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Molecular developmental biology II. Formation of
the body pattern, organs and appendages
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A C. elegans development from perspektive of
the individual cell

• Nematode worm consist of only about 1000 somatic
  cells and 1000-2000 germ cells. In every
  individual worm a given precursor cell follows
  the same pattern of cell divisions, and with a
  few exception the fate of the descendant cell can
  be predicted from its position in the lineage.
  This degree of stereotyped precision is not seen
  in the development of larger animals. Therefore
  it seems that cell fate is strictly following an
  internal program in the Nematode, but in fact
  cell-cell interactions are just as important here
  as in other animals.

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Segregation of P granules into founder cell of
the C. elegans germ line
Upper row shows cells stained with DNA specific
fluorescent dye in the nuclei
Lower row shows cells stained with antibody
against P granules (ribonucleoprotein particles,
directed by the partitioning defective gene, par)
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Asymmetric cell divisions play a role in the
early development of nematoda embryo

• The matternal-effect genes define early
  patterning, product of par (partitioning
  defective) gene help to transports
  ribonucleoprotein particles, P-granules to the
  posterior pole of the egg, the P-bodies are
  concentrated in a single cell until the 16 cell
  stage, this cell give rise to the germline
• Progenitors of the muscle, skin and neurons are
  already singled out in the 8-cell stage when the
  other cells are still totipotent

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Cell responsiveness to developmental signals
change over time

• if one of two daughter cells meets Notch signal
  and reacts to it by the same token it can become
  resistant to this signal
• in the next cell divison decendants of the other
  daughter cell will receive signal via the Notch
  pathway and in this case only they and not the
  progeny of the other resistant daughter cell
  differentiate into i.e. pharinx cells

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Do cells count how many cell divisions they have
to make for their development?
The determined neuroblasts undergo a defined
number of divisions.
But what happens if the cell has not gain
determination yet?
Is there a builtin clock of cell divisions in
cell development?
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• Timing of cell divisons and development the
  effect of lin-14 mutations on the change of the
  number of cell divisons. The loss of function of
  lin-14 will differentiate into adult already in
  the second larval stage, and the lin-14 gain of
  funkcion will be differentiated only in the fifth
  larval sage. On the top of the lin and let gene
  cascades there are RNA producing genes which acts
  on the other genes according to the principal of
  RNA interference (RNAi)

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• (The mechanism of RNA interference)
• formation of double stranded DNA
• dicing of dsRNS into siRNS (small interfering
  RNS)
• binding of RISC (RNA-induced silencing complex)
  protein
• unwinding of siRNS
• single stranded small RNAs with RISC binds to
  target RNA
• The double stranded RNA is cleaved by RNase

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Do count cells their divisions in timing their
internal program?

• Mutations of lin has proven that cells do not
  make such a thing! (cells can reach the terminal
  differentiation with less or more division)
• Drosophila, vertebrates, and mammalian examples
  are in line with this
• naturally in case of less cell divisions
  developmental defects might occur, simply because
  a single undivided cell can not differentiat in
  two ways at once

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Apoptosis is frequent in development

• The programed cell death happenes in a perfectly
  predictable way in a nematode (i. e. which cells
  die at what time in what parts of the body)
• In other animals, because of technical reasons,
  it is more difficult to follow apoptosis since
  the suiciding cells quickly disappear, their
  remnants may be swallowed by the neighgbouring
  cells

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The body pattern of Drosophila melanogaster

• it has 100 times as many cells as the nematoda,
  more parallels with our body structure
• the number of genes is less, 14 thousands in D.
  m. (nematoda19 thousands)
• it has more non coding DNA than the nematode, 10
  thousands bp per gene (nematoda 5 thousands bp)
• smaller genom, but larger combinatorial
  variability
• the Drosophila genes have mammalian homologs
  but there is less redundancy, (i.e. Duplications)
  in the fly
• suprisingly, Drosophila provided the key to
  understanding of the molecular genetics of our
  development

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• Drosophila 6-4 head, 3 thorax, 9 abdominal
  segment, first the parasegments are formed (half
  a segment out of register with the segments)
• Egg polarity regulating genes poszterior
  (nanoslocalised RNS), anterior (bicoidloc.RNS),
  terminal (Torsotransmembr. rec.), dorsoventral
  (Tolltransmembr. rec.) - ekto-, meso-, endoderm
  formation
• Gap genes mark out coarse subdivision (approx.
  6, i. e. Krüppel T1-A1)
• Pair rule genes every second parasegment,
  approx. 7 (fushi tarazu, even-skipped)
• Segment polarity genes mutations result in
  mirror image duplication of parasegments ( about
  10 kinds, i.e. gooseberry)
• Homeotic selector genes

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The nature of the body pattern regulating genes

• 75 of them is coding for transcription
  regulating protein, which acts on other genes
• Their expression is built on each other and
  formes a pattern where the cells remember their
  position and create determination
• The homeotic selector genes permanently
  distinguish one parasegment from another

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The homeotic selector genes

• Antennapedia complex it makes the head and
  thorax parasegments different (mutants forms
  antenna instead of a leg)
• bithorax complex controls the difference between
  the thorax and abdominal segments (in some
  mutants an extra wing is formed instead of the
  small haltera, - a fly with four wing)
• they determine the positional value of the cells
• they are expressed almost exactly in the order of
  their position on the chromosomes, the gene
  located in more posterior position is always
  dominant
• They form the antero-posterior axis
• their pattern will be stabilised by two other
  groups of genes Polycomb and trithorax

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The homeotic genes

• they are found in every investigated animals
  (hidras, nematoda, mollusc, mammalians)
• Ordered in complexes, there are four HOX
  complexes in mammals, each complex has homologue
  genes in the Drosophila Hox complex (HoxC) and
  probably have been formed by duplication
• their head-tail pattern is well distinquished in
  mammalians rhombencephalon (cerebellum, pons,
  hindbrain)
• the Hox complex is about 500 million years old

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The limb development

• Vertebrate or arthropode limbs look different but
  develop by a similar mechanism
• The precursor of the limb in the common ancestor
  might have been a protruding mouth organ
• In the developing limb bud of vertebrates every
  Drosophila genes expressed in wing development
  has a homolog

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The imaginal discs in Drosophila

• Consist of undifferentiated epithel cells, these
  are the sources of the formation of eye, antenna,
  wing, leg, genitals etc. in holometabolus insects
• They are formed already in larvae
• They grow and develop their internal pattern as
  the larva grows
• In metamorphosis they evert (turn inside out) and
  help to form the epidermal layer of the adult
• Studying transplantations of imaginal disc
  revealed a great deal about determination

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The compartments in the Drosophila imaginal wing
disc
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Molecules regulating vertebrate limb bud
development

• The scanning microscopic picture of the limb bud
  of a 4 d chick embryo from dorsal view (the
  somites are on the left)
• Expression of regulatory proteins that control
  patterning in a vertebrate bud

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The organogenesis gaining the final shape of
the differentiated organs

• The expression of many genes are directed by
  master genes (i.e. MyoD, myogenin, myf-5 and MRF4
  regulate myogenesis)
• The regulatory proteins that determine cell type
  are often belong to the helix-loop-helix (HLH)
  family of transcription factors, and their
  control is directed by the Notch signal pathway
  with frequent feedback loops

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Thesaurus

• Somatic cell line, germ cell line
• Polarity genes
• Gap genes
• Pair-rule genes
• Segment polarity genes
• Homeotic genes
• Master genes