The On’s and Off’s of Gene Expression

1. The On’s and Off’s of Gene Expression

2. Gene expression is regulated • The fact: • Virtually every cell in an organism contains the same DNA and the same genes. • The big questions: • Why are liver cells liver and not brain? • Why are leaf cells leaf and not root? • The answer: • Not all genes are expressed in all cells.

3. Why regulate gene expression? • To conserve energy • To respond to intracelluar signals • To respond to environmental conditions

4. How does gene regulation occur? Genes can be regulated anywhere in the process of information transfer. DNA mRNA protein protein function Pierce, B. 2005. Genetics, a conceptal approach. 2nd Ed. WH Freeman.

5. Organization of a transcription unit • Promoter: site for RNA polymerase binding • RNA coding region • Transcription termination signals Pierce, B. 2005. Genetics, a conceptal approach. 2nd Ed. WH Freeman.

6. What makes a promoter? In prokaryotes, DNA sequences--located about 10 and about 35 bases upstream of the transcription start site--serve as binding sites for RNA polymerase. Pierce, B. 2005. Genetics, a conceptal approach. 2nd Ed. WH Freeman.

7. In prokaryotes, regulation is pretty simple • Prokaryotic transcription is controlled by binding of RNA polymerase to the promoter. • Two options • If RNA polymerase is bound, gene is transcribed. • The gene is “on”. • The gene is “expressed”. • If RNA polymerase does not bind, gene is not transcribed. • The gene is “off”. • The gene is “not expressed”,

8. How is RNA polymerase binding controlled? • Negative regulation • Interferes with RNA polymerase binding • Binding of repressor proteins • Positive regulation • Promotes RNA polymerase binding • Binding of activator proteins.

9. Negative regulation example: lac genes • In Escherichia coli, the lac genes are needed to use the sugar lactose as a carbon source. • Expression of the lac genes is regulated. • The genes are expressed only if lactose is in the growth medium. • The genes are not expressed if glucose is present instead. • When glucose is present, the lac genes are turned off by a repressor protein.

10. The lac repressor binds DNA • lac repressor binds to a DNA sequence called the operator. • The operator sequence overlaps the promoter. • When repressor binds to the operator, it interferes with RNA polymerase binding to the promoter. Pierce, B. 2005. Genetics, a conceptal approach. 2nd Ed. WH Freeman.

11. Eukaryotic gene regulation is complex • Three types of RNA polymerase. • RNA polymerase I transcribes the large ribosomal RNAs. • RNA polymerase II transcribes mRNAs. • RNA polymerase III transcribes tRNAs and small ribosomal RNAs. • Each polymerase recognizes its own promoter.

12. RNA polymerase II promoters • Core of the promoter contains binding sites for basal transcription machinery • Regulatory part of the promoter contains binding sites for regulatory proteins Pierce, B. 2005. Genetics, a conceptal approach. 2nd Ed. WH Freeman.

13. Transcription Factors • Positive activators of transcription • Basal transcription factors bind to core promoter or to each other to facilitate binding RNA polymerase II. This positions RNA polymerase II for transcription. • Regulatory transcription factors bind to regulatory promoter elements. Their binding permits transcription by RNA polymerase II.

14. Assembly of basal transcription machinery Pierce, B. 2005. Genetics, a conceptal approach. 2nd Ed. WH Freeman.

15. Initiation of transcription Pierce, B. 2005. Genetics, a conceptal approach. 2nd Ed. WH Freeman.

16. Modular nature of regulatory promoters Different sequence motifs can be “mixed and matched” to recruit binding by various transcription factors. Pierce, B. 2005. Genetics, a conceptal approach. 2nd Ed. WH Freeman.

17. Expression of transcription factors • Constitutive • Expressed all the time, in many cell types • Examples include factors for: glucose metabolism; RNA and protein synthesis; ATP synthesis • Regulated • Expressed at specific times, in specific places • Examples include factors that respond to various signals (internal or external)

18. Anthocyanin pigment synthesis in corn • Synthesis of purple anthocyanin pigments in corn can occur in the kernel or in the plant. • The tissue-specificity is due to expression of different regulatory transcription factors. Karen Cone

19. Tissue-specificity of anthocyanin synthesis • Kernel-specific • Genes in the pigment pathway are turned on by the colored kernel (C1) transcription factor. • C1 is only expressed in the kernel, not in the plant. • Plant-specific • Genes in the pigment pathway are turned on by the purple plant (PL1) transcription factor. • PL1 is expressed only in plant organs, not in the kernel.

20. Environmental regulation of anthocyanin synthesis Light-dependent (“sun-red”) pigmentation MaizeGDB.org

21. What causes the “sun-red” pigmentation? turns on expression Light Light-dependent Transcription Factor pl1 gene LRE* TATA Transcription promoter * Light-responsive element

22. Other environmental cues and effects • Water • Too much (flooding): wilting, yellowing, death • Too little (drought): wilting, yellowing, death • Wilt in motion http://plantsinmotion.bio.indiana.edu/plantmotion/vegetative/veg.html http://www2.hawaii.edu/~coffee/drought2.jpg Coffee plant under drought stress

23. Other environmental cues and effects • Nitrogen deficiency Coffee plants: Normal on left, nitrogen-deprived on right http://www2.hawaii.edu/~coffee/y-nitrogen1.jpg

24. Other environmental effects on gene expression Light effects on development of bean plants http://facstaff.bloomu.edu/chamuris/concepts2/pics/bean_etiol1.jpg

25. Pathogen-induced changes in gene expression Susceptible pepper plant infected with bacterial pathogen Disease resistant pepper plant inoculated with bacterial pathogen Disease spreads through leaf. Expression of plant resistance gene limits spread of bacteria. http://www.apsnet.org/education/LessonsPlantPath/BacterialSpot

26. Examples of intracellular modulators of gene expression • Hormones • Auxin in plants regulates phototropism (growth towards light) • Sex hormones in animals control sexual development • Growth factors • Small molecules • cyclic AMP • Ca++ • Lipids

27. Summary • All cells of an organism contain the same DNA and thus the same genes • Not all of the genes are expressed in every cell. • Some genes (housekeeping) are expressed in many cell types. • Expression of most genes is regulated in response to environmental or cellular signals.