Gene Expression and Regulation

• Gene expression is the process by which information from a gene is used in the synthesis of a functional gene product such as a protein
• Transcription


  The process of making messenger RNA (mRNA) from a DNA template by RNA polymerase
• Transcription revisited

  Likely order of events in producing a mature mRNA from a pre-mRNA
• Transcription: 3 DNA-dependent RNA polymerases

  1. Pol I synthesizes 45S rRNA precursor, found in nucleoli (45S→18S, 28S, 5.8S rRNAs)

  2. Pol II synthesizes mRNA precursors, some snRNAs

  3. Pol III synthesizes 5S rRNAs, tRNAs, small nuclear RNAs (snRNAs)
• Relative cellular RNA abundance

  • Ribosomal RNAs (rRNAs) ~ 90%
  • Transfer RNAs (tRNAs) ~ 5%
  • Messenger RNAs (mRNAs) ~ 2%
  • The rest (~3%): Signal recognition particle (SRP) RNA, Small nuclear RNAs (snRNAs), Small nucleolar RNAs (snoRNAs), Micro RNAs (miRNAs)
• RNA polymerase II

  • 2 large subunits have regions of homology with ß and ß’ subunits of E. coli RNAP
  • Largest subunit is phosphorylated on its COOH-terminal domain (CTD) which is needed for transition from initiation to elongation
  • CTD also interacts with other proteins
  • Does not bind DNA by itself, requires other proteins to bind to promoter first
• Cis-regulatory elements (CREs) are regions of non-coding DNA which regulate the transcription of neighbouring genes
• Cis-regulatory elements (CREs) consist of

  • Promoters
  • Enhancers
  • Silencers
• Promoters


  • Relatively short sequences of DNA which include the site where transcription is initiated and the region approximately 35 bp upstream or downstream from the initiation site
• RNAP II promoters

  • Class-II promoters have 4 components: Upstream element(s), TATA Box (at approx. –25), Initiation region (includes the first transcribed nt, +1), Downstream element
• Upstream elements: Class II promoters

  • GC boxes (GGGCGG and CCGCCCC) stimulate transcription in either orientation, may be multiple copies, must be close to TATA box
  • CCAAT box stimulates transcription, binds CTF (Cat-box transcription factor)
• Upstream elements in Class II promoters

  • Promoters
  • GC boxes (GGGCGG and CCGCCCC)
  • CCAAT box
  • TATA box
• GC boxes

  • Stimulate transcription in either orientation
  • May be multiple copies
  • Must be close to TATA box
• CCAAT box

  • Stimulates transcription
  • Binds CTF (Cat-box transcription factor)
• TATA box of Class II promoters

  • Defines where transcription starts
  • Required for efficient transcription for some promoters
  • Bound by TBP – TATA box binding protein (in complexes like TFIID)
• Enhancers


  • Influence (enhance) the transcription of genes on the same molecule of DNA, stimulate transcription
• Silencers

  • Can bind transcription regulation factors (proteins) called repressors and inhibit the transcription
  • Orientation-independent
  • Position-independent (mostly)
  • Can work at a distance from promoter core
  • Enhancers have been found all over
  • Bind regulatory transcription factors
• Promoter elements not required for transcription initiation

  • CAAT box
  • GC box
  • Other gene-specific elements (light-responsive, nutrient-responsive, etc.)
  • Enhancer elements
• Transcriptional regulation

  • Controlling the rate of gene transcription, for example by helping or hindering RNA polymerase binding to DNA
  • Upregulation, activation, or promotion – increase the rate of gene transcription
  • Downregulation, repression, or suppression – decrease the rate of gene transcription
• Transcription factors for Class II promoters

  • Basal factors
  • Upstream factors
  • Inducible (regulated) factors
• Biological roles of transcription factors include basal transcription regulation, development, response to intercellular signals, response to environment, and cell cycle control
• The CRT/DRE response element responds to dehydration and cold-induced transcription factors (CBF)
• Eukaryotic transcription factors have a modular structure with DNA-binding domains, transcription-activating domains, and can have more than one of each type of module
• DNA-binding domains can recognize specific DNA sequences or have a general affinity to DNA
• Enhancers enhance transcription by acting on the promoter in cis and each enhancer has its own binding protein
• Enhancers regulate tissue
• Possible locations of a location in DNA

  • Upstream
  • Downstream
  • Intron
  • Exon
  • Untranslated region
• Enhancers enhance transcription by acting on promoter in cis
• Each enhancer has its own binding protein
• Enhancers are trans-regulatory activating factors
• Sequence of enhancers is variable
• Enhancers regulate tissue-specific and temporal expression of genes
• Enhancers activation from a distance

  3 possible models: Factor binding induces supercoiling of the promoter DNA, sliding of the complex to the promoter, looping out of DNA between enhancer and promoter
• Enhancer can work from downstream and upstream region
• TATA Binding Protein (TBP)

  Transcription factor with DNA-binding domains allowing it to bind directly to a cis-regulatory element
• Extreme trans-acting effectors of transcription: TAL effectors are from plant pathogenic bacteria Xanthomonas, secreted by bacteria when they infect, bind with plant promoters to express genes beneficial for the bacteria
• Some trans-acting elements prevent transcription
• Chromatin modification can regulate transcription by modifying chromatin (histones); highly transcribed genes have less condensed chromatin
• Basic unit of chromatin is the nucleosome: 4 different histones in the core (H2a, H2b, H3, H4 x 2 = octamer), 146 bp of DNA wrapped around core, histone H1 on outside
• Histone acetylation causes localized unpacking of nucleosomes, enhancing factor binding to DNA; de-acetylated histones bind DNA more strongly, nucleosomes condense into a solenoid inhibiting factor binding to DNA targets