Lecture 9

Question 1

What are the two main problems faced during segmentation

Answer 1

Organism needs to increase from one cells to many cells and makes these cells different from each other

Question 2

Segmentation is highly conserved in all organisms, T or F

Answer 2

T

Question 3

Where is segmentation easily visible in the Drosophila embryo

Answer 3

IN the anterior abdomen

Question 4

When does segmentation occur in Drosophila embryos

Answer 4

Around 24 hours after fertilisation

Question 5

The Drosophila embryo must set up all of the body axes in order to undergo segmentation, T or F

Answer 5

F – the anterior-posterior and dorsoventral axes have already been partially established in the oocyte by the adult fly. The embryo just needs to define and redefine this pattern

Question 6

What was the contribution of Christiane Nüsslein-Volhard and Eric Wischaus to the study of Drosophila

Answer 6

They undertook a saturation genetic screen to identify all the genes involved in development and patterning of the larval cuticle. This lead to the identification of 4332 embryonic lethal mutations and 139 complementation groups involved in patterning

Question 7

Most genes in development require both alleles to be functional to be expressed properly and provide normal function (haploinsufficient), T or F

Answer 7

F – most genes involved in development are haplosufficient

Question 8

Describe how complementation testing can be implemented to determine if mutations lie in the same gene or different ones

Answer 8

If you cross two parent individuals that both have the same mutant phenotype you can determine if these mutations lie in the same gene or different genes. If the progeny produced by breeding these two heterozygotes do not show the mutant phenotype, then the mutations are said to complement each other. If 25% of the progeny do show the mutant phenotype, then the mutations of the parents must lie in the same gene and thus fail to complement each other

Question 9

Recall the hierarchy of genes that dictate patterning in the Drosophila embryo

Answer 9

Maternal genes–> Gap genes–> Paired-rule genes–> Segment polarity genes

Question 10

What type of gene is gooseberry

Answer 10

Segment polarity gene

Question 11

What type of gene is bicoid

Answer 11

Maternal gene

Question 12

What type of gene are knirps, giant, Krupel and tailess

Answer 12

Gap genes

Question 13

Paired and fushi tarazu are hox genes, T or F

Answer 13

F – they are paired-rule genes

Question 14

Where is paired expressed in the Drosophila embryo

Answer 14

In alternating parasegments

Question 15

Gooseberry is expressed in alternating parasegments, T or F

Answer 15

F – it is expressed in all parasegments

Question 16

Genes higher up the patterning hierarchy have a greater influence over segmentation, T or F

Answer 16

T

Question 17

Give an example of another maternal gene, other than bicoid and explain where its expressed and its role in development

Answer 17

Nanos is another maternal gene. It is expressed in posterior embryo and is responsible for patterning the posterior larvae

Question 18

What is seen in bicoid mutants and why is this

Answer 18

Bicoid mutants develop without head structures. This is because bicoid is at its highest concentration at the anterior end and dictates formation of the head structures

Question 19

What is unique about bicoid and its role in development

Answer 19

Bicoid is a morphogen but is also in itself a transcription factor, unlike other maternal and patterning genes that are usually just transcription factors.

Question 20

What is the role of bicoid in establishing the anterior-posterior axis prior to fertilisation

Answer 20

Bicoid mRNA is deposited in the anterior oocyte by the adult female fly in a process called maternal loading. This leads to the localisation of bicoid protein at the anterior region of the embryo

Question 21

Describe the bicoid gradient in the early Drosophila embryo

Answer 21

Bicoid has its highest concentration at the anterior region of the embryo and drops away towards the middle

Question 22

What unique feature of the Drosophila embryo allows the bicoid gradient to be established easily and accounts for the simple early patterning

Answer 22

At this stage in development the Drosophila embryo is a syncytial blastoderm whereby many nuclei are contained in the same cytoplasm. This allows the bicoid protein to diffuse easily through the embryo and establish a gradient easily

Question 23

Explain the transplantation experiments carried out on the Drosophila embryo and what they showed

Answer 23

Transplantation of wild type Drosophila embryo cytoplasm into a bicoid mutant was sufficient to rescue some of the head structures. Transplantation of wild type cytoplasm to an ectopic site in the middle of the embryo lead to a duplication of the embryo with head structures developing in the middle flanked by thoracic segments either side. This shows that bicoid is both necessary and sufficient to dictate head structure formation

Question 24

What happens if both copies of the bicoid gene are mutated, why is this

Answer 24

If both copies of bicoid are mutated then the embryo will try to pattern normally using other maternal genes that are functional such as nanos. However the resultant patterning would still be abnormal

Question 25

What is the effects of forced overexpression of bicoid

Answer 25

Shunting of the segments towards the posterior end

Question 26

With morphogens, the ligand doesn’t act as the signal to dictate differentiation as such but it is in fact the gradient of the morphogen that encodes information on cell fate, T or F

Answer 26

T

Question 27

How can bicoid expression be visualise in situ

Answer 27

Fusing reporter genes for chaperones such as Hsp70 or genes easily stained for such as LacZ to the promoter sequence for bicoid allows easy visualisation. Wherever bicoid is expressed the reporter gene will be to as its under the control of the same promoter sequence. These reporter genes can then be easily visualised

Question 28

What attribute of morphogens accounts for there ability to be used to carry out multiple different functions

Answer 28

The there are different affinity sites for transcription factor morphogens such as bicoid. Some sites in the DNA have low affinity for the transcription factor ligand and so will only elicit binding at really high concentrations. However, other sites will have high affinity and will always elicit binding of the morphogen regardless of concentration

Question 29

What is the role of gap genes

Answer 29

Subdivide the embryo into different parts once the basic pattern is established

Question 30

What is significant about gap gene interactions

Answer 30

The gap genes actually also interact and repress each other as well as their targets

Question 31

Hunchback is a gap gene that regulates points in the embryo where other gap genes are switched on, what is significant about its expression with regards to other maternal gene(s)

Answer 31

Hunchback expression directly mirrors bicoid

Question 32

How are the paired-rule genes expressed in the Drosophila embryo

Answer 32

Expressed in alternating parasegments whereby their expression is controlled stripe by stripe

Question 33

What is paired-rule gene expression dependant on

Answer 33

Interactions of positively and negatively acting transcriptional regulators, many of which are gap genes

Question 34

Even-skipped is a paired-rule gene only expressed in parasegments 3. What conditions are required for even-skipped expression

Answer 34

Very low concentrations of giant and kruppel, high concentrations of hunchback and a little bit of bicoid

Question 35

Describe the expression pattern of segment polarity genes

Answer 35

Segment polarity genes are expressed in all 14 parasegments

Question 36

What has happened in the Drosophila embryo by the time that the segment polarity genes are expressed, and what are the implications of this

Answer 36

Cellularisation has occurred meaning that signalling pathways dictating patterning and development now need to get more complicated. Factors used for signalling now need to be secreted

Question 37

Engrailed is a segment polarity gene that is always expressed in the very anterior part of the segment, T or F

Answer 37

F – this was initially thought to be the case but in fact, engrailed is aways expressed most posteriorly in the segment

Question 38

What common phenotype is seen in wingless and hedgehog mutants in Drosophila and why is this

Answer 38

Both mutants are said to have a lawn of denticles in the larvae. This is because both genes maintain each other, hh maintains wg which supresses denticle development

Question 39

What is the function of wingless and hedgehog together

Answer 39

They work together to inhibit the formation of the denticles

Question 40

The segment polarity gene engrailed is switched on by wingless, what is the role of engrailed

Answer 40

Engrailed then switches on hedgehog

Question 41

Explain how wingless and hedgehog interact

Answer 41

Hedgehog signals in a paracrine manner to the adjacent cell to switch on wingless expression. Wingless then acts on its own receptors in an autocrine manner to upregulate is expression but also acts via engrailed to upregulate hedgehog too

Question 42

Hh and wg feedback onto each other to maintain their expression and refine segment borders, T or F

Answer 42

T

Question 43

The hedgehog-wingless signalling loop is present in in the posterior parts of the segments, T or F

Answer 43

T

Question 44

Wg is expressed uniformly throughout the Drosophila embryo, T or F

Answer 44

F – its expressed as a gradient

Question 45

What is the role of the Hox genes (homeobox domain containing genes)

Answer 45

Hox genes provide identity once the segments have already formed

Question 46

Hox genes are transcription factors, T or F

Answer 46

T

Question 47

From which genes do the hox genes receive input from

Answer 47

Pair rule and gap genes

Question 48

What is unique about the expression of the hox genes

Answer 48

They are expressed along the anterior-posterior axis in the same order in which they lie in the genome

Question 49

What happens due to a of loss of function in a hox gene

Answer 49

The segment will adopt a different identity, this is known as a homeotic transformation

Question 50

What is meant when Drosophila are referred to as long germ band organisms

Answer 50

All 14 segments are defined at once. All tissues are present they just need defining during development. This process is relatively quick (24hours in Drosophila)

Question 51

How do short germ band organisms develop differently to long germ bands

Answer 51

Similar process to short band organisms in the anterior head and thoracic segments. However, the abdominal segments are added sequentially by a region known as the posterior proliferative disc budding off segments as it gets smaller

Question 52

Short germ band segmentation is more complex and slower than that of long band organisms, T or F

Answer 52

T

Question 53

What signalling pathway mediates the segmentation of short band organisms

Answer 53

Delta-Notch

Question 54

Explain how the segmentation clock of delta, notch and Hes1 accounts for segmentation in short germ band organisms

Answer 54

Notch and delta signal to each other in a juxtacrine manner. Activation of the Notch receptor leads to the expression of Her/Hes1 which inhibits delta expression in the nucleus of that cell. At high concentrations Her also inhibits itself. This mechanisms acts as a molecular oscillator or clock whereby notch activity rises and then triggers the expression of Her. Her then switches off delta and then itself. This causes notch to rise again and the process repeats. This pattern the segments at exactly the right time as the organism grows based on the time taken to produce Her mRNA and then translate it and switch delta off and then itself

Question 55

How is the segmentation clock relevant to vertebrates

Answer 55

Humans are short band organisms. The primitive streak in vertebrates also use the notch pathway and its oscillations to produce and pattern body segments