mod 7

• Gene expression


  Whether a gene is being transcribed, not whether the protein encoded by the gene is present in the cell
• Reasons for regulating gene expression

  • Cost: wasteful to make a protein that is not required
  • Transcription and translation use a lot of energy
  • Harmful: presence of a protein at the wrong time could impair cell function
• When genes are turned on and off

  • In response to the external environment
  • Temporally, to carry out specific cellular or organismal programs
  • Development of multicellular organisms
  • Response to internal environment
• Gene expression control


  1. Transcriptional

  2. Translational

  3. Posttranslational

  4. RNA processing

  
• Method to identify bacterial genes

  1. Gene transfer from one bacterium to another for recombination

  2. Conjugation

  3. F-plasmid

  4. Mapping by time of entry with Hfr strains

  5. Transduction

  6. Transfer of genes via bacteriophage intermediate

  7. Mapping by cotransduction frequency

  8. Transformation

  9. Lateral transfer of genes in bacteria

  
• Constitutive gene expression


  Expressed continually

  
• Inducible gene expression


  Expression is turned on in response to a certain factor

  
• Key approaches to genetic analysis in bacteria

  • Forward genetics
  • Reverse genetics

  
• Chromosomes


  Structures that contain genetic material in eukaryotic cells
• Repressible gene expression


  Expression is turned off in response to a certain factor
• Tree of life


  • Thermotagales
  • Flavobacteria
  • Cyanobacteria
  • Purple bacteria
  • Gram positives
  • Green nonsulphur bacteria
  • Pyrodictium
  • Thermoproteus
  • Methanococcus
  • Methanobacterium
  • Methanosarcina
  • Haloarchaea
  • Animals
  • Fungi
  • Plants
• Metaphase chromosomes


  • Formed after DNA replication and before cell division
  • Sister chromatids held together most tightly at centromere

  
• Transcription factors (TFs)


  • Sequence-specific DNA binding proteins
  • Bind to the DNA of the genes they regulate
  • Binding sites primarily located outside of the transcribed region
  • Have DNA binding, regulatory, and activation/repression domains

  
• Karyotype


  Chromosome complement of a cell or organism
• Bacteria are very successful
• Negative regulation of transcription in bacteria

  Repressor binds to operator adjacent to promoter, blocking RNA polymerase binding

  
• Bacteria differ from Eukaryotes

  • Chromosomes
  • Location of DNA
  • Lack of membrane bound organelles
  • DNA tertiary structure
  • Recombination
  • Concept of 'species' is more vague. Individuals within the same species may show high variability in their genes

  
• Karyotyping


  • Characterizing chromosomes within a species by length, location of centromere, banding pattern
  • Used by cytogeneticists to identify chromosomal abnormalities linked to disorders

  
• Positive regulation of transcription in bacteria

  Activators bind adjacent to weak promoters, helping recruit RNA polymerase

  
• Bacteria provided key insights into genetics
• Chromosome banding techniques


  • Giemsa staining (G-banding)
  • Fluorescent in situ hybridization (FISH)
  • Spectral karyotyping (SKY)
• Common ancestor with eukaryotes – similarities and key differences
• Regulation of TF activity

  • Constitutively expressed TFs have their activity regulated
  • Transcriptionally or translationally regulated TFs are selectively expressed

  
• Syntenic chromosomal regions


  Regions between species that contain the same genes
• E. coli


  The standard bacterium used in most lab studies
• Lac operon


  Model system for understanding transcriptional regulation in bacteria and eukaryotes

  
• Growth in the lab does not necessarily reflect how bacteria grow in the wild
• Chromosome breakage and repair

  1. Cell hates double-strand breaks

  2. Non-homologous end-joining (NHEJ) following breaks

  3. Illegitimate recombination between repetitive DNA sequences
• Biofilms


  Groups of microorganisms that adhere to a surface usually embedded in a secreted matrix. Heterogeneous population of cells.

  
• Lac operon


  • Encodes enzymes for lactose utilization
  • Negatively regulated by LacI repressor
  • LacI repressor activity is negatively regulated by allolactose

  
• Chromosomal rearrangements


  • Deletion
  • Duplication
  • Inversion
  • Translocation

  
• Auxotroph


  Cannot grow in the absence of a chemical (wild-type can grow without)
• lacI and lacO mutations


  Constitutive expression of lac operon genes

  
• Prototroph


  Usually refers to the wild-type that can grow in the absence of the chemical
• Chromosomal rearrangements


  Can change the amount of genetic information or disrupt gene function
• cis-acting elements


  Influence gene expression only when on the same chromosome and usually close by

  
• Conjugation


  1. Natural, via physical contact

  2. Donor cell transfers DNA through a connecting tube

  3. Process is controlled by a specialized plasmid

  
• Consequences of chromosomal rearrangements

  • Deletion - loss of genetic information, potential haploinsufficiency
  • Duplication - extra copies of genes, potential genome imbalance
  • Inversion - can suppress recombination, disrupt genes
  • Translocation - can fuse genes, change gene regulation
• trans-acting factors


  Can influence expression of genes on homologous and same chromosome
• Transformation


  Uptake of DNA from the environment