molecular patterning during development

Question 1

what are 3 principles governing cell differentiation?

Answer 1

• Generative program (The embryo does NOT contain a description of the adult, rather it contains a generative program for making it)
• Regulatory proteins work together as a “committee” to control the expression of a eukaryotic gene
• Combinations of a few gene regulatory proteins can generate many different cell types during development

Question 2

what is the process of cell differentiation?

what does cell differentiation involve?

what does it achieve?

how is it charcterised?

Answer 2

• the process by which embryonic cells become different from one another, which happens very early on soon after fertilisation
• Involves the emergence of cell types such as muscle, nerve, skin and fat cells
• Is the achievement of a stable terminal differentiated state (not just transitory differences)
• Is characterized by the profile of proteins expressed in that cell

Question 3

list the 3 steps of differentiation from a single fertilised egg to a multipotent stem cell

Answer 3

1. single fertilised egg through cell divisions into morule stage to form totipotent stem cell
2. then to blastocyst stage, there are two types of cell: placenta cells and the inner cell mass (which becomes the individual) - they are now pluripotent
3. from pluripotent, they differentiate into different organ systems - they are now multipotent

Question 4

what is the order of the heirarchy of stem cells?

Answer 4

totipotent to pluripotent to multipotent

Question 5

define potency

Answer 5

potency - the entire repertoire of cell types a particular cell can give rise to in all environments

Question 6

totipotent - define toti

pluripotent - define pluri

multipotent - define multi

Answer 6

• Totipotent: ‘toti’ = whole. eg. Cells of the very early mammalian embryo;
  identical and unrestricted; can give rise to any cell of the body (EMBRYONIC)
• Pluripotent: ‘pluri’ = more. eg. Inner cells of the blastocyst; less potent;
  can give rise to many cell types but not all (EMBRYONIC)
• Multipotent: ‘multi’ = many. e.g. Blood stem cells;
  they give rise to cells that have a particular function (ADULT)

Question 7

cells are committed - this step is a decision that restricts cell fate. what are the 2 stages of commitment?

Answer 7

First stage: specification (reversible)
1. Capable of differentiating autonomously if placed in isolation BUT can be respecified if exposed to certain chemicals/ signals.

Second stage: determination (irreversible change)
2. Cell will differentiate autonomously even when exposed to other factors or placed in a different part of the embryo.

Question 8

in what 2 ways does a naive cell (totipotent) become specified (differentiated)?

Answer 8

• Intrinsic signal – cell autonomous signal tells the cell ‘who is it’
• Extrinsic signal - a chemical or molecule in the
  environment gives the cell spatial information, tells the
  cell ‘where it is’

Question 9

what does determination imply?

Answer 9

Determination implies a stable change - the fate of determined cells does not change

Question 10

describe the progress of a cell during development?

what is competence?

Answer 10

1. cytoplasmic determinants or induction
2. loss of competence* for alternative fates
3. cell specific gene expression

competence - the ability of a cell to respond to the chemical stimuli (a cell can lose competence by changes in surface receptor or intracellular molecules)

Question 11

what do patterns in chromatin determine?

what is this called?

describe this mechanism:

Answer 11

patterns in chromatin determine the determination of differentiation

this is known a Bivalent chromatin - mechanism basis for these fate decisions

• only occurs in developmental master regulator genes
• bivalent chromatin occurs in the promoters of these genes
• it is a pattern of either open/closed chromatin structure (chemical change of histone proteins)
• early on the genes are poised - still need to make the decision to be expressed or not^

Question 12

a combination of a few regulatory proteins can generate many cell types:

where are these proteins acting within the cell?

what are the 2 factors which these sites bind onto?

Answer 12

where are these proteins acting within the cell:
- the promoter region
- these regulatory genes are transcription factors used to direct gene expression

note that most of the sites can bind either a stimulatory transcription or inhibitory factor but not both at once. thus the balance of stimulatory and inhibitory transcription factors in a cell determine how strongly it is expressed:

Question 13

describe endochondrial ossification

Answer 13

• bones form as an early event
• bones are formed by forming mesochyme
• which eventually becomes hard bone with chondrocytes and osteoblasts
• ossification occurs at growth plates

Question 14

describe intermembranous ossification

Answer 14

• Intramembraneous ossification is the formation of bone in fibrous connective tissue (which is formed from condensed mesenchyme cells)
• The process occurs during the formation of flat bones such as the mandible and flat bones of the skull
• the mesoderm is one of the early embryonic germ layers
• mesenchyme generalised embryonic connective tissue derived from mesoderm

Question 15

what 2 things do HOX genes determine?

where are they expressed?

Answer 15

HOX genes determine the body axis + the position of limbs across it as well as the shape of bones

HOX genes are a related group of genes that are expressed along the long axis of the embryo from head to tail.

Question 16

During embryonic development HOX genes determine the body axis and the position of the limbs along the body axis - Intrinsic factors

what are the products of HOX genes?

what 3 axes is limb growth regulated?

Answer 16

• The products of HOX genes belong to a class of proteins known as transcription factors, which bind to DNA, and thereby regulate the transcription of other genes (e.g. TBX5, TBX4).
• Once the cranio-caudal position is set, limb growth is regulated along three axes:
  1. Proximo-distal axis
  2. Antero-posterior axis
  3. Dorso-ventral axis

Question 17

when do upper limb buds appear and where?

when do lower limb buds appear and where?

Answer 17

• Upper Limb buds appear on approximately day 24 between somites C5-T1
• Lower limb buds appear on approximately day 28 between somites L1-S2
  • By week 8 all major components of the limbs are present
  • Medial rotation of the LL is complete

Question 18

rotation of the limbs:

when do limbs rotate and in which directions and by how much?

where do the flexor compartments lie in upper limb and lower limb?

Answer 18

• Development of fore and hind limbs is similar:
• In week 7 the forelimbs rotate 90° laterally and the hind limbs rotate 90 ° medially.
• Results in the flexor compartments being anterior in the upper limb and posterior in the lower limb

Question 19

describe the descriptive terms used in embryology growth

Answer 19

Question 20

proximo-distal development (wrist to fingers)

what is proximo-distal development controlled by?

what 2 things does a limb bud consist of?

Answer 20

• controlled by Apical Ectodermal Ridge AER
• limb bud consists of :
  
  • core of mesenchyme derived from parietal layer of lateral plate mesoderm
  • ectoderm which forms the outer covering of the limb (epidermis)
    ectoderm is thickened at the ‘apex’ of the developing limb to form the Apical ectodermal ridge

Question 21

what does the AER do in limb development (proximo-distal development)

Answer 21

1. it induces the underlying tissue to remain as a population of undifferentiated mesochymal, rapidly proliferating cells- known as the PROGRESS ZONE
2. As cells move further away from the AER they will begin to differentiate into cartilage and muscle
3. This differentiation results in proximo-distal development

Question 22

what are 4 controllers within Proximo-distal development?

Answer 22

1. HOX-8 controls the position of the limb on the long axis of the body
2. Initiation of outgrowth of the fore limb is controlled by the TBX5 gene and FGF-10
3. AER secretes FGF4 and FGF8 to maintain the progress zone and the further development of the proximodistal axis
4. As growth progresses, mesenchymal cells are left behind the advancing ridge (and its influence) and so they begin to differentiate

Question 23

what is the development of the antero-posterior axis (cranio-caudal axis) (thumb to little thinger) controlled by?

what does it ensure?

what does ZPA express?

Answer 23

• controlled by ZPA (zone of polarizing Activity)
• cluster of cells near the posterior border of the limb form the ZPA which regulated the AP axis
• it ensures that the thumb grows on the cranial (anterior) side of the limb bud
• ZPA expresses the protein sonic hedgehog (SHH). ZPA moves distally with the AER

Question 24

how is the dorsal-ventral axis (from back to front) formed?

Answer 24

• BMPs in the ventral ectoderm induce EN1
• EN1 represses WNT7 restricting its expression to the dorsal limb ectoderm
• WNT7 induces LMX1 which then specifies the cells to be dorsal

Question 25

how do HOX genes determine the shape of bones? how are upper limb bones and lower limb bones different? what is the expression of the HOX genes dependent on?

Answer 25

Variations in the combinations of HOX genes ensure that the: * Upper and lower limbs are different TBX5 (upper limb), TBX4 (lower limb) * Patterns for the proximal (arm) middle (radius and ulna) and distal (hand) are defined Expression of the HOX genes is dependent on SHH, FGFs and WNT7a

Question 26

in limb development, what is in charge of the shape?

Answer 26

apoptosis

Question 27

define: - amelia - meromelia - phocomelia - micromelia

Answer 27

- **Amelia:** complete absence of the limbs - **Meromelia:** partial absence of the limbs - **Phocomelia:** absence of long bones - **Micromelia:** segments are abnormally short

Question 28

thalidomide: how did thalidomide cause an increase in limb abnormalities? what were the babies which were born associated with? what did the thalidomide cause?

Answer 28

thalidomide was prescribed as a sleeping pill and due to this, there was an increase in incidence of limb abnormalities 12,000 babies survived with phocomelia (flipper like limbs) associated with intestinal atresia and cardiac abnormalities (Hot oram syndrom) thalidomide caused: amelia: prolonged or early exposure - leading to total loss of vessels, widespread cell death and all signalling lost phocomelia: short exposure - leading to loss of blood vessels, uniform or localised cell death, loss or partial loss of ZPA and AER signalling which later recovers to distalise remaining tissue

Question 29

what is hot oram syndrome caused by? list 4 limb deformities

Answer 29

- TBX5 mutations lead to defects in limb development causing upper limb deformities and heart defects: brachydactyly - short digits syndactyly - fused digits due to failure of apoptosis polydactyly - extra digits cleft foot - lobster claw deformity