lecture 23

Question 1

What is pattern formation?

Answer 1

• the process by which a spatial and temporal pattern of cellular activities is organised within the embryo so that a well ordered structure develops
• development of a body plan, axes

Question 2

What happens if there are errors in pattern formation?

Answer 2

• can have severe consequences for the development of an organism
• e.g. duplicated thorax in fly
• fusions of digits, knuckles

Question 3

What are simple stages in development of an organism?

Answer 3

• zygote
• cleavage
• blastula
• 1. gastrulation
• 2. neurulation
• neurula
• segmentation (somitogenesis)
• determination of segment identity
• Hox genes

Question 4

What is the definition of determination?

Answer 4

Formal definition

• determination implies a stable change in the internal state of a cell such that its fate is now fixed, or determined
• a determined cell will follow that fate when grafted into other regions of the embryo
• cells often “know” where they are positioned in a developing embryo before morphological differentiation occurs
• they are determined

Question 5

What is an experiment that shows the pre-somitic mesoderm has a positional identity prior to somite formation?

Answer 5

• with chick embryos
• the segmental plate mesoderm is determined as to its position along the anterior-posterior axis before somitogenesis
• when segmental plate mesoderm that would ordinarily form thoracic somites is transplanted into a region in a younger embryo (caudal to the first somite) that would ordinarily give rise to cervical (neck) somites
• the grafted mesoderm differentiates according to its original position and forms ribs in the neck

Question 6

By what is the positional identity of somites along the antero-posterior axis specified?

Answer 6

• Hox genes
• e.g. head to tail separated to four regions: W, X, Y, Z
• gene I expressed in W, X, Y, Z
• gene II expressed in X, Y, Z
• gene III expressed in Y, Z
• gene IV expressed in Z
• therefore each region has/is defined by the expression of certain numbers of these genes

Question 7

What are Hox genes?

Answer 7

• are transcription factors
• contain a DNA-binding region of approx. 60 amino acids called the homeodomain (contain a helix-turn-helix DNA-binding motif)
• the DNA sequence of 180 base pairs that encodes the homeodomain is called the homeobox (hence the name)
• first identified in the fruitfly, Drosophila
• required for anterior-posterior positional identity in flies and mammals (and all other segmented animals)

Question 8

Why are they called homeobox genes?

Answer 8

• homeobox received their name because mutations in these genes can result in homeotic transformations
• i.e. one body segment is transformed into another with different positional identity

Question 9

Are clusters of homeotic genes only found in the fly?

Answer 9

• no
• while clusters of homeotic genes were first identified in the fly
• similar clusters were subsequently identified in vertebrates
• known as Hox complexes
• in the mouse there are four Hox clusters that have arisen during evolution of vertebrates by duplication of an ancestral cluster
• therefore, corresponding genes within the four clusters resemble each other closely (known as paralogs)
• the similarity is most pronounced within the homeobox

a point of confusion:
- most genes that contain a homeobox are not part of a Hox complex and are not associated with homeotic transformations

Question 10

What are orthologs cf paralogs?

Answer 10

• ortholog = similar gene between species

- paralog = similar gene with in an organism’s/species’ genome

Question 11

How are Hox genes aligned in the drosophila?

Answer 11

• along chromosome from 3’ to 5’
• labial - expressed in the head
• expression of genes aligns with their position along the chromosome
• matches anterior to posterior structures of the fly
• collinearity
• theoretically there’s no reason why this should be the case in genes

Question 12

What are features of hox gene expression?

Answer 12

• Hox genes typically have sharp anterior boundaries of expression and less well defined posterior boundaries
• “anterior” genes are expressed first
• genes found more 3’ in the cluster are expressed more anteriorly
• collinearity

Question 13

How do we know that Hox genes influence segment identity?

Answer 13

• appear to be expressed in the right regions
• particular Hox genes specify particular regions in mouse and chick, even if the number of segments in each region is different
• e.g. boundary between expression of hox 5 and hox 6 is found at the boundary between cervical and thoracic vertebrae in both the chick and the mouse despite the chick having 14 and the mouse having 7 cervical vertebrae (or sometimes that give rise to these vertebrae)

Question 14

What is posterior dominance?

Answer 14

• aka posterior prevalence
• the more posteriorly expressed Hox genes tend to inhibit the action of Hox genes expressed more anteriorly to them
• hence back to our 4 region example
• in region Z, gene I, II, III and IV are expressed but if gene IV inhibits all the three genes anterior to it, the only functioning gene in region Z is therefore gene IV

Question 15

What do you have to consider when doing KO mice for hox genes?

Answer 15

• they have paralogs, so if you want to see knockout function you have to ko all paralogs

Question 16

What happens in KO mice?

Answer 16

• the Hox region knocked out will express the more anterior phenotype

Question 17

What happens in overexpression of a particular hox gene?

Answer 17

• anterior to posterior transformation

Question 18

Have Hox genes evolved? What does this mean?

Answer 18

• yes hox gene complexes have evolved through evolution
• changes in Hox gene expression may have assis ted evolution of specialised body segments
• e.g.
  X (branchroot?): whole lot of similar body segments that express three different hox genes throughout
  grasshopper - separation and distinct expression of Hox genes leading to distinct body segments

the combination of Hoxc-6 and Hoxc-8 promotes rib formation on vertebrae - hence snakes do not develop forelimbs (hindlimbs were lost via a different mechanism)

Question 19

What other axis do hox genes also pattern (other than anterior-posterior)?

Answer 19

• Hox genes also pattern proximo-distal axes
• e.g. limbs
• hox expression along limbs that are characterised by different bone structures
• e.g. losing a hox gene will result in loss of a particular segment of limb (e.g. ulna/radius – Hoxa11, Hoxd11-deficient mutant)
• HoxD13 mutant – mutations in bones of wrist and hand