1. Gene regulation Ch 8 pp 280 -292

2. 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)

3. Definition 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.

4. 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

5. The 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)

6. Act 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

7. 08_16_anterior_posteri.jpg

8. Act 2 Labeling of the 4 proteins - Bicoid, Hunchback, Giant, Kruppel with fluorescent dyes reveals that they are not randomly located inside the cell...

9. 08_17_4.gene.reg.prot.jpg

10. Act 2 continued The nuclei are therefore bathed in differing concentrations of these four proteins - depending on their position in the cell.

11. 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

12. 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!

13. 08_18_reporter.gene.jpg Isolate the different regions from the promoter and test in embryo.

14. 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.

15. 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’

16. 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

17. 08_15_Reg. proteins.jpg

18. Combinatorial 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

19. Maintaining the status quo - with respect to gene regulation- through cell division

20. 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

21. When things go wrong…

22. 08_25_eye.on.leg.jpg Activation of the wrong regulator can have devastating effects One can move cells to new locations may lead to monsters!!

23. 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!!