Developmental Biology

Question 1

Why do the eggs of different species used as embryological models look so different? And what are the differences between mammalian eggs and other eggs?

Answer 1

Differences in the reproductive systems between the species
Mammalian eggs are typically small with very little yolk as the embryo is typically nourished by the placenta rather than non-mammalian eggs where most of the nourishment comes from the yolk sac

Question 2

What are the advantages of studying c.elegans?

Answer 2

Transparency (allowing dyes to be used to track cell lineage)
Full cell lineage is known
Roughly the same number of genes as humans (though a more compact genome)
Many of these gene can be controlled through RNA interference

Question 3

What can be learnt from c.elegans?

Answer 3

The initial assymetric developments leading to the formation of the body axis
Early development involves cellular division of zygote to blastomeres with changes in cell number but also change to smaller size
Early cleavage is not dependant on zygotic DNA but rather maternal mRNA
Zygotic genes are activated at different times in different organisms
During initial cell division in devlopment there is no growth phase

Question 4

What are the features of the first 3 cell divisions?

Answer 4

P0 is a totipotent cell polarised to animal and vegetative space
The first division gives rise to a large AB cell followed by a P1 cell
The second round of division causes AB to divide into ABa and ABp while P1 divides into P2 and EMS
The third division causes P2 to divide into P3 and C, while EMS divides into MS and E

Question 5

What drives the early asymmetric cell divisions?

Answer 5

Feedback from cell positions rather than an autonomous process

Question 6

What is the mechanism of asymmetric cell division?

Answer 6

Cytoplasmic determinants are segregated

Mitotic spindle must form across division of determinents

Question 7

How do PAR genes control early asymmetric cell division in c.elegans?

Answer 7

PAR3 determines if there is rotation of the mitotic spindle, if PAR 3 is absent the rotation occurs (as is the case with p1, were the rotation allows segregation along the anterior/posterior axis)
PAR3 expression is determined by PAR2 as it restricts its function

Question 8

What is segregated into the AB cell?

Answer 8

PAR3 and PAR6, and PKC-3 via co-localisation at the anterior cell cortex before and during the first mitosis

Question 9

What is segregated into the P1 cell?

Answer 9

PAR-2 and PAR-1 localise to the posterior cell cortex and segregate into P1 upon mitosis.

Question 10

What are the early cytoplasmic signals received by the AB cells in C.elegans embryo?

Answer 10

ABp cell differentiates in response to an interation between GLP1 Receptor and Apx1 ligand from P2
(ABa does not differentiate as it is in the incorrect position to recieve Apx1)

Question 11

What are the early cytoplasmic signals received by the EMS cell in C.elegans embryo?

Answer 11

EMS is driven to differentiate via a Mom2 ligand, Mom6 receptor complex from P2
consequently the pop1 transcription factor is downregulated at the posterior side of EMS
Therefore the resultant MS contains high levels of Pop1 and the resultant E contains low levels

Question 12

What is unique about drosophila embryo cleavage?

Answer 12

The cellular cleavage results in formation of a syncytium where there is a shared cytoplasm and multiple nuclei due to division
These nuclei then migrate to the periphery of the syncytium while eventually membranes grow to cause the formation of individual cells

Question 13

What is the role of the nurse cells in the drosophila embryo?

Answer 13

Synthesis of maternal mRNAS and delivery to the follicle

This allows formation of mRNA gradients within the oocyst causing differential protein expression

Question 14

What is an example of the maternal mRNA leading to the formation of differential protein expression in drosphila?

Answer 14

The maternal gene bicoid regulates the expression of the zygotic gene hunchback asbicoid is a transcription factor that can bind to the promoter of the hunchback gene and switch on its transcription
With no copies, theres no gradient of bicoid protein set up and also no gradient of hunchback
With 1 copy, theres a small bicoid gradient and a small hunchback gradient
With 2 copies theres a large bicoid gradient and a large hunchback gradient

Question 15

What is the heirachy of gene expression in drosophila?

Answer 15

maternal genes are switched on first, which act to regulate the expression of the zygotic gap genes, which regulate the pair-rule genes, which regulate the segment polarity genes, which regulate the selector genes. This results in the embryo being broken down and specified into progressively smaller and smaller parts.

Question 16

What are the unique features of early development in mammals?

Answer 16

Slow cell development
No maternal factors
Rotational cleavage
Asymmetric cell division generates two cells which inhabit 2 distinctive microenviroments
Cleavage is asynchronous

Question 17

What is the process between ovulation and implantation?

Answer 17

An oocyte is rleaased from the ovary during ovulation
the egg then travels down the fallopian tube where it is fertilized
Male and female pronuclei fuse causing the development of morula
Morula hatches out of the zona pellucida leading to the formation of a blastocyst which can implant on the wall of the uterus leading to the formation of a placenta

Question 18

What is the process of compaction?

Answer 18

The phase inbetween the 8 cell ball and the morula when cells increase in number but not in size leading to them flattening against each other which is mediated through the action of cell junctions containing e-cadherins, in this way the first epithelial layer was formed as the asymmetric cleavage results in the generation of more trophoblast cells and not more cells in the inner mass

Question 19

What are the functions of e-cadherins?

Answer 19

They allow the formation of adherens junctions and through their action with beta catenin can send signals to the cytoplasm

Question 20

What is achieved by the process of compaction?

Answer 20

Two layers of cells have been formed, the external trophoblast which will form the first epithelium which will become the placenta, as well as the inner cell mass which will eventually become the embryo proper and th amnion

Question 21

What occurs in the process of cavitation?

Answer 21

The formation of the blastocoel cavity
this occurs via passive Na+ movement across the apical membrane, it is then actively pumped across the basolateral membrane
Cl- and HCO3- is also pumped allowing for the preservation of electroneutrality
The change in osmotic pressure generated by the movement of electrolytes drives the movement of water this results in formation of the amniotic fluid

Question 22

What determines the final fate of cells in the morula?

Answer 22

The position of the cells and the number determines their fate.
If there are two few cells then they are all on the outer layer and no embryo will develop
Location will determine if it gives rise to the extra embryonic structures like the placenta or to the embryo proper

Question 23

When to cells stop becoming totipotent and only pluripotent? and what is the difference between these terms?

Answer 23

Totipotent means the cell can give rise to both the extra embryonic structures and all the cells in the body while pluripotent cells can only give rise to all the cells in the embryo proper
The cells reach this differentiation point when they undergo compaction

Question 24

What is gastrulation?

Answer 24

Cells gain motility and move inside the blastocoel cavity via the blastopore
This process is initiated by bottle cells constricting their apical ends resulting in a buckling of the epithelial sheet

Question 25

What is the process of gastrulation?

Answer 25

bottle cells invaginate to form the blastopore lip,
surface and adjacent cells involute to form endoderm and mesoderm,
surface stretching by epiboly,motility and repacking of internal cells

Question 26

How do cells change shape and gain motility in the process of gastulation?

Answer 26

Regulation and alteration of the cytoskeleton, typically mediated through the action of constriction through actin filaments

Question 27

What is the process of gastrulation in the sea urchin?

Answer 27

Blastula cells at the vegetal pole transform from epithelial to mesenchymal cells (primary mesenchymal cells PMCS) and ingress through the basal lamina into the blastocoel.

(B) Then there is an invagination of a central disc of cells, with some apical constriction, which pushes the cells inwards and begins the formation of the archenteron.

© The archenteron elongates to about 2/3 of the way up to the animal pole by convergent extension – a repacking of cells that results in an elongated shape. Once there, the cells at the advancing tip extend filopodia, search out the blastocoel surface.

(D) These filopodia form a kind of cable to tow the archenteron completely across the blastocoel.

(E) The archenteron roof finally touches the roof of the blastocoel and will become the future mouth of the sea urchin. Finally some involution of cells at the anal pore is evidenced, which populate the archenteron mainly along its posterior half. Also some proliferation

Question 28

What are the 8 morphogenic maneuvers performed by in embryo development?

Answer 28

Epiboly
Intercalation
Convergent extension
Invagination
Involution
Migration
Ingression
Proliferation

Question 29

What occurs in epiboly?

Answer 29

Flattening and spreading of cells,
allows for greater surface area
caused by increased cell-substratum adhesion

Question 30

What occurs in intercalation?

Answer 30

Cells from different layers spread out to form a single sheet
Results from changes in cell-cell adhesions
Forms multilayered tissues

Question 31

What occurs in convergent extension?

Answer 31

Elongation of tissue mass by extensive and concerted intercalation of cells
Medaited by changes in both cell-cell adhesions and cell-substratum adhesions

Question 32

What occurs in invagination?

Answer 32

Loacalized areas of cell constriction causing buckling and bending of the sheet

Question 33

What occurs in involution?

Answer 33

Folding of the cell layer under the original cell layer after invagination

Question 34

What occurs in migration?

Answer 34

Cells move away from the edges of a coherent mass due to a loss of cell-cell adhesion

Question 35

What occurs in ingression?

Answer 35

Cells detach from the epithelial layer to migrate into the extracellular matrix
Mediated by changes in cell-cell and cell-matrix adhesions

Question 36

What occurs in proliferation?

Answer 36

Cell multiplication which spreads a cell layer leading to localized growth, folding and buckling

Question 37

What is the Spemann organizer?

Answer 37

A region which express signalling molecules and their anatagonists
This specifies where the dorsal blastopore lip will form, as well as determining the anterior/posterior axis

Question 38

What is the Nieuwkoop centre?

Answer 38

Region of the blastula which specifies where the spemann organiser will form
the Nieuwkoop centre is specified when a sperm enters the egg, due to a cortical rotation of the cytoplasm under the egg membrane the Nieuwkoop centre is specifed as directly opposite the site of sperm entry and the future dorsal side
The spemann organizer and blastopore will develop in a region just above the Nieuwkoop centre

Question 39

What are the differences between an epithelial cell and a mesenchymal cell?

Answer 39

Mesenchymal cell are surrounded by extracellular matrix, have a flattened phenotype and a capable of migration
Epithelia cells however are polarised, contain specialised junctions and have keratin filaments

Question 40

What are somites?

Answer 40

Transient structure made of mesoderm which is closest to the neural tube
form the vertebrae and ribs, skeletal muscle of the back, body wall and limbs, amongst other things. They are also important in determining the migratory pathways of spinal nerve axons and other cells

Question 41

What are the processes involved in somite formation?

Answer 41

Somites begin to form from the anterior end and progressively bud off in an A-P direction.
The process of somite formation is one of epithelialisation, and then once formed, the epithelial cells then undergo EMT and de-epithelialise.

Question 42

What is an example of an EMT transition that occurs in Sea Urchin Embryos?

Answer 42

Formation of mesenchymal cells by ingression in to the blastopore
This involves the breakdown of the cells connection to other cells and to the surrounding hyaline layer (frequently seen in sea urchin species)
Holes form in the basal lamina, while the cells round and bleb to allow for ingression, the hole is quickly repaired after this movement

Question 43

What is the general EMT process?

Answer 43

Constriction of an adherens belt at the apical end of the cell
Down regulation of cell-cell and cell-ECM adhesion molecules
Up regulation of cell-mesenchymal adhesion molecules
Cell squeezes out of epithelial layer

Question 44

What are the signals which control EMT events in the drosophila?

Answer 44

Twist and Snail which are controlled by maternal proteins such as dorsal
Dorsal activates Twist and Snail expression
Twist acts to activate mesodermal genes, via the Rho GTPase pathway, which results in actin-myosin based cell changes.
Snail acts to inhibit genes that are associated with non mesodermal phenotypes, such as cell cycle proteins and also the E-cadherin/catenin adhesion complex genes.

Question 45

How are cell culture models useful in determining signals involved in an EMT event?

Answer 45

They can allow the effect of specific molecular signals to be determined as they can be genetically modified easily

Question 46

What induces EMT?

Answer 46

extracellular signals – growth factors, matrix molecules
 need specific receptors
 GTP-binding proteins (Ras, Rho, Rac)
 activation of MAP kinase and PI3 kinase
 src tyrosine kinase proteins

Question 47

What are the three ways in which adherens junctions can be dissociated?

Answer 47

decrease amount of E-cadherin (gene mutation, promoter repression)
phosphorylation of b-catenin (inhibits binding to cadherins)
inhibiting proteins that associate cadherins with actin (Rho effectors)

Question 48

How are cell-ECM junctions broken down in EMT?

Answer 48

reduction in integrin receptors on cell surface
interference with integrin function (inside-outside signalling)
induced protease activity (eg matrix metalloproteinases)

Question 49

What are the actions of the Slug/Snail transcription factor family?

Answer 49

zinc finger proteins
activated through Ras/MAPK signaling
causes EMT in MDCK cells in culture
represses E-cadherin transcription

Question 50

Where do we see conversion of mesenchyme to epithelium and how does this occur?

Answer 50

In the development of the kidney
The metanephrogenic mesenchyme induces the ureteric bud to elongate and branch.
The tips of the ureteric bud branches induce the metanephrogenic mesenchyme to form epithelial aggregates, Each aggregated nodule of about 20 cells will proliferate and differentiate into the intricate structure of a renal nephron (where blood is filtered).
At the junction between the ureteric bud tips and the epithelialising nodules, the nodules break down the basal lamina of the ureteric bud and fuse with them. This allows material to pass from the newly formed tubule into the ureteric bud, which becomes the collecting duct for urine.

Question 51

What signals induce metanephric mesenchyme to epithelialize?

Answer 51

FGF and LIF. FGF alone can’t induce epithelialisation, but in combination with LIF (leukaemia inhibitory factor), tubules will form

Question 52

What is the definition of a stem cell?

Answer 52

Cells that undergo asymmetric cell division to form one daughter that goes on to differentiate, and one daughter that retains its stem-cell properties
In this way Stem cells both maintain their own population, while creating a constant supply of specialized cells

Question 53

How does the mammalian epidermis demonstrate layers of differentially differentiated cells?

Answer 53

Right next to the basal lamina there are stem cells, these undergo asymmetric cell division leading to transient amplifying cells which will eventually detach from the basal lamina and form the intermediate layer where cells are still large and active, eventually these will form the dead differentiated cells that have lost their nuclei and become filled with keratin and involucrin

Question 54

How do neuroblasts undergo asymmetric cell division in drosophila?

Answer 54

Neuroblasts undergo asymmetric cell division to give rise to another neuroblast and a smaller ganglionic mother cell which will differentiate to form two neurons
This asymmetric cell division is driven by the protein concentrations of Miranda (localised to the basal side in anaphase after being in the apical side during interphase) and numb (localized to the basal side during anaphase)

Question 55

What are the key development points during neuroblast development?

Answer 55

Presumptive neuroblast in the ventrolateral ectoderm accumulates numb.
This presumptive neuroblast delaminates from the ventrolateral ectoderm into the embryo.
The mitotic spindle rotates 90 degrees such that its in an apical-basal orientation and
Numb is localised to the basal GMC
After division into another NB and the GMC, the NB again begins to accumulate numb and divides further into another NB and GMC
The GMC divides into two neurons, each of which accumulate numb

Question 56

How can stem cells be used to generate a transgenic mouse?

Answer 56

Isolate single cells from a 
blastocyst of parents
Clone and grow a single cell for many 
generations. These are ES 
(embryonic stem) cells,
Transfect ES cells with a transgene – including an
antibiotic resistance marker,
Inject selected cells into blastocysts of 
Different parents
Resulting pups will
be chimeras

Question 57

What keeps stem cells potent?

Answer 57

Oct 4, as well as Lif which activates STAT3, Nanog