Gene Regulation during Development

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Gene Regulation during Development
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Cells from different parts establish different
program of gene expression , and most
differential gene expression is regulated at the
level of transcription initiation .
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This chapter ,we are going to learn gene
Regulation during Development

• Outline
• Section 1.Three strategies by which cells are
  instructed to express specific sets of gene
  during development
• Section 2. Example of the three strategies for
  establishing different gene expression
• Section 3. The molecular biology of Drosophila
  embryogenesis

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Section 1

• The three major ways strategies used to instruct
  genetically-identical cells to express distinct
  sets of genes and thereby differentiate into
  diverse cells types are
• 1.mRNA localization
• 2.cell-to-cell contact
• 3.Signaling through the diffusion of a secreted
  signaling molecule

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mRNA localization

• Principal distributing mRNA asymmetrically
  during cell division? daughter cells inheriting
  different amount of the regulator ?following
  different pathways
• of development
• The roles of those
• mRNA are encoding
• RNA-binding proteins
• or cell signaling
• molecules and
• transcriptional activators and repressors .

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The common mechanism for localizing mRNAs isan
mRNA can be transported form one end to the other
by means of an adapter protein ,which binds to a
specific sequence within the 3UTR of an mRNA and
cytoskeleton. So the mRNA adapter complex can be
moved with the cytoskeleton.
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Cell to cell contact

• Principal a cell producing extracellular
  signaling proteins, then the given signal is
  recognized by specific receptor on the surface of
  
• recipient cells
• and triggers
• changes in
• gene expression
• in the recipient.

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This communication from the cell surface to the
nucleus often involves signal transduction
pathways . There are a few basic features of
these pathways

• 1. sometimes
• ligand-receptor
• interactions induce
• an enzymatic
• cascade the ultimately
• regulatory proteins
• already present in
• the nucleus.

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2.activated receptor cause the release of
DNA-binding proteins from the cell surface or
cytoplasm into the nucleus
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3.Upon cleavage,the intracytoplasmic domain of
the receptor is released from the cell surface
and enters the nucleus, to associate with
DNA-binding proteins and then influence
transcript
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Signaling through the diffusion of a secreted
signaling molecule
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A recurring theme in development is the
importance of a cells position within a
developing embryo or organ in determining what it
will become.The influence of location on
development is called positional
information.Signaling molecules that control
position information are sometimes called
morphogens
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The most common ways of establishing positional
information A small group of cells synthesize
andsecrete signals ? the signal distribute
in an extracellular gradient ? cells in
different positions receive different signal
concentrations ? levels of activated
transcription factor differ ? lead to the
expression of different sets of genes
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Section 2Examples of the Three Strategies for
Establishing Differential Gene Expression
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EXAMPLE 1 the localized Ash 1 repressor controls
mating types yeast by silencing the HO gene
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The yeast S.cerevisiae can grow as haploid cells
that divide by budding. Which give rises to two
cellsmother cell and the daughter cell.
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SWITCHING ISCONTROLLED BY THE PRODUCT OF THE
HOGENE

• A mother cell and its daughter cell can exhibit
• different mating types. This difference arises
• by a process called mating-type switching. After
  budding, a mother cell can switch mating type.

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It is kept silent in the daughter cell due to
the selective expression of a repressor called
Ash1. The ash1 gene is transcribed in the mother
cell during budding.But the encoded RNA becomes
localized within the daughter cell through the
following process.
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• During budding ,the ash1 mRNA attaches to the
  growing ends of microtubules .movement is
  directed and begins at the - ends of the
  filament and extends with the growing ends.

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• The ash1 mRNA transport depends on
• the binding of the She2 and She3
• adapter proteins to specific
• sequences contained within the 3UTR

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• Once localized within the daughter cell, the ash1
  mRNA is translated into a repressor protein that
  binds to, and inhibits the transcription of, the
  HO gene . This silencing of OH expression in the
  daughter cell prevents that cell from undergoing
  mating-type switching

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EXAMPLE 2

• Localized mRNAs can establish
• differential gene expression among the
  genetically-identical cells of a developing embryo

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Just as the fate of the daughter cell is
constrained by its inheritance of the ash1 mRNA
in yeast, the cells in a developing embryo can be
instructed to follow specific pathways of
developmentthrough the inheritance of localized
mRNA.Such as the cionadevelopment.
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This is the Ciona life cycle. They are
hermaphrodites and possess both sperm and eggs,
then fertilize to develop into a new individual.
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Early cleavage in AscidiansThe fertilized,
1-cell ascidian embryo contains a number of
localized determinants that control the
development of different tissues. For example,
the yellow determinants is inherited by cells
that form the tail muscles.
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In the case of muscle differentiation in sea
squirts, a major determinant for programming
cells to form muscle is a regulatory protein
called Macho-1.the Macho-1 mRNA becomes
localized in the fertilized egg.
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EXAMPLE 3

• Cell to cell contact elicits differential gene
  expression in the sporulating bacterium ,
  B.subtilis.

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Under adverse condition,the bacterium Bacillus
subtilis can formspores First, a septum is
formed within the sporangium then the septum
produces two cells forespore (smaller one)
and the mother cell (larger one).The mother cell
can help the forespore to form a spore. And the
forespore can also influences the expression of
genes in the neighboring mother cell.
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The forespore contains an active form of a
specific sfactor,sF (inactive in mother cell).
The spollR gene is activated by sF in forespore.
The SpollR protein triggers the activation of sE,
and leads to the recruitment of RNA polymerase
and the activation of specific genes in mother
cell.
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EXAMPLE 4

• A skin-nerve regulatory switch is controlled by
  notch signaling in the inset CNS

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We must first describe the development of the
ventral nerve cord in insect embryos.

• It arises from a
• sheet of cells called
• neurogenic ectoderm,
• then forms two
• major cell type
• neurons and skin
• cells (or epidermis).

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The developing neurons contain a signaling
molecule Delta on their surface, which binds to a
receptor Notch on the skin cells. The activation
of the Notch receptor by Delta renders them
incapable of developing into neurons.
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The developing neuron does not express neuronal
repressor genes. These genes are kept off by a
DNA-binding protein called Su(H) and associated
repressor proteins. The Delta-notch interactions
cause neuronal repressor genes to express.
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In all vertebrate embryos, there is a stage when
cells located along the future back to move in a
coordinated fashion toward internal regions and
form the neural tube.Cells located in the
ventralmost region of the neural tube form a
specialized structure called the floorplate.
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Section 3

• The molecular biology of drosophila embryogenesis

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Development of germ cell

• Polar granules located in the posterior
  cytoplasm of the unfertilized egg contain germ
  cell determinants, and the Nanos mRNA. Nuclei
  begin to migrate to the periphery. Those that
  enter posterior regions sequester the polar
  granules and form the pole cells, which form the
  germ cells.

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Ligation experiment is the first evidence that
has shown that cell fate specification is
controlled by localized maternal determinants
that are deposited into the egg during oogenesis.

• When a hair is used to separate the anterior and
  posterior halves of early embryos, then
  determinants emanating from the anterior pole
  fail to enter posterior regions. As a result, the
  embryos develop into abnormal files that lack
  thoracic structure.

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In contrast, when the hair separates older
embryos, then the determinant already entered
posterior regions and a normal thorax forms.
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Embryogenesis of the fruit fly,drosophila
melanogaster,has been studied extensively as a
relatively simple model for cell differentiation
in animals .
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During fertilization ,a single sperm cell enters
a mature egg ,and the haploid sperm and egg
nuclei fuse to form a diploid nucleus.
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Overview of Drosophila Development

• 1.After fertilization, the diploid ,zygotic
  nucleus undergoes 10 rapid cleavages,the embryo
  is called a syncitium at this point, because it
  is a single cell with multiple nuclei. Acturally
  after 8 cleavages ,the resulting 256 nuclei begin
  to migrate to the outer edge of the cell. During
  this migration,the nuclei undergo two more
  cleavages,resulting 1024 nuclei.

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2.About 90 minutes after fertilization, most
nuclei have reached the cortex. These nuclei
undergo 3 more cleavages
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3.From 2-3 hours after fertizilation, cell
membrances form between nuclei ,and embryo
transforms into a cellular blastoderm.
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When the nuclei reach the edge of the cell ,they
are totipotent. Just after cellularization,the
nuclei have been irreversibly determined to
differentiate into specific tissues in the adult
fruit fly. The location each nucleus determines
its fate.
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A morphogen gradient contrals Dorsal-Ventral
patterning of the Drosophila embryo

• This is determinated by a regulatory protein
  called Dorsal.After fertilization, Dorsal enters
  nuclei in ventral and lateral regions but remains
  in the cytoplasm in dorsal regions.
• Regulated nuclear transport of the Dorsal protein
  is controlled by a cell signaling molecule called
  Spätzle. This signal is distributed in a
  vintral-to-dorsal gradient within the
  extracellular matrix.

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• After fertilization, Spätzle binds to the cell
  surface toll receptor . Toll is activated to a
  greater or lesser extent by the degree of
  receptor occupancy in a given region of the
  syncitial embryo.

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Toll signaling causes the degradation of a
cytoplasmic inhibitor, Cactus, and the release of
Dorsal from the cytoplasm into nuclei.This leads
to the formation of a corresponding Dorsal
nuclear gradient in the ventral half of the early
embryo.
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We now consider the regulation of three different
target genes that are actived by
high,intermediate,and low levels of the Dorsal
protein-twist,rhomboid,and sog.
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The highest levels of Dorsal protein activate the
expression of the twist gene . The reason for
this is that the twist 5regulatory DNA contains
two low-affinity Dorsal binding sites.
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The intermediate levels of the Dorsal protein
activate the rhomboid gene. The rhomboid
5flanking region contain a 300bp enhancer
location about 1.5kb 5of the transcription start
site. But it is kept off in the mesoderm by a
transcriptional repressor called Snail.
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The lowest levels of the Dorsal protein activate
the sog gene. The Snail repressor precludes
activation of sog expression in the mesoderm.
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Segmentation is initiated by localized RNAs at
the anterior and posterior poles of the
unfertilized egg.

• The unfertilized egg contains two localized
  mRNAs, bicoid in anterior regions and osker in
  posterior regions.

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The osker mRNA encodes an RNA-binding protein
that is responsible for the assembly of polar
granule, which contral the development of tissues
that arise from posterior regions of the early
embryo.
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During the formation of the Drosophila egg,
polarized microtubules are formed that extend
from the oocyte nucleus and grow toward the
posterior plasm. The oskar mRNA binds adapter
proteins that interact with the microtubules,
thus transport the RNA to the posterior plasm.
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The bicoid UTR causes it to be localized to the
anterior pole while the distinct oskar UTR causes
localization in the posterior plasm. An
engineered oskar mRNA that contains the bicoid
UTR is localized to the anterior pole. This
mislocalization of oskar causes the formation of
pole cells in anterior regions.
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The bicoid gradient regulates the expression of
segmentation genes in a concentration-dependent
fashion.

• There are peak levels of the
• Bicoid protein in anterior
• regions, intermediate levels in
• certral regions, and low levels
• in posterior regions.
• Orthodenticle is activated only
• by high levels of the Bicoid
• gradient in the head
• hunchback is activated by both
• high and intermediate levels in
• the head and thorax.

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In central regions of the embryo, the
orthodenticle gene is off. In contrast, hunchback
is on. These is contral by the levels of Bicoid
protein.
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Hunchback expression is also regulatd at the
level of translation

• The translation is blocked in posterior regions
  by RNA-binding protein called Nanos. After the
  Nanos mRNA is translated, the protein diffuses
  from posterior regions to form a gradient. The
  translation of the maternal hunchback mRNA is
  arrested by the Nanos protein. The Nanos gradient
  thereby leads to the formation of a reciprocal
  Hunchback gradient in anterior regions.

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The gradient of hunchcack repressor estiblishes
different limits of gap gene expresion.

• High levels of Hunchback are required for the
• repression of Krüppel, whereas intermediate and
• low levels
• repress the
• expression of
• knirps and
• gaint, respectively.

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Besides, the number of Hunchback repressor sites
may be a more critical determinant for distinct
patterns of Krüppel, knirps and gaint.
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Hunchback and gap proteins produce segmentation
stripes of gene expression.

• A culminating event in the regulatory cascade
  that
• begins with the localized bicoid and oskar mRNAs
  is
• the expression of a pair-rule gene called
  even-skipped.

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The eve locus contains over 12kb of regulatory
DNA. The 5 regulatory region contains two
enhancers. These control the expression of
stripes 2,3,and 7. Each enhancer is 500bp in
length. The 3 regulatory region contains three
enhancers. These contain the expression of
stripes 1,4,and 6. The five enhancers produce
seven stripes of eve expression in the early
embryo.
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Regulation of eve stripe 2

• a. The 500bp enhancer contains a total of twelve
  binding sites for the Bicoid, Hunchback,
  Krüppel,and Giant proteins.
• b. There are high levels of the Bicoid and
  Hunchback proteins in the cells that express eve
  stripe 2. The borders of the stripes are formed
  by the Giant and Krüppel proteins

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Two distinct modes of transcriptional repression.

• the binding of Krüppel repressor
• to the Kr1 and
• Kr3 sites
• Precludes
• the binding
• of Bicoid to
• overlapping
• sites.

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b.The binding of Krüppel to the Kr2 sites does
not interfere with the binding of the Bicoid to
adjacent sites. So Krüppel mediates repression by
recruiting the CtBP repressor protein.
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Gap repressor gradients produce many stripes of
gene expression

• The enhancer that controls
• the expression of eve stripe
• 4 is also repressed by
• Hunchback and Knirps.
• Differential regulation of the
• stripe 3 and stripe 4
• enhancers by opposing
• gradients of the Hunchback
• and Knirps repressors.

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Different enhancers work independently of one
another in the eve regulatory region due to
short-range transcriptional repression.