding sites (and regulatory proteins) than prokaryotes
  *

Question 2

Methods of transcriptional regulation

Answer 2

1. alternative promoters
   
   • may be tissue specific
   • one promoter leads to differing exons
2. transcripition factor regulation
   
   • binding of hormones/receptors, growth factors to promoter elements of genome (DNA/RNA)
   • eg phosphorylation
     
     • Signal Transducers and activators of transcription (STAT)
     • eg JAK-STAT:
       
       1. JAK is a kinase - phosphorylates tyrosine on STAT creates dimer -
       2. dimerization allows passage through nuclear pores -
       3. binds to DNA and activates transcription
3. Tissue-specific expression
   
   • eg homeobox gene expression in C. elegans

Question 3

regulatory proteins

Answer 3

• activators
  
  • bind to enhancers
    
    • position varies (1kb up/downstream) or within gene (Intron)
    • orientation independent
      
      • promoters depend on orientation
  • increase rate of transcription
  • generally function through recruitment
• Repressors
  
  • bind to silencers
  • decrease rate of transcription
  • many different mechanisms

Question 4

Insulators

Answer 4

• induces formation of heterochromatin to create a boundary between euchromatin -
  
  • prevents inappropriate activation of adjacent genes by enhancers
    *

Question 5

activator

Answer 5

• bind to enhancers
• position varies (1kb up/downstream) or within gene (Intron)
• orientation independent
• promoters depend on orientation
• increase rate of transcription generally function through recruitment
  
  • recruit polymerase indirectly
    
    • recruit mediator complex,
    • recruit TFIID complex
    • some activators recruit nucleosome modifiers

Question 6

types of regulator binding domains

Answer 6

• helix turn helix
• homeodomain
• zinc finger

Question 7

helix turn helix motif (regulator binding)

Answer 7

• most common motif in prokaryotic primary proteins
• third helix interacts through polar side chains with the major groove of DNA
• flexible loop separates two dimerization helices
• minor interaction with phosphate backbone
• region of helix in major groove DNA - rich in basic R and K residues

Question 8

Homeodomain

Answer 8

• many bind as heterodimers
• AA from helix 3 H-bonds in major groove
• helix 1 also h-bonds
• usually leads to body segmentation
  
  • transcription factors recognize different body segments
    *

Question 9

Zinc finger

Answer 9

• 2 beta strands and a single alpha helix
• stabilized by
  
  • hydrophobic core
  • coordiation of a zinc ion by 2 cysteines or 2 histidines
• occur in multiples (2 to >5)
  
  • connected by flexible peptide linkage
    *

Question 10

combinatorial regulation

Answer 10

• when two trnascription factors are required to activate each gene
• cooperative/synergistic activation:
  
  • synergy: (greater than additive)
• heterodimerization:
  
  • eg. Jun can form homodimers and heterodimers
    
    • heterodimers 10X higher binding affinity
  • eg. Fos cannot bind DNA by itself

Question 11

eukaryotic repressors: ways they work

Answer 11

1. compete with activators for DNA binding
2. bind to activators, inhibiting activation
3. interact with transciptional machinery and actively repress transcription
4. deacetylate histones or recruit deacetylases

Question 12

histone code hypothesis

Answer 12

histone flexible tails can be modified

• modifications influence packing of chromatin
  
  • done by interacting with different proteins
  • modifications can change
  • associated with protein structure and gene expression
  • proteins detect modifications
    
    • eg chromodomain
    • eg bromodomain
• examples
  
  • DNA methylation: silences genes
    
    • eg CGCGCGCG sequences causes methylation
    • methylation recruits packing proteins
    • 5-methyl of cytosine:
  • deacetylated: inactivated
  • acetylated: activated/open configuration