Eukaryotic Genomes: Organization, Regulation and Evolution.

1
Chapter 19

• Eukaryotic Genomes Organization, Regulation and
  Evolution.

2
Chromatin

• The DNA-protein complex found in eukaryotes.
• It is much more complex in eukaryotes than in
  prokaryotes.

3
The DNA Within Cells

• It undergoes a variety of changes as it proceeds
  through the cell cycle.
• Recall, in prophase its highly diffuse, but as
  the cell prepares to divide, it becomes highly
  condensed.
• Proteins called histones are responsible for the
  first level of DNA packing in chromatin.
• The mass of histone is nearly equal to the mass
  of DNA.

4
DNA-Histone Binding

• DNA is negatively charged, and histones contain a
  high proportion of positively charged aas and
  enable easy binding of the histones to the DNA.

5
DNA-Histone Binding

• Histones play a very important role in organizing
  DNA and they are very good at it.
• Thus, this is a likely reason why histone genes
  have been conserved throughout the generations in
  the course of evolution.
• The structure of histones are very similar among
  eukaryotes and between eukaryotes and prokaryotes.

6
DNA-Histone Binding and DNA Packing

• Electron micrographs show unfolded chromatin and
  they look like beads on a string.
• These beads are referred to as nucleosomes (the
  basic unit of DNA packing), and the string is DNA.

7
The Nucleosome and DNA Packing

• A nucleosome is a piece of DNA wound around a
  protein core.
• This DNA-histone association remains in tact
  throughout the cell cycle.
• Histones only leave the DNA very briefly during
  DNA replication.
• With very few exceptions, histones stay with the
  DNA during transcription.

8
Nucleosome Interaction and DNA Packing

• The next level of DNA packing takes place between
  the histone tails of one nucleosome/linker DNA
  and the nucleosomes to either side.
• The interactions between these cause the DNA to
  coil even tighter.
• As they continue to coil and fold, eventually the
  DNA resembles that of the metaphase chromosome.

9
DNA Packing

• Movie

10
Heterochromatin Vs. Euchromatin

• During interphase, some of the DNA remains
  condensed as you would normally see it in
  metaphase. (centrosomes, telomeres, and some
  other regions of the chromosome).
• This is called heterochromatin to distinguish it
  from euchromatin which condenses and relaxes with
  the cell cycle.
• Heterochromatin is rarely transcribed.

11
The Structural Organization of Chromatin

• The structural organization of chromatin is
  important in helping regulate gene expression.
  Also, the location of a genes promoter relative
  to nucleosomes and to sites where DNA attaches to
  the chromosome scaffold or nuclear lamina can
  also affect whether it is transcribed or not.
• Research indicates that chemical modification to
  the histones and DNA of chromatin influence
  chromatin structure and gene expression.

12
Acetylation

• There is a lot of evidence supporting the notion
  that the regulation of gene expression is, in
  part, dependent upon chemical modifications to
  histones.
• When an acetyl group is added to the histone
  tail, the histones become neutralized and the
  chromatin loosens up.
• As a result, transcription can occur.

13

• The enzymes that interact with histones are
  closely associated with, or are components of
  transcription factors that bind to promoters.

14
Methylation

• Addition of a methyl group to a histone tail
  leads to condensation of the chromatin.

15
Histone Code Hypothesis

• This hypothesis states that the specific
  modifications of histones help determine
  chromatin configuration thus influencing
  transcription.

16
DNA Methylation

• DNA methylation is completely separate from
  histone methylation, but may be a way in which
  genes become inactivated.
• Evidence
• Inactivated X chromosomes are heavily methylated.
• In many cells that have inactivated genes, the
  genes are more heavily methylated than in cells
  where the genes are active.

17
Control of Eukaryotic Gene Expression

• Recall the idea of the operon and how it
  regulated bacterial gene expression.
• The mechanism of gene expression in eukaryotes is
  different.
• It involves chromatin modifications, but they do
  not involve a change in DNA sequence. Moreover,
  they can be passed on to future generations by
  what is known as epigenetic inheritance.

18
Epigenetic Inheritance

• Epigenetic inheritance occurs when traits are
  passed on and do not involve the nucleotide
  sequences (proteins, enzymes, organelles).
• It also seems to be very important in the
  regulation of gene expression.
• The enzymes that modify chromatin are integral
  parts of the cells machinery that regulates
  transcription.

19
Chromatin Modifying Enzymes

• These provide initial control of gene expression.
• They make the region of DNA more or less able to
  bind DNA machinery.
• Once optimized for expression, the initiation of
  transcription is the most universally used stage
  at which gene expression is regulated.

20
Recall,

• Eukaryotic genes have promoters, a DNA sequence
  where RNA polymerase II binds and starts
  transcription.
• There are numerous control elements involved in
  regulating the initiation of transcription.
• 5 caps.
• Poly-A tails.

21
Also,

• RNA modifications help prevent enzymatic
  degradation of mRNA, allowing more protein to be
  made.

Movie
22
Recall,

• RNA processing involves 3 steps
• 1. Addition of the 5 cap.
• 2. Addition of the poly-A tail.
• 3. Gene splicing.
• Removal of introns and splicing together of exons.

23
Recall,

• The transcription initiation complex assembles on
  the promoter sequence.
• RNA polymerase II proceeds to transcribe the gene
  making pre-mRNA.
• Transcription factors are proteins that assist
  RNA polymerase II to initiate transcription.

24
RNA Processing

• Movie

25
Eukaryotic Gene Expression

• Most eukaryotic genes are associated with
  multiple control elements which are segments of
  non-coding DNA that help regulate transcription
  by binding certain proteins.
• These control elements are crucial to the
  regulation of certain genes within different
  cells.

26
Eukaryotic Gene Expression

• Only after the complete initiation complex has
  assembled can the polymerase begin to move along
  the DNA template strand, producing a
  complementary strand of DNA.

27
Eukaryotic Gene Expression

• In eukaryotes, high levels of transcription of a
  particular gene at the appropriate time depends
  on the interaction of control elements with other
  proteins called transcription factors.
• Enhancers and activators play important roles in
  gene expression.
• Enhancers are nucleotide sequences that bind
  activators and stimulate gene expression.

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Enhancer-Activator Interaction and Eukaryotic
Gene Expression

• When the activators bind to the enhancers, this
  causes the DNA to bend allowing interaction of
  the proteins and the promoter.
• This helps to position the initiation complex on
  the promoter so RNA synthesis can occur.

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Eukaryotic Gene Expression

• Some specific transcription factors function as
  repressors to inhibit expression of a particular
  gene.
• Certain repressors can block the binding of
  activators either to their control elements or to
  parts of their transcriptional machinery.
• Other repressors bind directly to their own
  control elements in an enhancer and act to turn
  off transcription.

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(No Transcript)
31
Transcription Initiation

• Movie

32
Blocking Transcription

• Movie

33
Eukaryotic Gene Expression

• There are only a dozen or so short nucleotide
  sequences that exist in control elements for
  different genes.
• The combinations of these control elements are
  more important than the presence of single unique
  control elements in regulating the transcription
  of a gene.

34
Recall,

• Prokaryotes typically have coordinately
  controlled genes clustered in an operon. The
  operons are regulated by single promoters and get
  transcribed into a single mRNA molecule. Thus
  genes are expressed together, and proteins are
  made concurrently.

35
Control of Eukaryotic Gene Expression

• Recent studies indicate that within genomes of
  many eukaryotic species, co-expressed genes are
  clustered near one another on the same
  chromosome.
• However, unlike the genes in the operons of
  prokaryotes, each of the eukaryotic genes have
  their own promoter and is individually
  transcribed.
• It is thought that the coordinate regulation of
  genes clustered in eukaryotic cells involves
  changes in chromatin structure that makes the
  entire group of genes available or unavailable.

36
Control of Eukaryotic Gene Expression

• More commonly, co-expressed eukaryotic genes are
  found scattered over different chromosomes. In
  these cases, coordinate gene expression is
  seemingly dependent on the association of
  specific control elements or combinations of
  every gene of a dispersed group.
• Copies of activators that recognize these control
  elements bind to them, promoting simultaneous
  transcription of the genes no matter where they
  are in the genome.

37
Control of Eukaryotic Gene Expression

• The coordinate control of dispersed genes in a
  eukaryotic cell often occurs in response to
  external signals such as hormones.
• When the steroid enters the cell, it binds to a
  specific intracellular receptor protein forming a
  hormone-receptor complex that serves as a
  transcription activator.

38
Control of Eukaryotic Gene Expression

• In an alternative mechanism, a signal molecule
  such as a non-steroid hormone or a growth factor
  bind to a receptor on a cells surface and never
  enter a cell.
• Instead, they control gene expression by inducing
  a signal transduction pathway.

39
Post-transcriptional Regulation and Control of
Gene Expression

• The mechanisms weve just discussed involve
  regulating the expression of the gene.
• Post-transcriptional regulation involves
  regulating the transcript after the mRNA has been
  made.
• These modes are unique to eukaryotes.

40
Alternative RNA Splicing and Control of Gene
Expression

• Alternative RNA splicing is a way in which
  different mRNA transcripts are produced from the
  same primary transcript.
• This is determined by which RNA segments are
  treated as introns and which are treated as exons.

41
Alternative RNA Splicing and Control of Gene
Expression

• Different cells have different regulatory
  proteins that control intron-exon choices by
  binding to regulatory sequences within the
  primary transcript.

42
Alternative Mechanisms to Control Gene Expression

• Protein processing is the final spot for
  controlling gene expression.
• Often, eukaryotic polypeptides undergo further
  processing to yield a functional protein.
  Regulation can occur at any of the sites of
  protein modification.

43
Protein Processing

• Movie