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MUTANTS genetic variation in human development Lecture 7 Fall 2006 Bennington College. Overall comments for Manar Maget/Bodyshock Essay. Really great work - amazing variety of public health issues raised great insights and discussion of Manar’s situation and the - PowerPoint PPT Presentation
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MUTANTSgenetic variation in human development
Lecture 7
Fall 2006Bennington College
Overall comments for Manar Maget/Bodyshock Essay
Really great work - amazing variety of public health issues raised
great insights and discussion of Manar’s situation and the
Bodyshock program in general
things to work on:Attention to detail - make sure to address/answer allof the questions the assignment asks for.
PLEASE read over your paper BEFORE you print it out.Nothing says “I couldn’t be bothered to check my workahead of time” like hand-written corrections at the lastminute…
For the sake of my quickly declining eyesight, pleaseat least 1.5 space everything (I still have no vision plan…)
POLYDACTYLY
(aka “digital enhancement”)
The “normal” number of digits variesgreatly between vertebrate species:
humans, mice, cats, dogs, elephants - 5
pigs - 4camels - 2horses - 1
Relatively common for extra digits to occur:
1:3000 Europeans (usually thumbs)
1:300 Africans (usually pinkies)
extra pinkie finger extra pinkie toe
Same guy, by the way…
POLYDACTYLY
POLYDACTYLY
bonus thumbs
POLYDACTYLY
usually genetic
generally dominant
runs in families over many generations
What governs the formation of the “correct” number of digits?
John Saunders and Mary Gasseling addressed this question usingsurgically manipulated chicken embryos
For correct limb and digit development, three specialized cell clusters are of primary importance: the apical ectodermal ridge (AER), the progress zone (PZ), andthe zone of polarizing activity (ZPA).
limb bud
Saunders and Gasseling
The result was a palindromic duplication of the wing and digits mirror image polydactyly
Postulated that the ZPA secreted a polarizing morphogen
They named this area of mesodermal tissue the Zone of Polarizing Activity
morphogen intensity
numerous potential candidates for the morphogen that dictates digit number and polarity
Numerous mouse models for polydactyly usedto investigate possible morphogen candidates
Doublefoot
Sasquatch
Extra Toes
Crick, A. et al., Developmental mechanisms underlying polydactyly in the mouse mutant Doublefoot. J. Anat., 202: 21-26, 2003.
skeletal phenotype of right hindlimbs in WT andDoublefoot mutant mice at embryonic day 17.5
Doublefoot
In Doublefoot mutant mice, the expression pattern of SHH itself is normal, but there is ectopic expression of SHH signaling pathway components
ectopic expression - when a gene is expressed in cells that it is not normally expressed in.
Doublefoot
The sonic hedgehog signaling pathway
SHH
SHH receptor(Patched - PTC)
CELL
NucleusCytoplasm
is activated by the binding of SHH to PTC
activated smoothened signals for the expression of SHH target genes
Smoothened
Crick, A. et al., Developmental mechanisms underlying polydactyly in the mouse mutant Doublefoot. J. Anat., 202: 21-26, 2003.
Patched expression in embryonic day 10.5 embryos
SHH expression in embryonic day 10.5 limb buds
Doublefoot
Blanc, I. et al., Unusual pattern of sonic hedgehog expressionin the polydactylous mouse mutant Hemimelic extra-toes. Int. J. Dev. Biol. 46: 969-974, 2002.
Mouse extra-toes mutant stained for SHH expression at embryonic day 11.5
WTforelimb
Hx +/-forelimb
Hx +/-hindlimb
Extra toes
Blanc, I. et al., Unusual pattern of sonic hedgehog expressionin the polydactylous mouse mutant Hemimelic extra-toes. Int. J. Dev. Biol. 46: 969-974, 2002.
forelimb paw (7 digits) hindlimb paw (6 digits) hindlimb skeleton
Appearance of newborn polydactylic limbs in mouse extra-toes mutant
Extra toes
To initiate gene expression (or in some cases, to prevent gene expression), regulatory proteins (homeobox proteins and other transcription factors)bind to DNA upstream of the start of the target gene at regulatory elements.
The most recent evidence suggest that the Hx extra-toes mutation is in a regulatory element upstream of the SHH gene.
Sharpe et al., Identification of Sonic hedgehog as a candidate gene responsible for the polydactylous mouse mutant Sasquatch. Current Biology. 9: 97-100.
WT Ssq/-strong weak
variable polydactyly phenotypesassociated with the Sasquatch mutation (all pictures are of the hindlimb)
Ssq/-
adult
embryo
Sasquatch
Sharpe et al., Identification of Sonic hedgehog as a candidate gene responsible for the polydactylous mouse mutant Sasquatch. Current Biology. 9: 97-100.
forelimb
hindlimb
Sasquatch
Shh
ShhFgf8
Hoxd13
WT WT+/- +/- -/-
WT
WT +/- +/- +/- +/-
-/--/--/--/-
Sharpe et al., Identification of Sonic hedgehog as a candidate gene responsible for the polydactylous mouse mutant Sasquatch. Current Biology. 9: 97-100.
forelimb-------------------hindlimb------------------
Sasquatch mutation induces ectopic expression of Shh, elevated expression of FGF8, and ectopic expression of Hoxd13
Sasquatch
Sonic hedgehog seems to be a good candidate for theZPA morphogen…
-altered regulation (too much or too little Shh produced)
- abnormal pattern of expression (ectopic expression)
- expression of other components in the Shh signaling pathway altered
Sonic hedgehog seems to be a good candidate for theZPA morphogen…
- Altering Shh levels can result in mirror image polydactyly in chicken wings
- Shh mutant mice lack paws (similar to acheriopody)
- expression of other components in the Shh signaling pathway altered
mutations in either a transcription factor itself or in its the DNA of its regulatory elements can alter function
There are at least 10 genes that, when mutated,affect the activity of the Shh signaling pathway
- forebrain separation
- facial geometry
- digital determination
crosstalk and feedback between signaling pathwaysmake it complicated to determine exactly whatcontrols what
for example: If the AER and FGFs secreted from it fail, the result is failure of limb formation
one role of Shh is to maintain and shape the activityof the AER
one function of the AER is to maintain and shapeShh production if the ZPA
Signaling pathways are not independent, but rather are interconnected and often reciprocal in nature
The role of Programmed Cell Death in fine tuning digit formation
~ day 32 first signs of bone formation are visible - “condensation”
first to develop are those bones closest to the body:
humerus --- radius, ulna ---wrist and palm bones --- digits
by day 38 the ends of the limb buds look like paddles
the ectoderm between the paddles forms involutions and the extra cells “die off” to clearly define the 5 digits
Programmed Cell Death - occurs not only during development but also in some adult tissues
used to balance cell proliferation to maintain constant cell numbersin tissues that undergo cell turnover:
liverblood cells (~5 x 1011 killed daily)
also used as a defense mechanism to protect against virusesand DNA damage that can lead to cancerous cells
During development, Programmed Cell Death is used extensively:
gets rid of larval tissues during insect and amphibian metamorphosis
gets rid of excess neurons (up to 50% of all neurons initially made end up being destroyed - the one that make their proper connectionswith their target cells cause those cells to secrete growth factors whichblock initiation of the programmed cell death pathway
eliminates the extra tissue between digits during finger and toe formation (so we don’t end up with webbed hands and feet like ducks…)
Apoptosis - the distinct series of cellular changes that occurs during Programmed Cell Death
Apoptotic cells and fragments are effectively removed by macrophagesand neighboring cells
partly due to “eat me” signals normally not displayedon the cell surface (certain lipid moieties, for example,phophatidylserine)
Apoptosis is different than accidental cell deathfrom injury - those cells will swell and burst, releasing their contents into the extracellularspace and causing inflammation.
Digit formation also depends upon Hox genes (homeotic genes)
Hox gene mutations result in:
short big toes and bent pinkies (single mutant allele of Hoxd13)synpolydactyly (extra and often fused digits)missing forearm bones, fingers, and toes (deletion of 9 Hox genes)
Hox gene mutations also affect other appendages that grow outward from the body (i.e. genitalia)
What can Hox genes tell us about our origins?
are our limbs ≈ fins?
are our fingers ≈ fin rays?
very different type of bone…
what about “closer: relatives, the lobe-finned fishes?
lungfish
coelacanth
the lobe-finned fishes seem to have cognatesof our humerus, radius and ulna.
even have some small bones that could becognates of our digits
made of the right kind of bone…
Pros:
Cons:the geometry is all wrong
But still the fin buds have AER, ZPA, FGFs, Shh, and Hox genes
fin -vs- limb
Hoxd13 (or fish orthologue) is expressed in the ZPA of both
in fin buds, Hoxd13 only expressed for a short timeand over a short range
in limb buds, Hoxd13 stays on much longer and reachesall the way across the outermost part of the ZPA
fin -vs- limb
Does Hoxd13 have the power to specify our digits?
Do ∆hoxd13 mice have fins instead of arms and paws with digits?
NO! but they do have small deformed digits and 6 instead of 5
Suggests that perhaps the common ancestor of modern land-dwellingvertebrates had more digits than what we now have and that Hox geneshave evolved to trim down and define this number
Some contenders for this common ancestor (from the Devonian era ~360 million years ago)
Acanthostega
8 digits
Some contenders for this common ancestor (from the Devonian era ~360 million years ago)
Icthyostega
6 or 7 digits