REVIEW SESSION

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REVIEW SESSION

• Wednesday, September 15 530 PM
• SHANTZ 242 E

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Gene Regulation
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Gene Regulation

• Gene expression can be turned on, turned off,
  turned up or turned down!
• For example, as test time approaches, some of you
  may note that stomach acid production increases
  dramatically. due to regulation of the genes
  that control synthesis of HCl by cells within the
  gastric pits of the stomach lining.

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Gene Regulation in Prokaryotes

• Prokaryotes may turn genes on and off depending
  on metabolic demands and requirements for
  respective gene products.
• NOTE For prokaryotes, turning on/off refers
  almost exclusively to stimulating or repressing
  transcription

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Gene Regulation in Prokaryotes

• Inducible/Repressible gene products those
  produced only when specific chemical substrates
  are present/absent.
• Constitutive gene products those produced
  continuously, regardless of chemical substrates
  present.

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Gene Regulation in Prokaryotes

• Regulation may be
• Negative gene expression occurs unless it is
  shut off by a regulator molecule
• or
• Positive gene expression only occurs when a
  regulator mole turns it on

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Operons

• In prokaryotes, genes that code for enzymes all
  related to a single metabolic process tend to be
  organized into clusters within the genome, called
  operons.
• An operon is usually controlled by a single
  regulatory unit.

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Regulatory Elements

• cis-acting element The regulatory region of the
  DNA that binds the molecules that influences
  expression of the genes in the operon. It is
  almost always upstream (5) to the genes in the
  operon.
• Trans-acting element The molecule(s) that
  interact with the cis-element and influence
  expression of the genes in the operon.

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The lac operon

• The lac operon contains the genes that must be
  expressed if the bacteria is to use the
  disaccharide lactose as the primary energy
  source.
• To be used as an energy source, lactose must be
  cleaved into glucose and galactose. The glucose
  is then available for metabolism (glycolysis).
• Note glucose is the preferred energy substrate.

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Negative Control

• The genes in the lac operon are normally turned
  off, and only expressed when a repressor molecule
  is removed from the regulatory region.
• This repressor is removed only in the presence of
  lactose

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The lac Operon
Regulatory Region
Repressor gene
Structural Genes
PPromoter OOperator
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Structural Genes

• Structural genes are those that encode for the
  enzymes that do the metabolic work.
• LacZ b-galactosidase, cleaves lactose into
  glucose and galactose
• LacY Permease, promotes entry of lactose into
  cell
• LacA Transacetylase, thought to reduce toxicity
  of byproducts of lactose metabolism

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Structural Genes

• In prokaryotes, all the structural genes within
  an operon are usually transcribed as a single
  mRNA, then the genes are independently translated
  by ribosomes.

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LacIThe Repressor

• LacI is the regulatory molecule.
• When there is no lactose present in the cell,LacI
  binds to the Operator element and blocks binding
  of RNA polymerase to the Promoter element.

X
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LacIThe Repressor

• When lactose IS present, the genes to metabolize
  lactose must be expressed.
• Lactose itself causes LacI to dissociate from the
  operator, which frees up the promoter region,
  allowing RNA polymerase to bind, and
  transcription begins.
• Lactose is the inducer molecule for the lac
  operon.

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Induction of the lac operon
Lactose
Binding of lactose causes a change in the shape
of LacI
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Induction of the lac operon
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What happens if you mutate LacI?

• LacI encodes the lac repressor, which keeps the
  operon shut off in the absence of lactose.

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What happens if you mutate LacI?

• Inactivation of LacI would be called a
  constitutive mutation, because the genes of the
  lac operon would be on all the time even if there
  is no lactose present (removed repression).

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Positive Control of the lac Operon

• A further increase in transcription of the lac
  operon occurs if a molecule called
  catabolite-activating protein (CAP) also binds
  the promoter region.

CAP facilitates the binding of RNA
polymerase, and therefore increases transcription
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Positive Control

• Remember, glucose is the preferred substrate.
• CAP exists in the state that will bind the
  promoter ONLY when glucose is absent.

This is the form CAP takes when there is no
glucose
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Positive Control

• When glucose is present, CAP exists in a state
  that will NOT bind the promoter of the lac
  operon.

X
This is the shape CAP takes when glucose
is present. It cannot bind the promoter in this
shape
X
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Regulation of the lac Operon

• So, transcription is regulated as follows
• Off when lactose is absent (repressed)
• Active when lactose is present as well as glucose
  (de-repressed)
• Really active when lactose is present but glucose
  is absent (activated)

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Gene Regulation in Eukaryotes
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Differences between Prokaryotes and Eukaryotes

• 1. DNA is a lot more complicated in
  eukaryotestheres a lot more of it and its
  complexed with proteins to form chromatin
• 2. Genetic information is carried on multiple
  chromosomes
• 3. Transcription and translation are physically
  separated

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Differences between Prokaryotes and Eukaryotes
(cont.)

• 4. Eukaryotic mRNA is processed prior to
  translation
• 5. Eukaryotic mRNA is much more stable (not as
  easily degraded)
• Gene expression can be controlled at the level of
  translation!
• 6. Different cell types express different genes

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Chromatin Remodeling

• Chemical alteration of the histone proteins of
  chromatin facilitates or inhibits access of RNA
  polymerases to DNA promoters.

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Recruitment of Co-activators

• Remember enhancer elements? These are binding
  sites for molecules that influence formation of
  the RNA polymerase initiation complex.
• Enhancer elements may have DNA sequences for both
  positive and negative regulators of
  transcription.

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Enhancers

• The presence or absence of regulators is
  determined by the cells environment, metabolic
  state, developmental state and/or the presence or
  absence of signal molecules.
• The net effect of all the information available,
  summed up by the regulators present, dictates the
  transcription efficiency of RNA polymerase from a
  given promoter.

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DNA Methylation

• Chemical modification of DNA by adding or
  removing methyl (-CH3) groups from the DNA bases,
  usually cytosine.
• The presence of the methyl group alters the shape
  of DNA, which influences the binding of proteins
  to the methylated DNA.

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DNA Methylation

• Typically, increased methylation decreases
  transcription efficiency.
• In mammalian females, one X chromosome is
  inactivated (only one of the X chromosomes is
  used to drive transcription). The inactivated X
  chromosome has much more methylation than the
  active chromosome.

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Post-Transcriptional Regulation

• Alternative Splicing

Exon 1
Exon 2
Exon 3
Exon 4
Exon 5
1.
Exon 1
Exon 2
Exon 3
Exon 4
Exon 5
2.
Exon 1
Exon 2
Exon 4
Exon 5
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Post-Transcriptional Control

• RNA Stability
• Stability sequences
• Instability sequences
• Translation efficiencyincreased translation
  increases stability