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Summary Lectures molecular principles of development

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  • Course
  • Molecular Principles of Development (NWIBB084B)
  • Institution
  • Radboud Universiteit Nijmegen (RU)
  • Book
  • Principles of Development

All lectures of the course molecular principles of development (NWI-BB084B) are covered in the document (using powerpoint slides, additional information from the lectures and the book). The book used for this course was: molecular principles of development, the 6th Revised edition. The pictures inc...

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  • Unknown
  • February 13, 2020
  • 51
  • 2019/2020
  • Summary

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  • development
  • biology
  • radboud
  • medical
  • molecular principles

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  • Test Bank for Principles of Development 6th Edition by Lewis Wolpert, Cheryll Tickle, Alfonso Martinez Arias - Complete,Elaborated & Latest Test Bank. All Chapters [1-14] Included & Updated
  • Test Bank for Principles of Development 6th Edition by Lewis Wolpert, Cheryll Tickle, Alfonso Martinez Arias - Complete, Elaborated & Latest . All Chapters [1-14] Included & Updated
  • Test Bank For Principles of Development 6th Edition By Lewis Wolpert; Cheryll Tickle; Alfonso Martinez Arias 9780198800569 Chapter 1-14 Complete Guide .
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  • Radboud Universiteit Nijmegen (RU)
  • Biologie
  • Molecular Principles of Development (NWIBB084B)
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Lectures molecular principles of
development
2019-11-07 Basic concepts
Forward genetics
Choose mutations (phenotypical) and look for the genes belonging to these mutations. Drosophila
sperm was randomly mutated  search for the mutations with the help of a visual marker
(phenotype).
Patterning genes
There are different types of patterning genes in embryos:
1. Gap gene. Loss results in a reduced number of segments.
2. Pair-rule gene (even-skipped). Loss allows only odd-numbered segments to develop.
3. Segment polarity gene. Loss leads to segments with similar head and tail ends.
The patterning genes allow the genes to become segmented.
Hox genes
Important for patterning, deciding which part of the developing body the genes are present in.

Drosophila melanogaster
Life cycle
Fertilized egg  cleavage  syncytial blastoderm  gastrulation  larval stage  adult fly. The
process from fertilization to adult flies only takes 9 days.
A syncytium forms in early development. Sperm enters through the micropyle (anterior). The
drosophila egg is elongated: the micropyle on one end, the germ line on the other end. The
drosophila egg has polarity, which is not the case in all species.

Syncytium: early in development the nuclei divide but the cells do not. This leads to a ‘bag of
cytoplasm  syncytium. The syncytium allows for diffusion of proteins.
At a certain moment the syncytium will cellularize.
 First the nuclei are in the centre  move to the periphery  cellular blastoderm.
The pole cells will give rise to the germ line.

If there is a syncytium, molecules can freely diffuse: local source + diffusion.
Gastrulation and segmentation
Cleavage  blastoderm  gastrulation  germ band elongation  head involution and dorsal
closure  differentiation.

Gastrulation is the first morphogenetic process of development: process where the ball of cells turns
into an organism that has polarity and axes. Gastrulation can be defined as the main process in
development. At the end of gastrulation there is obvious morphological segmentation. Patterning is
important for this segmentation.

Drosophila versus vertebrate development
Maternal axes and symmetry Syncytium and Early embryonic cell
cleavages division time
Drosophil Anterior-posterior, AP Nuclear divisions 9 minutes
a Dorsal-ventral, DV Syncytium
Zebrafish Animal-vegetal (radial symmetry) Meroblastic divisions 15 minutes

, Yolk syncytial layer
Mammals No polarity (point symmetry)
Xenopus Animal-vegetal (radial symmetry) Holoblastic divisions 25 minutes
Mouse Holoblastic divisions 15-20 hours

There is a syncytium in drosophila. In zebrafish there is a syncytium underneath the embryo: the yolk
syncytial layer. This because zebrafish undergoes meroblastic divisions  no division through the
whole egg, but on top of the egg. Frogs and mice undergo holoblastic divisions  right through the
middle of the embryo.
When considering genome size and division time (relative speed) zebrafish is extremely fast.

Embryonic patterning
Embryonic patterning is the process of establishing positional information at the molecular level
among similar cells. It helps cells respond differently depending on where they are. The cells look
identical but start to differentiate in different ways.
Patterning establishes body axes:
- Dorsal-ventral (DV).
- Anterior-posterior (AP).
- Medial-lateral/left-right.
This all starts with polarity/symmetry-breaking. Some eggs are already patterned in some way.
Symmetry breaking can be done in multiple ways:
 Asymmetric cell divisions.
 Molecular gradients.

Patterning is not the same as differential gene expression. Gene expression differences, are not the
same as patterning. Patterning: the process which established differential gene expression (among
otherwise similar cell) that is directly related to the position in the embryo  e.g. AP, DV, LR. Gene
expression differences are however involved in gene expression. The patterning process will lead to
differential gene expression.
Patterning is an early developmental process.

Maternal genes
Maternal factors set up the body axes: AP and DV. In Drosophila, the oocyte is already patterned
before fertilization. Some RNAs are localized within the oocyte. When they are translated, they form
a local source and the proteins diffuse.
- Bicoid (anterior) inhibits caudal translation from maternal mRNA  both maternal.
Maternal genes are expressed in the oocyte before fertilization. A lot of the maternal genes are
expressed in gradients in the Drosophila embryo.
Zygotic genes
As soon as the oocyte starts to express its own genes, it starts to respond to the activity of maternal
proteins.
 Increased specificity and position.
 When knowing only the concentration of Bicoid and caudal, you can know exactly where in
the embryo you are  Bicoid (anterior) inhibits caudal translation from maternal mRNA.
Examples of zygotic genes:
- Gap genes: usually there is a gap in their expression pattern across the embryo.
- Pair-rule genes.
- Segmentation genes.
- Selector genes.
The activity of the proteins is organised in such a way, that it induces increased diversity. Normally,
there are multiple domains of expression of a particular gene.

,  Activity of both maternal and gap genes produces a striped pattern across the embryo.
Other examples of expression in early drosophila zygote:
- Transcription factor Fushi tarazu: embryonically expressed.
- Transcription factor Even-skipped: embryonically expressed.
- Transcriptional regulator Snail.
There are molecular differences between cells before morphological differences arise.

Germ layers
There are three germ layers formed in the embryo. They are the most basic cell types.
Endoderm Stomach, colon, liver, pancreas, urinary
bladder, part of urethra, epithelial parts of the
trachea, lungs, part of the pharynx, thyroid,
parathyroid, intestines.
Mesoderm Muscle (smooth + striated), bone, cartilage,
connective tissue, adipose tissue, circulatory
system, lymphatic system, dermis, genioto-
urinary system, serous membranes, notochord.
Ectoderm Surface epidermis: hair, nails, lens of the eye,
sebaceous glands, cornea, tooth enamel,
anterior pituitary, epithelium of mouth and
nose.
Neural crest: peripheral nervous system,
adrenal medulla, melanocytes, facial cartilage,
dentin of teeth.
Neural tube: brain, spinal cord, posterior
pituitary, motor neurons, retina.

From the three basic cell types, all cell types of the adult body are formed.

Fate maps
Fate maps tell what the progeny of cells will be. A cell will be fluorescently labelled and is followed
through development as it will stay fluorescent as it divides.
Faces in development:
1. Cell has not received any information on its progeny but they are positioned at a certain
place  fate.
2. The cell has received information on its progeny but it can still change  specification.
3. The cells’ progeny cannot change any more  determination.
If not determined, cells can adapt to a new environment.

 Transplantation of a tissue at an early stage  the cell changes fate to its new environment.
The cells were not determined yet.
 Transplantation of a tissue at a later stage  the cell won’t change its fate to its new
environment. The cells were already determined.

In situ hybridization
Detection of a protein: immunohistochemistry using antibodies.
Detection of RNA: in situ hybridization with RNA probes.
Whole mount in situ hybridization is performed in the whole embryo.

, Working mechanism: cloning a gene of interest  produce RNA of that + providing labelled
nucleotides. The labelled nucleotide in the RNA can be followed through development. For this
purpose, DIG labelling is used. The RNA has to be complementary to the wanted RNA. The
complementary RNA can anneal to the wanted RNA.
Working on the embryos:
- Fixing the embryos.
- Making the embryos permeable, so the probe can get into the fixed embryo.
- The probe is annealed to the RNA present in the embryo, the rest is washed out.
- The DIG molecule can be detected in the embryo.

Test questions
What is a syncytium and how does it influence the early development in Drosophila?
Nuclei without cell membranes, allowing for diffusion of proteins to pattern the embryo.

Wat is the essence of a forward genetics screens?
Introduction of random mutations, followed by screening of phenotypes.
Reverse genetics:
Introduction of a specific deletion in a gene of interest using homologous recombination.

How do you know you have reached saturation (all genes identified) in a forward genetics screen?
Most genes that you find, you find more than once.

Most of the variation in embryonic cell division time between species may be best explained by:
External versus intra-uterine development. Zygotes that develop external, unprotected by a parent,
have to develop more fast to survive.



2019-11-09 Axes and germ-layers 1
Early development
Start of development: cleavage division. The embryo does not grow in the beginning, but the cells
cleave  the size become smaller. The size of an oocyte is very diverse between organisms. A human
oocyte is about 0.1 mm in diameter (the largest cell in the human body). Sperm is the smallest cell in
the human body  just a package of DNA. The human oocyte is small in comparison to the oocytes
of other animals.
The maternal to zygotic transition (MZT)
The early stages are largely driven by maternal gene products: RNAs and proteins that were already
present in the oocyte. The earliest cleavage divisions, the embryonic/zygotic genome is not
transcriptionally active. The embryo does not transcribe its own genes  embryonic development is
completely relying on maternal genes.
After a while, the genes of the embryo are being transcribed: the maternal to zygotic transition
(MZT).
- The maternal RNA degrades.
- The zygotic genome activation (ZGA).
It is a variable amount of time and cleavage divisions between species for the MZT to occur:
- In mice it takes about 24 hours for the MZT to occur: 2 cell stage.
- In drosophila it takes about 1.5 hours for the MZT to occur: 10 cycles of division.

In externally developing species, the MZT is also called the mid-blastula transition (MBT). In these
species the MZT happens at the mid-blastula stage. Term MBT is used in flies, fish and frogs. What
happens during MBT:

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