Developmental Biology II(Molecular Mechanisms of Development)

Anke van Eekelen, PhDTelethon Institute for Child Health Research

Basic Principles of Development

1- Cell Division • multicellular organism

2- Pattern Formation • Defining the Axes: Body Plan

• initiating Germ Layer Formation

3- Morphogenesis • Formation of 3 Germ layers

4- Cell Differentiation • Blood, Muscle, Nerves …

5- Growth • maturity

Cleavage

Gastrulation

Organogenesis

Event Principle Outcome

Developmental events

are controlled by differential gene expression,

……..which drives cascades of gene-regulatory events,

…………………..which define differential cell behaviours,

………………………………which underlie major developmental processes

crucial to study gene expression & protein function

DNA

mRNA

Protein(e.g.transcription factors)

Differential gene expression:- type of enhancer

- processing of mRNA intodifferent splice product

- protein modification to createfunctional proteins

TF = transcription factordevelopmental specificcascades of gene regulatory events

Transcriptional protein complex:

= downstream gene(e.g. new TF!!)

During cleavage, gastrulation and cell differentiation:

‘Autonomous’ signals Cell Conditioning signals

Maternal & Zygotic determinants

Genes encoding Genes encodingtranscription factors secreted proteins(or factors affecting translation)

direct effect on gene indirect effect on gene expressionexpression in same cell in another (target ) cell

Wolpert: “The early development of Drosophila is probably the best understood developmental system of all animals, at the molecular level. This is due largely to a saturation mutagenesis screen for embryonic pattern defects carried out in the 1970s by C. Nusslein-Volhard and E. Wieschaus (Nobel Prize in 1995). Approximately 100 genes were identified which can account for most of the pattern formation and morphogenetic events of the early embryo. This set of genes complement a different set of genes studied by E. Lewis, whose activities were involved in specifying the identity of segments observable in the adult fly. Together, these genes became the Rosetta stones that allowed fly development to be deciphered. At the time, no one suspected that they would provide the keys to understanding development in all animals, yet that is what has occurred; even humans use the genetic networks first discovered in flies, albeit in sometimes very different ways. For this reason, Drosophila development deserves special attention in studies in developmental biology.”

Wolpert, Beddington, Jessell, Lawrence, Meyerowitz and Smith (2002)“Principles of Development” 2nd edition (Oxford University Press)

P (p

oste

rior)

bicoid (oocyte mRNA encoding a TF)

Nanos (translation repressor)

Patterning of A-P axis during cleavage event (Fruit fly) A

(ant

erio

r)

Putting A and P determinants together…..

hunchbackmRNA

Hunchbackprotein

Zygotic hunchback

The expression of each transcription factor in developmentat the right time in the right location

is regulated by a complex interactive network of gene regulatory events

Over time the type of expressed developmental specific TF and other determinants change!

Nature of protein : TF (different families of TFs based on structure)secretion proteinmembrane coupled receptor

Cell Signalling regulation of development

Interactive network ofdeterminants regulating

gene expression in neighbouring cells

e.g. Segment polarity genes

Cell Autonomous regulation of development

Interactive network ofdeterminants regulating gene expression within

cellse.g. Gap genes

Similarity between invertebrates and (higher)vertebrates:- many common genes- maternal determinants in oocyte- overlap in cascades of gene-regulatory events driving development

Why study flies……if we are mostly interested in development of mammals like

mice and humans?

Homeotic selector genes• Hom (flies) vs Hox in (mammals)• Homeobox family of TFs: highly conserved homeodomain DNA binding region• Specify order of type of structures along a line

(“memory” for the A-P order already established by earlier developmental specific genes)

Fruit fly: 1 cluster of Hox genesmammals: 4 Hox clusters, each preserving the homologies and order along the chromosome of the fly

cluster.

• Secreted factors noggin and chordin both bind to the TGF-beta type growth factor BMP-4.• The secreted factor Frizbee is related to Frizzled, the cell membrane receptor for Wnt-family

growth factors. It acts as a competitive inhibitor of Wnt/Frizzled signalling.

Example of the molecular mechanism underlying initiation of gastrulation in the frog:

The somite (Pax3 +ve) is target of numerous known signals from all its adjacent tissues at

the end of gastrulation and neurulation,

that sub-divide it into regions expressing distinct gene signatures. These regions are

specified for distinct fates.

Example of the molecular mechanism underlying mesodermal tissue differentiation (somite)in the chicken:

Various commonly used Signalling Pathways in developmental events of

tissue organization and organogenesisG

row

th fa

ctor

sign

alin

gH

edge

hog

sign

alin

g

Wnt

sign

alin

g

0

Organizer AER:• controls proximal to distal (ie shoulder to finger tip) growth & development

• morphogen = diffusible growth factors FGF4 and FGF 8

Tbx-5

Tbx-4

Proximal distal

mesenchym

ectoderm

AER

Example of the molecular mechanism underlying mesodermal limb formationin the chicken:

Example of cascade of gene regulatory events in heart development(organogenesis)

Example of crucial regulatory genes in muscle development

HLH TFs

How to study molecular mechanisms underlying development ?

previously- Mutagenesis in fruit fliescurrently- Genetic manipulation - gene knockout loss of function

Tutorial in week 6

Future research will be inclusive of Alternative ways to influence gene expression ?-Epigenetics

Lecture in week 5