Lecture 10 - Transcription

• Mutations: sustainer of life and cause great suffering

  • Source of all genetic variation, which further provides raw material for evolution
  • Source of many diseases and disorders
• Mutations are also useful for probing fundamental biological processes

  • Identifying mutant genes that alter development are useful for identifying the function of that gene
  • Example: Wnt genes involved in patterning the vertebrate axis now known to be at the center of colon cancer
• Types of Gene Mutations (based on their molecular nature)

  • Base substitutions
  • Transition
  • Transversion
  • Insertions and Deletions
  • Frameshift mutations
  • In-frame insertions and deletions
  • Expanding Nucleotide repeats
  • Increase in the number of a copies of a set of nucleotides
• A base substitution alters a single codon
• An insertion or a deletion alters the reading frame and may change many codons
• Stop codons are very prevalent in the genome, so a frame shift due to an insertion or deletion could result in a stop codon soon after the mutation
• Phenotypic effects of Mutations

  • Forward mutation: wild type-->mutant type
  • Reverse mutation: mutant type --> wild type
  • Missense mutation: amino acid --> different amino acid
  • Nonsense mutation: sense codon-->nonsense codon
  • Silent mutation: codon --> synonymous codon
  • Neutral mutation: no change in function
• No mutation results in the wild type protein produced
• Missense mutation results in the new codon to encode a different amino acid; there is a change in amino acid sequence
• Nonsense mutation: The new codon is a stop codon; there is premature termination of translation
• Silent mutation: The new codon encodes the same amino acid; there is no change in amino acid sequence
• Different mutations have different effects on the gene product\
• The effects of single base mutations vary enormously

  • silent (no effect)
  • nonsense (results in truncation)
  • missense (changes one amino acid)
• The central dogma of biology is DNA --> RNA --> Protein

  • with transcription and translation happening in between the arrows
• The genome

  • 6 billion bases of DNA distributed over 23 pairs of chromosomes
  • Only ~5% actually codes for protein
• Ribosomal RNA (rRNA) is in both prokaryotic and eukaryotic cells. In eukaryotes its located in the cytoplasm and its function is the structural and functional components of the ribosome (translation).
• Messenger RNA (mRNA) is in both prokaryotic and eukaryotic cells. In eukaryotes it is located in the nucleus and cytoplasm and its function is to carry genetic codes for proteins
• Transfer RNA (tRNA) is in both prokaryotic and eukaryotic cells. In eukaryotes it is located in the cytoplasm and its function is to help incorporate amino acids into polypeptide chain (translation).
• Is the Synthesis of an RNA molecule from a DNA template

  • The template strand: the transcribes strand
  • The transcription unit (a promoter, RNA-coding sequence, terminator (end))
• Terminator: a sequence of nucleotides that signals where transcription is to end. It is incorporated into the RNA, so transcription stops only after the terminator has been incorporated into the RNA.
• Upstream: A promoter sequence is located upstream of the transcriptional start site
• Downstream: The termination sequence is located downstream of the transcriptional starting site
• Only one of the DNA strands will be the template for RNA
• The template strand is in the 3' --> 5' orientation
• The template strand is transcribed (or read by the polymerase)
• RNA strand is synthesizes in the 5' -->3' direction
• RNA strand is complementary to the template strand
• RNA strand is similar to the non-template strand (except U's are substituted for T's)
• Genes can be located on the + or - strand on DNA
• Eukaryotic Transcription:

  • Promoter region binds to transcription factors that are specific for initiating transcription of that particular gene (not every gene is turned on all of the time)
  • Transcription factors recruit the polymerase to initiate transcription
  • Highly organized and regulated process
• In transcription, new RNA nucleotides are added at the 3' end. The transcript grows from the 5' end to the 3' end.
• Steps:

  1. Initiation of RNA synthesis does not require a primer
  2. New nucleotides are added to the 3' end of the RNA molecule
  3. DNA unwinds at the front of the transcription bubble...
  4. ...and then rewinds
• RNA polymerase II is the enzyme responsible for generating the majority of mRNA for making proteins