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BIOL 3542 > Final Exam - Limb Development > Flashcards

Final Exam - Limb Development Flashcards

1
Q

Specification of limb bud region

A
  • Is specified in the lateral plate mesoderm
  • The overlying ectoderm aids in patterning and outgrowth
  • Hox genes are involved in positioning the limb region and identity
  • Hox-b5: When function is missing there is a shift in the placement of the forelimb, the shoulder girdle towards the neck
  • Pitx1: Transcription factor that is expressed only in the hind limb, is necessary and sufficient for hindlimb identity

-specified: describes the developmental status of a group of cells that when isolated and cultured in a minimal medium will developing according to their normal fate

  • limb development initiates as small buds on the side of the embryo
  • Anterior buds give rise to forelimbs (wings in chick)
  • Posterior limbs give rise to hind limbs
  • The limb bud consists of mesoderm (originating from the lateral plate mesoderm) and an overlying layer of ectoderm
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2
Q

Lateral plate mesoderm

A
  • where the limb bud region is specified
  • mesoderm in vertebrate embryos that lies lateral and ventral to the somites and gives rise to the tissues of the heart, kidneys, gonads, blood and limb connective tissue
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3
Q

Hox genes

A
  • involved in positioning the limb region and identity
  • a family of homeobox-containing genes that are involved in patterning the antero-posterior axis
  • are clustered on the chromosomes in one or more gene clusters or complexes
  • combinatorial expression of different Hox genes characterizes different regions or structures along the axis
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4
Q

Hoxb5

A
  • involved in positioning the forelimb region and identity
  • When function is missing there is a shift in the placement of the forelimb
  • The shoulder girdle towards the neck
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5
Q

Pitx1

A
  • Transcription factor that is expressed only in the hind limb
  • Is necessary and sufficient to specify hindlimb identity
  • Pitx 1 mutants in hindlimbs show a transition to forelimb identity (take it away, you get two forelimbs)
  • A major consequence of Pitx 1, Tbx 5 and Tbx 4 expression is the upregulation of Fgf10 and Fgf8
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6
Q

Tbx5

A
  • in the forelimb
  • Are T-box transcription factors (related to brachyury)
  • limb bud development through control of Fgf production
  • The placement and expression of Tbx 5 and 4 are under the control of the Hox genes
  • A major consequence of Pitx 1, Tbx 5 and Tbx 4 expression is the upregulation of Fgf10 and Fgf8
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7
Q

Tbx4

A
  • in the hindlimb
  • are T-box transcription factors (related to brachyury)
  • limb bud development through control of Fgf production
  • The placement and expression of Tbx 5 and 4 are under the control of the Hox genes
  • A major consequence of Pitx 1, Tbx 5 and Tbx 4 expression is the upregulation of Fgf10 and Fgf8
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8
Q

Apical ectodermal rigde (AER)

A
  • Essential for limb bud outgrowth and proximal-distal patterning of the limb
  • The AER is a ridge of ectoderm that runs along the dorsal-ventral boundary of the limb
  • Maintains a region of cells just proximal to it in the mesoderm, the Zone of proliferating and undifferentiated cells
  • As cells leave this zone, they begin to differentiate
  • Differentiation occurs in the proximal to distal direction as the limb extends along this axis
  • A key signal produced by the AER is Fgf 8
  • Fgf 8 is sufficient for limb bud growth
  • Beads that have been soaked in Fgf 8 and applied to the limb region can mimic AER function
  • Fgfs are necessary for limb bud outgrowth
  • AER is required for outgrowth and if removed, outgrowth stops
  • Cells behind it stopped proliferating
  • Cells leave the proliferating zone, they differentiate and proximal structures form first then progressively more distal
  • Removal of the AER at later stages of limb growth result in formation of more distal structures

-a thickening of the ectoderm at the distal end of the developing chick and mammalian limb bud, which is essential for limb bud outgrowth and for correct patterning along the proximo-distal axis of the limb

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9
Q

Zone of proliferating undifferentiated cells

A
  • a region of cells maintained by the apical ectodermal ridge (AER) and is just proximal to it
  • these cells are proliferating (dividing) and are undifferentiated
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10
Q

Limb bud outgrowth

A
  • Fgfs are necessary for limb bud outgrowth
  • A key signal produced by the AER is Fgf 8 and is sufficient for limb bud growth
  • AER is required for outgrowth and if removed, outgrowth stops
  • Cells behind it stopped proliferating
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11
Q

Fgf-8

A
  • A major consequence of Pitx 1, Tbx 5 and Tbx 4 expression is the upregulation of Fgf10 and Fgf8
  • a key signal produced by the apical ectodermal ridge (AER) that is sufficient for limb bud growth
  • Beads that have been soaked in Fgf 8 and applied to the limb region can mimic AER function
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12
Q

Fgf-10

A

-A major consequence of Pitx 1, Tbx 5 and Tbx 4 expression is the upregulation of Fgf10 and Fgf8

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13
Q

AER removal experiment

A
  • AER is required for outgrowth and if removed, outgrowth stops
  • Cells behind it stopped proliferating
  • Cells leave the proliferating zone, they differentiate and proximal structures form first then progressively more distal
  • Removal of the AER at later stages of limb growth result in formation of more distal structures
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14
Q

Progress zone model

A
  • Cells interpret their position based on the amount of time they are present in the region behind the AER
  • Progress zone=undifferentiated and proliferating cells
  • Evidence?
  • AER outgrowth is tightly linked to proximal-distal patterning
  • Fgf functions to maintain the progress zone (removing this, they stop dividing, know where they are in the limb)
  • When the AER is removed, cells can still interpret their position as though the full limb had formed
  • If the progress zone is not maintained, more distal structures cannot form
  • X-ray treatments were used to kill cells in the progress zone during limb bud outgrowth
  • This results in loss of proximal but not distal structures
  • Cells in the progress zone after treatment do not have time to replenish the zone until later outgrowth (used in cancer treatments)
  • Cells that leave the zone are too few to generate mid proximal structures

-in the timing model of vertebrate limb development, an area at the tip of the limb bud where cells acquire positional values

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15
Q

Two-signal model

A
  • Diffusible signals in the early limb pattern the cells to specify proximal-distal values
  • Elements differentiate progressively as they grow out
  • Evidence?
  • Fgf expressed in the AER signals distal identity
  • Retinoic acid (RA) is expressed by the body wall muscles and opposes the Fgf gradient to signal proximal identity
  • High gradient in the flank of the animal
  • RA regulates a transcription factor called Meis in the proximal region
  • Meis expression in distal regions is sufficient to create limb truncations
  • Fgf mutations result in a truncated limb with proximal and distal structures but NO intervening structures
  • Fgf and RA pattern early limb to generate most distal and proximal structures FIRST
  • Limb outgrowth generates a region that can adopt an intermediate fate
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16
Q

Killing cells of the progress zone (X-ray irradiation)

A
  • used to prove the Progress Zone Model
  • X-ray treatments were used to kill cells in the progress zone during limb bud outgrowth
  • This results in loss of proximal but not distal structures
  • Cells in the progress zone after treatment do not have time to replenish the zone until later outgrowth (used in cancer treatments)
  • Cells that leave the zone are too few to generate mid proximal structures
17
Q

FGF

A
  • used to prove the two signal model
  • Fgf expressed in the AER signals distal identity
  • Retinoic acid (RA) is expressed by the body wall muscles and opposes the Fgf gradient to signal proximal identity
  • =OPPOSING GRADIENTS
  • High gradient in the flank of the animal
  • RA regulates a transcription factor called Meis in the proximal region
  • Meis expression in distal regions is sufficient to create limb truncations
  • Fgf mutations result in a truncated limb with proximal and distal structures but NO intervening structures
  • Fgf and RA pattern early limb to generate most distal and proximal structures FIRST
  • Limb outgrowth generates a region that can adopt an intermediate fate
18
Q

Retinoic acid

A
  • used to prove the two signal model
  • Fgf expressed in the AER signals distal identity
  • Retinoic acid (RA) is expressed by the body wall muscles and opposes the Fgf gradient to signal proximal identity
  • =OPPOSING GRADIENTS
  • High gradient in the flank of the animal
  • RA regulates a transcription factor called Meis in the proximal region
  • Meis expression in distal regions is sufficient to create limb truncations
  • Fgf mutations result in a truncated limb with proximal and distal structures but NO intervening structures
  • Fgf and RA pattern early limb to generate most distal and proximal structures FIRST
  • Limb outgrowth generates a region that can adopt an intermediate fate

-small non-protein secreted signalling molecule with many roles in development

19
Q

Meis transcription factor

A
  • Retinoic acid regulates a transcription factor called Meis in the proximal region
  • Meis expression in distal regions is sufficient to create limb truncations
20
Q

Zone of polarizing activity (ZPA) / polarizing region

A
  • is located in the posterior region of the limb bud and is necessary for anterior to posterior patterning
  • Is mesoderm tissue
  • Sonic hedgehog (Shh) is a ligand secreted protein in the ZPA (ie. Morphogen)
  • There is a concentration gradient of Shh in the limb (High in the posterior, Low in the anterior)
  • [Shh] specifies A-P positional identity
  • If ZPA is transplanted within a new position in the bud, this is able to induce a new A-P pattern
  • Is a true organizer
  • Able to induce surrounding tissue of the host to change fate
  • ZPA produces a morphogen (Shh) that forms a gradient
  • Cells interpret [Shh] to develop a particular structure (e.g. digit 2)
  • The graft of ZPA (donor) would form a mirror image if placed in the anterior
  • A weaker signal (i.e. less cells in the graft) lowers [Shh] and induces predictable structures in intermediate to posterior identity
  • Similar results are seen with a shorter period of time that the transplant is left in place

-in developing chick and mouse limb buds, an area at the posterior margin of the bud which produces a signal specifying position along the antero-posterior axis

21
Q

Sonic hedgehog (Shh)

A
  • a ligand secreted protein in the Zone of Polarizing Activity (ie. Morphogen)
  • There is a concentration gradient of Shh in the limb (High in the posterior, low in the anterior)
  • [Shh] specifies A-P positional identity
  • ZPA produces a morphogen (Shh) that forms a gradient
  • Cells interpret [Shh] to develop a particular structure (e.g. digit 2)
  • a weaker signal (i.e. less cells in the graft) lowers [Shh] and induces predictable structures in intermediate to posterior identity
22
Q

Organizer

A

-a signalling center that directs development of the whole embryo or of part of the embryo such as a limb
the zone of polarizing activity is a true organizer, able to induce surrounding tissue of the host to change their fates

23
Q

Morphogen

A
  • any substance active in pattern formation whose spatial concentration varies and to which cells respond differently at different threshold concentrations
  • Sonic hedgehog (Shh) secreted ligand and protein from the zone of polarizing activity is an example of a morphagen due to cells interpreting the different concentration levels of Shh
24
Q

ZPA grafting experiments

A
  • If ZPA is transplanted within a new position in the bud, this is able to induce a new A-P pattern
  • Is a true organizer
  • Able to induce surrounding tissue of the host to change fate
  • ZPA produces a morphogen (Shh) that forms a gradient
  • Cells interpret [Shh] to develop a particular structure (e.g. digit 2)
  • The graft of ZPA (donor) would form a mirror image if placed in the anterior
  • A weaker signal (i.e. less cells in the graft) lowers [Shh] and induces predictable structures in intermediate to posterior identity
  • Similar results are seen with a shorter period of time that the transplant is left in place
25
Q

Hox gene expression

A
  • hox gene expression reflects positional information along the proximal-distal and anterior-posterior axis
  • Anterior-Posterior axis: curved concentration with Hoxd9 (most anterior), Hoxd9-11, middle and Hoxd9-13, (most posterior
  • Proximal-distal axis: vertical lines with Hoxa9 (most posterior), Hoxa9-11 (middle) and Hoxa9-13 (most distal)
26
Q

Bone identity

A
  • Hox9=humerus
  • Hox10=humerus , radius, ulna and carpus
  • Hox11=radius, ulna, carpus and phalanges
  • Hox12=carpus
  • Hox13=palanges

BIOL 3542 (10 decks)

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