Gene regulationGene regulation

Ch 8

pp 280 -292

What will I learn?What will I learn?

THAT;

1. Gene regulation is at the heart of development

2. The most important part of a gene is its regulatory region, because;1. It determines how, when, and by how much a

gene is activated

2. The combination of proteins which bind to this region determine activation (or repression)

Definition of gene regulationDefinition of gene regulation

• Regulation of gene expression (gene regulation) is the cellular control of the amount and timing of the appearance of the functional product of a gene.

The Story of ‘Eve’The Story of ‘Eve’

• This example illustrates why gene regulation is fundamental to development

• The Players– Drosophila embryo– Even-skipped gene– Regulatory proteins

• Bicoid• Hunchback• Giant• Kruppel

The setupThe setup

• In Drosophila, the fertilized egg initially performs many rounds of mitosis without cell division - resulting in a cell with many nuclei

• The embryo is 400 um long and 160 um wide• It has anterior (that will eventually develop

into the head) and a posterior (that will develop into the abdomen)

Act 1Act 1

• Removing the cytoplasm from the anterior will result in the failure to develop a head

• Replacing this cytoplasm with some taken from the posterior of another embryo will result in an embryo with two tails

08_16_anterior_posteri.jpg

Act 2

• Labeling of the 4 proteins - Bicoid, Hunchback, Giant, Kruppel with fluorescent dyes reveals that they are not randomly located inside the cell...

08_17_4.gene.reg.prot.jpg

Act 2 continued

• The nuclei are therefore bathed in differing concentrations of these four proteins - depending on their position in the cell.

Act 3

• Even-skipped - ‘EVE’

• It is a master regulatory gene whose product is a master regulatory protein

• This genes’ promoter region has the ability to bind to all four of the regulatory proteins

Act 4 The Reporter

• A reporter gene is an artificial DNA construct used to reveal information

• Lac Z reporter gene is used extensively• GAL (galactosidase) - Hydrolyzes colorless

galactosides to yield colored products. • Attach different parts of the promoter region of

the ‘eve’ gene to LacZ to determine if that region is bound and activated by proteins.

• Check the web on this please!

08_18_reporter.gene.jpgIsolate the different regions from the promoter and test in embryo.

08_19_eve.stripe.2.jpg

The same STRIPE 2 region actually has binding sites for all four of regulatory proteins - Bicoid and Hunchback are activators of this region and Giant and Kruppel are repressors.

Curtain

• The regulatory region of ‘Eve’ extends more than 20,000 bp

• It is thought to bind more than 20 different proteins

• It is very sensitive to the position of the gene (nucleus) within the developing giant cell

• The different concentrations of the different proteins impact on the expression of ‘Eve’

Gene Expression

• Regions huge distances from the gene have an effect on the activation (or repression) of that gene

• These regions bind proteins that then interact, by DNA looping, with the local promoter regions

• A combination effect is seen - and thus the term combinatorial control is used

08_15_Reg. proteins.jpg

Combinatorial controlCombinatorial control

• Not just by how much are genes regulated but also when.

• It is akin to an orchestra….

• It is fantastic coordination at each gene locus

Maintaining the status quo- with respect to gene

regulation- through cell division

Maintaining the status quo- with respect to gene

regulation- through cell division

08_24_chromatin.state.jpg

Inactive genes are kept inactive by regulatory proteins bound to the DNA, and active genes are kept active! How, see below

When things go wrong…When things go wrong…

08_25_eye.on.leg.jpgActivation of the wrong regulator can have devastating effects

One can move cells to new locations may lead to monsters!!

Other examples of developmental issues.

Here either antenna, or proboscis is growing on the head,Or dual thorax segments with a pair of wings on each!!