Chapters 19 - Genetic Analysis of Development:

Development

Development refers to interaction of then genome with the cytoplasm and external environment to produce a programmed sequence of typically irreversible events.

Differentiation

Differentiation refers to the formation of cell types, tissues, and organs through specific gene regulation. A single cell with one genotype produces a variety of specialized tissues and organs.

Development and differentiation can be studied at many levels:

1. Morphology

2. Biochemistry

3. Genetics

Model systems for the study of development and differentiation and why they are studied:

• Drosophila melanogaster (fruit fly)

• Long tradition of study.• Lots of mutations affecting development identified.

• Caenorhabditis elegans (nematode)

• Genome is small (97 Mb)• # and lineage of body cells are known• Genetic crosses and selfings are easy• Body is transparent

• Brachydanio rerio (zebrafish)

• Embryos are transparent.• Large #s of fish can be bred.• Screening techniques are well-developed.

• Arabidopsis thaliana (plant)

• Small, easy to cross and analyze large numbers of progeny.• Many developmental mutations identified.

Fruit Fly (Drosophila melanogaster)

NematodeCaenorhabditis elegans

Fig. 19.2Showing development from the two-cell stage to the adult.

Fig. 19.4, Zebrafish (Brachydanio rerio)

Fig. 19.3, Arabidopsis thaliana

Genetic regulation of development in Drosophila:

Fig. 19.18,

Developmental stages of Drosophila(10-12 days)

Egg

Larva (3 instars)

Pupa

Adult

Embryonic development in Drosophila:

• Development begins with fertilization.

• Prior to fertilization, molecular gradients exist within the eggs. Polar cytoplasm occurs at the posterior end---example of maternal effect.

• 2 nuclei fuse after fertilization to form a zygote.

• 9 mitotic divisions occur without cell division, and after 7 divisions, some nuclei migrate to the polar cytoplasm (posterior) creating germ-line precursors.

• Other nuclei migrate to the cell surface and form blastoderm precursor.

• 4 more mitotic divisions occur and all nuclei are separated by cell membranes.

Fig. 19.19, Embryonic development in Drosophila.

Subsequent development depends on two processes:

Fig. 19.20,

Adult segmentation reflectEmbryo segmentation

1. Anterior-posterior and dorsal-ventral molecular gradients exist in the egg---mRNAs and proteins placed in egg by mother confer maternal effect.

2. Formation of (1) parasegments and (2)embryonic segments, which give rise to (3) adult segments.

Three major classes of genes control development and differentation

*Mutations identified by presence lethal or abnormal structures during development.

1. Maternal effect genes

2. Segmentation genes

3. Homeotic genes

1. Maternal effect genes

Expressed by the mother during egg production; they control polarity of the egg and the thus embryo.

bicoid gene

• Regulates formation of anterior structures (mutants possess posterior structures at each end).

• Gene is transcribed during egg production, and expressed after fertilization.

nanos gene

• Regulates abdomen formation (mRNAs collect in posterior of the egg).

torso gene

• Transcription and translation occur during egg production.

• Occurs throughout the eggs, but is only active at the poles.

Fig. 19.24Distribution of bicoid mRNA and protein in the egg

A = Anterior

P = Posterior

2. Segmentation genes:

Determine the segments of the embryo and adult, and thus divide the embryo into regions that correspond to the adult segmentation patterns.

1. Gap genes Subdivide the embryo along the anterior-posterior axis.

Mutation results in the deletion of several adjacent segments.

2. Pair rule genes Divide the the embryo into regions, each containing

parasegments.

Mutations cause deletions of the same part of a pattern in every other segment.

3. Segment polarity genes Determine regions that become segments of larvae and

adults

Mutants possess parts of segments replaced by mirror images of adjacent half segments.

Fig. 19.25, Functions for segmentation genes defined by mutations.

3. Homeotic genes:

• Homeotic genes specify the body part to develop at each segment.

• Adult body parts develop from undifferentiated larval tissues called imaginal discs.

• Homeotic mutants develop a different body part at a particular segment (imaginal disc) than the usual body part.

• Different homeotic gene groups share similar sequences of ~180 bp called homeoboxes that code proteins.

• Homeoboxes regulate development and produce proteins that bind upstream of the gene units.

• Homeotic gene complexes are abbreviated Hox.

• Hox genes also specify body plans in vertebrates and plants.

Fig. 19.21, Locations of homologous imaginal discs in larva and adult.

Fig. 19.26

Examples of homeotic Drosophila mutant with the bithorax mutation

Fig. 21.27, 2nd edition

Antennapedia and aristapedia mutants

Fig. 19.1, Second set of eyes in place of antennae:

Fig. 19.28, Organization of bithorax homeotic genes in a 300kb region of the Drosophila genome.

T = thoracic A = abdominal

Fig. 19.29

Homologous Hox gene clusters occur in Drosophila and the mouse.

How do development biologists study differential expression of genes during development and differentiation?

Immunofluorescene assays that bind to specific mRNAs and proteins.

How do development biologists study differential expression of genes during development and differentiation?

Quantitative real-time RT-PCR of cDNA from mRNA transcripts.

http://www.nature.com/nbt/journal/v28/n5/full/nbt0510-421.html

RNA-Seq

Ribosome Profiling – sequencing of ribosome-bound mRNAs

How do development biologists study differential expression of genes during development and differentiation?

Gene knockout using transformation or transduction, or other gene silencing techniques like RNAi.

http://ja.wikipedia.org/wiki/RNAi RISC = RNA-induced silencing complex