8' Control of Gene Expression 2

1
8. Control of Gene Expression (2)
2
Control of Gene Expression

• Repression of Transcription
• Cells also possess negative regulatory elements
• Mechanisms
• Binding to promoter elements
• Blocking assembly of the preinitiation complex
• Inhibiting binding or functioning of
  transcriptional activators
• Modifying DNA and its interaction with
  nucleosomes
• Some transcription factors activate some genes
  and repress others

3
Control of Gene Expression

• Repression of Transcription - Mechanisms
• Binding to promoter elements
• Blocking assembly of the preinitiation complex

4
Control of Gene Expression

• Repression of Transcription - Mechanisms
• Inhibiting binding or functioning of
  transcriptional activators

5
Control of Gene Expression

• Repression of Transcription
• DNA Methylation
• Methyl groups may be attached to cytosine (C5
  position)
• Methyltransferases
• Methyl groups provide a tag
• In mammals always part of a symmetrical sequence
• Concentrated in CG-rich domains
• Often in promoter regions
• Methylation of promoter DNA highly correlated
  with gene repression

6
Control of Gene Expression

• Repression of Transcription
• DNA Methylation
• Maintains a gene in inactive state rather than
  initiating gene repression Example
• Inactivation of genes of one X chromosome in
  female mammals occurs prior to a wave of
  methylation
• Shifts throughout life in DNA-methylation levels
• Early Zygote most methylation tags removed
• Implantation a new wave of methylation occurs
• Important example Genomic Imprinting

7
Control of Gene Expression

• Repression of Transcription
• DNA Methylation Genomic Imprinting
• Certain genes are active or inactive during early
  development
• Depending on whether they are paternal or
  maternal genes
• Eg IGF-2 is only active in the gene from the
  male parent
• The gene is imprinted according to parental
  origin
• Mammalian genome has gt 100 imprinted genes in
  clusters
• Imprinted due to selective methylation of one of
  the alleles

8
Control of Gene Expression

• Repression of Transcription
• DNA Methylation Genomic Imprinting
• In the early embryo the waves of demethylation
  and new methylation do not affect the methylation
  of imprinted genes
• Thus the same alleles are affected in the zygote
  through to the adult stage in the individual

9
Control of Gene Expression

• Repression of Transcription
• Chromatin structure and transcription
• DNA is not naked but wrapped around histone
  complexes to form nucleosomes
• How are transcription factors and RNA polymerases
  able to interact with DNA tightly associated with
  histones?
• Apparently nucleosome structure does inhibit
  initiation of transcription
• Initiation of transcription requires assembly of
  large complexes and nucleosomes block assembly at
  the core promoter

10
Control of Gene Expression

• Repression of Transcription
• Chromatin structure role of acetylation
• Genes which are actively transcribed are bound by
  histones which are acetylated
• Each of the histones has a flexible N-terminal
  tail
• Extends outside the core particle and the DNA
  helix
• Acetyl groups are added to lysine residues by
  enzymes
• Histone acetyl transferases (HATs)
• Acetylation has two functions
• Neutralize the positive charge on the lysine
  residues
• Destabilize interactions between histone tails
  and structural proteins

11
Control of Gene Expression

• Repression of Transcription
• Chromatin structure role of acetylation
• Some coactivators have HAT activity
• Links histone acetylation, chromatin structure
  and gene activation
• HAT activity of coactivator acetylates core
  histones bound to promoter DNA causing
• release of nucleosome core particles or
  loosening of histone-DNA interaction
• Subsequent binding of transcription factors and
  RNA polymerase
• Once transcription is initiated RNA polymerase
  is able to transcribe DNA packaged into
  nucleosomes
• Acetylation is dynamic enzymes also remove
  acetyl groups

12
Control of Gene Expression

• Repression of Transcription
• Chromatin structure role of deacetylation
• Removal of acetyl groups
• Histone deacetylases (HDACs)
• HDACs associated with transcriptional repression
• HDACs are subunits of larger complexes
  corepressors
• HDACs guided to regions of DNA by methylation
  patterns
• Example
• Inactive X chromosome of female
• Largely deacetylated histones
• Active X chromosome has a normal level of histone
  acetylation

13
Control of Gene Expression

• Repression of Transcription
• Chromatin structure Acetylation / Deacetylation

14
Control of Gene Expression

• Repression of Transcription
• Chromatin structure Acetylation / Deacetylation

15
Control of Gene Expression

• Processing-Level Control
• Recall that the formation of multigene families
  is a mechanism that generates protein diversity
• Protein diversity also generated via alternate
  splicing
• Regulates gene expression at the level of RNA
  processing
• A mechanism by which a single gene can encode two
  or more related proteins
• Most genes (and their primary transcripts)
  contain multiple introns and exons
• Often more than one pathway for processing of
  primary transcript

16
Control of Gene Expression

• Processing-Level Control
• Transcripts from approx 35 of human genes may be
  subjected to alternate splicing
• Simplest case a specific segment either spliced
  out or retained Example
• Fibronectin
• Synthesized by fibroblasts two additional
  peptides compared to that synthesized by liver
• Extra peptides encoded by pre-mRNA retained in
  fibroblast

17
Control of Gene Expression

• Translational-Level Control
• Wide variety of mechanisms affecting mRNA
  previously transported from the nucleus
• Subjects include
• Localization of mRNA in the cell
• mRNA translation
• Half-life of mRNA
• Mediated via interactions between mRNA and
  cytosolic proteins

18
Control of Gene Expression

• Translational-Level Control
• mRNA noncoding segments untranslated regions
  (UTRs)
• 5 UTR from methylguanosine cap to AUG
  initiation codon
• 3 UTR from termination codon to end of
  poly(A) tail
• UTRs contain nucleotide sequences which mediate
  translational-level control

19
Control of Gene Expression

• Translational-Level Control
• Cytoplasmic localization of mRNAs Example
  ferritin
• Translation regulated by iron regulatory protein
  (IRP)
• Activity of IRP dependent on cellular iron
  concentration
• At low iron concentration IRP binds the 5 UTR
• Bound IRP interferes physically with the binding
  of a ribosome to the 5 end of the mRNA
• At high iron concentration the IRP changes
  conformation and looses affinity for the 5 UTR

20
Control of Gene Expression

• Translational-Level Control
• Control of mRNA stability
• Half-life of mRNA is variable 10 minutes to 24
  hours
• Specific mRNAs are recognized in the cytoplasm
  and treated differentially
• mRNAs lacking the poly(A) tail are rapidly
  degraded
• Poly(A) tail is not naked mRNA but bound by the
  poly(A) binding protein (PABP)
• Each PABP bound to about 30 adenosine residues

21
Control of Gene Expression

• Translational-Level Control
• Control of mRNA stability
• PABP protects poly(A) tail from general nuclease
  activity
• But increases its sensitivity to poly(A)
  ribonuclease
• mRNA in cytoplasm is gradually reduced in length
  by poly(A) ribonuclease
• When the tail is reduced to approx 30 residues
• mRNA is rapidly degraded
• Degradation occurs from the 5 end
• Suggests two ends held in close proximity