Gene Expression: mRNA Transcription

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Gene ExpressionmRNA Transcription

• September 15, 2003

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The DNA is in Charge
Today
Ribosome
Protein
I
III
II
AGCTAGTCGATAGCTGATCTGCTGATGCTGATGCTGATCATGCTAGCTAG
CTAGCTAGCTAGCATACGTAGCGA
rRNA
tRNA
mRNA
(Nucleolus)
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Flow of Genetic Information
DNA
RNA Transcription
DNA Replication
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Todays Exciting Topics

• Role of Messenger RNA
• Basic Steps of Transcription
• Prokaryotic Transcription
• Some Differences in Eukaryotic Transcription
• Enhancers
• Posttranscriptional Processing

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rRNA and tRNA are Cogs in the Machinery

• rRNA is a structural part of the ribosome
• tRNA helps the protein machinery to read the mRNA
• Neither of these types of RNAs actually carries
  any information

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Messenger RNA

• Messenger RNA carries the information in the DNA
  to the protein translation machinery (ribosomes)
• Serves as the template for protein synthesis
• Which mRNAs are transcribed in a cell decide the
  fate of that cell since they dictate which
  information in the DNA is read by the protein
  translation machinery

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How do mRNAs Encode Protein?(Preview of Dr.
Hodels Lectures)

• Each three bases in the mRNA codes for a
  different one of the 20 amino acids
• In the ribosome these three bases are recognized
  by the tRNA that is charged with the
  appropriate amino acid
• That amino acid is then added to the peptide chain

Methionine
Phenylalanine
Serine
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3
UAGUUUUCC
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Todays Exciting Topics

• Role of Messenger RNA
• Basic Steps of Transcription
• Prokaryotic Transcription
• Some Differences in Eukaryotic Transcription
• Enhancers
• Posttranscriptional Processing

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RNA Transcription

• RNA transcription can be broken down into three
  different steps
• Initiation
• Elongation
• Termination

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Transcription
Coding DNA Template Strand
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(No Transcript)
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Regulation can occur at any of these steps

• Initiation- highly regulated step
• Elongation- the rate at which the mRNA is made
  can control how quickly its made
• Termination- premature termination can mean that
  the whole mRNA never gets made and neither does
  what it codes for
• Like receiving only part of the instructions on
  how to put together your easy to assemble
  bookcase/desk/whatever

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Todays Exciting Topics

• Role of Messenger RNA
• Basic Steps of Transcription
• Prokaryotic Transcription
• Some Differences in Eukaryotic Transcription
• Enhancers
• Posttranscriptional Processing

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Prokaryotic Transcription of mRNA

• In prokaryotes all RNA is transcribed by the same
  RNA polymerase
• There is no nucleus to separate the transcribed
  RNA from the protein translation machinery
• Translation start as soon as the RNA is
  transcribed
• This is called
• Coupled Transcription and Translation

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Coupled Transcription and Translation
RNA polymerase
Actively Transcribed Chromatin
Ribosomes
mRNA
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Initiation Prokaryotic

• Occurs at specific sites within the DNA called
  promoters
• Requires an initiation factor (s) that helps
• to recognize the promoter sequence
• Two step process
• Closed Complex
• Open Complex

Gene
Promoter
Transcription Start Site
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Initiation Prokaryotic Promoter Region

• RNA Polymerase Recognizes the Promoter with the
  help of an Accessory Protein called Sigma Factor
• First they bind to the Closed Helix and then a
  Change Occurs and the Helix Opens i.e.

Start Site
Closed Complex
Open Complex
-35 Region
-10 Region
mRNA
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Initiation can occur at Multiple Sites
Bacterial Chromosome
Direction of Transcription
Initiation site
RNA Fibrils
Termination site
Initiation site
RNA Fibrils
Termination site
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Initiation can Regulate the Amount of Different
mRNAs Made

• The number of initiation events at different
  promoters varies
• Promoters that closely match the Consensus
  Promoter are Strong Promoters
• Strong promoters can have an initiation event
  every two seconds
• Weak promoters can have one only once every ten
  minutes

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Strong (2 sec)
Weak (10 min)
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Elongation Prokaryotic

• Elongation phase occurs when s factors is lost
  from the complex
• RNA polymerase moves like a snowplow through the
  double helix with the help of topisomerases
• The average rate of elongation for prokaryotic
  RNA polymerase is about 40 nucleotides/second
• Rate of elongation can vary depending on the
  secondary structure of the RNA (loops or hairpins)

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Termination Prokaryotic

• Termination occurs at specific sites after all
  the information in the gene has been encoded in
  the mRNA
• Termination can be
• Rho-dependent (not understood)
• Rho-independent

Rho-independent Termination Site
Hairpin slows the polymerase And then weak AU
pairs allow polymerase to fall off
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Todays Exciting Topics

• Role of Messenger RNA
• Basic Steps of Transcription
• Prokaryotic Transcription
• Initiation, elongation, termination
• Some Differences in Eukaryotic Transcription
• Enhancers
• Posttranscriptional Processing

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Eukaryotic Transcription

• Has to be more control of how more complex
  genetic material is read to create more variety
  (multicellular)
• RNA has to be transcribed in the nucleus and then
  transported to the protein translation machinery
  in the cytoplasm before it can be read (compare
  to the Coupled transcription and translation of
  prokaryotes)

DNA
Nucleus
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Transcription of mRNA in Eukaryotes

• Steps involved are the same as in prokaryotes
• Initiation
• Elongation
• Termination
• Mediated by RNA polymerase II
• Very complex enzyme with many subunits

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Eukaryotic Intiation

• Initiation occurs at promoters as in prokaryotes-
  eukaryotic promoters are not well-characterized
  but have some well conserved elements- including
  the TATA box and CAAT box (both have AT pairs)
• In addition to the promoters there are region in
  the DNA called enhancers to which transcription
  factors bind and regulate which DNA is read and
  encoded in mRNA

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Transcription Factor Function
Pol
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Elongation and Termination

• Neither is understood in as much detail as in
  prokaryotes
• Elongation of the chain occurs by the addition of
  nucleotide to the 3-OH of the previous
  nucleotide
• Can be regulated by the rate at which nucleotides
  are added
• Termination occurs after a specific signal in the
  DNA

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Posttranscriptional ProcessingSplicing

• Often eukaryotic mRNAs have extra pieces in them
  called introns that have to be spliced out to
  make the mature message

Exons are spliced together to make the final mRNA
WHY?????
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More Posttranscriptional Processing

• Once the mRNA is made it has a long journey ahead
  to the cytoplasm
• mRNA gets processed to make it more stable
• Needs to be transported
• Needs to be relatively stable over time

AAAAAA
5
3
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(No Transcript)
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Why is eukaryotic mRNA modified?
AAAAAA
5
3
Prokaryotic mRNA
Eukaryotic mRNA
5
3
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Basic Scheme The Life of an mRNA
DNA
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Todays Exciting Topics

• Role of Messenger RNA
• Basic Steps of Transcription
• Initiation, elongation, termination
• Prokaryotic Transcription
• Some Differences in Eukaryotic Transcription
• Enhancers
• Posttranscriptional Processing

35
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