gene regulation

• constitutive gene expression (housekeeping): expression needed at all times in virtually all cells

  • usually negative feedback
  • homeostasis
• regulated gene expression: expression rises or falls according to cellular needs/signals

  • usually positive feedback
  • basal level -> constant
  • induction -> turned on
  • repression -> turned off
• gene regulation: prokaryotes

  • gene expression regulated primarily at the level of initiation
  • due to:
  • short mRNA 1/2 lives (minutes)
  • co-transcription/-translation (can start translation before transcription ends)
  • operons
  • no histones (no compaction/uncompaction)
  • no nucleus
• gene regulation: eukaryotes

  • more complex regulation requires; occurs at multiple levels

  1. chromatin state
  2. transcription factor binding (recruiting RNAP)
  3. transcript processing -> spliced in nucleus
  4. translation initiation/mRNA stability
• prokaryotic promotor consensus sequences

  • level of initiation
  • expression controlled by interactions of promotor elements with RNA polymerase and specific repressors/activators (eg sigma factors)
  • sigma factors are released after about +10 nt transcribed
  • prokaryotic promotor elements:
  • UP element (-40 to -60; A-T rich) strongly stimulates transcription.
  • TATAAT (-10) and TTGACAT (-35) regions. Help define transcription start and strand usage
  • all is designed to get RNA polymerase to start copying/making DNA
• prokaryotic activators and repressors

  • negative regulation
  • repressor binds operator at or near promotor to block transcription
  • when repressor leaves the signaling molecule falls off
  • positive regulation
  • activator binds adjacent to promotor to enhance transcription
• lactose operon (prokaryotes)

  • operon: several protein-producing genes are linked together under the control of a single promotor
  • lactose is present -> genes turned on
  • lactose is gone -> genes turned off
  • LacI, LacZ, LacY, LacA
  • CAP
• LacI: repressor
• LacZ: β-galactosidase

  lactose -> glucose + galactose

  first encoded protein in operon
• LacY: permease

  lactose -> cellular uptake
• LacA: transacetylase

  acetylates β-galactosidase
• cap: catabolite activator protein
• LacI -> CAP -> P -> O -> LacZ -> LacY -> LacA
• lac operon expression not needed: Lac repressor binds to operator, σ70-Pol can't bind promotor -> repression

  • -lactose, +glucose (low cAMP)
  • no operator bound, represor bound (blocks σ factor binding/transcription)
• low lac operon expression needed: lactose effector (allolactose) binds repressor -> repressor can't bind operator. σ70-Pol binds promotor -> low operon expression

  • +lactose, +glucose (low cAMP)
  • low expression
  • cAMP not bound
• high lac operon expression needed: lactose effector binds repressor -> repressor can't bind operator. σ70-Pol binds promotor. cAMP binds CAP -> CAP-cAMP binds adjacent to promotor -> stimulates high operon expression

  • +lactose, -glucose (high cAMP)
  • everything but repressor bound
  • cAMP increases with decrease of glucose
• under high glucose/high lactose conditions, what is the effect on lac operon transcription if there is a CAP site mutation?

  less RNA

  sigma can still bind
• mammalian gene and proximal promoter

  • enhancers: landing sequences to help promote/regulate transcription
  • proximal promoter: within 200bp of transcription start

  1. TATA box (TATAAA)
  2. -20 to -30 upstream of start +1; fixed position
  3. recruits TATA binding protein (TBP) complex to promoter
  4. major assembly point for transcription preinitiation complex (PIC)
  5. initiator region (INR): sequence where DNA is unwound. +1 usually within or near
  6. other activation elements
  7. critical for docking basal transcription apparatus
  8. tissue specific elements