Control Mechanisms

1. Control Mechanisms

2. (Laugh)

3. Review… • What do genes ultimately encode? • What is the intermediate between gene and polypeptide chain (protein)? • What are the various functions of proteins? • How many genes encode proteins? • Do you think all proteins are needed to be transcribed/translated at all times of the day? Why or why not?

4. So, cells need a way to turn genes on and off! • Transcription factors turn genes ‘on’ when required – what do we mean by ‘on?’  genes are expressed (transcribed and translated to protein) • This method of controlling the transcription and translation of genes is called gene regulation • Some proteins are always needed – called housekeeping genes

5. Levels of Gene Regulation • On your own, think of a way (identify a step or mechanism) where transcription or translation can be controlled (Hint: think of molecules that are involved in the production of proteins from genes)

6. As information flows from DNA to proteins… DNA  mRNA  protein activated protein transcription translation Post-translational modifications Transcriptional Control DNA  mRNA  protein  activated protein Translational Control DNA  mRNA  protein  activated protein

7. Four levels of Gene Regulation • Transcriptional – regulates which genes or the rate at which they are transcribed • Posttranscriptional – after the mRNA is transcribed, the modifications it undergoes can be regulated • Translational – regulates how often and how quickly mRNA is translated, which affects the time to activate/destroy mRNA • Posttranslational – regulate modifications that are made to newly formed proteins

8. Levels of Gene Regulation

9. Got Milk? • It was previously thought that the E. coliin our digestive system was responsible for our ability to digest lactose • However, it is now known that the enzyme lactase digests lactose • Nonetheless, the lac operon serves as an excellent model for understanding the concept of gene regulation superlaugh.com/dan/lactose.htm

10. A Prime Example – lac Operon Primary energy source in E. coli: Glucose However, E. coli may use other carbon sources (i.e.: lactose) if glucose is absent. lactose = Glucose + Galactose Therefore, E. coli can indirectly obtain glucose via cleavage of lactose by the enzyme β-galactosidase Note: β-galactosidase is only produced in the presence of lactose  WHY?

11. A Prime Example – lac Operon • Only in prokaryotes • The lac gene codes for a protein called β-galactosidase • To conserve energy, this enzyme is only made when lactose is present in the cell • When lactose is unavailable, the cell must block the production of β-galactosidase • What’s an operon? A region of bacterial DNA that codes for a series of functionally related genes that are transcribed into one RNA transcript • Let’s look at the anatomy of the lac gene to understand how this happens

12. P O The lac Operon • The gene that codes for β-galactosidase is part of an operon • An operon consists of structural genes that code for enzymes, promotor (P), and anoperator (O)

13. P O The lac Operon • Followed by the operon are a cluster of genes that code for proteins involved in the breakdown of lactose: lacZ, lacY, lacA

14. The lac Operon • lac Z • Z codes for the β-galactosidase enzyme that breaks down lactose lac Y -Y codes for a protein called galactoside permeasewhich transports lactose into the cell • lac A • A codes for transacetylase (function unknown )

15. There’s more to this operon… • lacI is a gene that is part of the operon but codes for a repressor protein; it has its own promoter • This repressor blocks the transcription of β-galactosidase by binding to the lactose operator (this prevents RNA polymerase from binding to the gene)  will any genes be transcribed?

16. P O

17. An Analogy… • Here's an analogy. A promoter is like a door knob, in that the promoters of many operons are similar. An operator is like the key hole in a door knob, in that each door is locked by only a specific key, which in this analogy is a specific regulatory protein (i.e. the repressor made by lacI).

18. So what does this all mean? • If lactose is not present, the repressor protein will be made to prevent the production of β-galactosidase and its associated proteins • What if lactose is present? β-galactosidase will be required! • What happens?

19. In the presence of lactose… • Lactose interacts with the repressor and causes it to change shape and release from its binding site (allosteric enzyme – what is this again?) • Thus, lactose itself is called the signal molecule or inducer

20. Allosteric Regulation http://library.thinkquest.org/28751/media/review/figure/allosteric.jpg

22. Think for a minute… • If scientists grew a culture of E.coli bacteria who are known to have a mutation in their lacI gene (so that the gene is defective), what would they observe in terms of lactose metabolism? • Loss of regulation, which causes constitutive production of β-galactosidase at all times despite the absence of lactose

23. Finally! It’s time to test your knowledge… Will you ace the test? Which one will it be?

24. Let’s see what you understood! 1. What are the stages of translation? – initiation, elongation and termination 2. What is it called when DNA is made into mRNA? mRNA to protein? – transcription and translation 3. Where does RNA Polymerase bind? – the promoter 4. What is an operon? – a region of bacterial DNA that codes for a series of functionally related genes that are transcribed into one RNA transcript

25. 1. If a certain gene contains 2 exons (each with 5 nucleotides) and 1 intron (containing 2 nucleotides), how many nucleotides will be in the mRNA transcript after splicing? a. 4 b. 8 c. 10 d. 12 2. From the following three nucleotides, what would be the mRNA codon and tRNA anticodon, respectively? 5’…….GCT…….3’ a. AGC, UCG b. UCG, AGC c. CGA, UCG d. TCG, TCG

26. 3. The following sequence depicts: DNA --X---> mRNA ------> protein ------> activated protein (NOTE: X = block in the pathway) a. Transcriptional control b. Post-transcriptional control c. Translational control d. Post-translational control 4. The following sequence depicts: DNA -- ---> mRNA --- X ---> protein ------> activated protein (NOTE: X = block in the pathway) a. Transcriptional control b. Post-transcriptional control c. Translational control d. Post-translational control

27. 5. The processes of transcription and translation occur in which parts of the eukaryotic cell, respectively? a. nucleus and cell membrane b. cytoplasm and extracellular space c. cytoplasm and nucleus d. nucleus and cytoplasm 6. Lactose cannot accumulate in an E. coli cell. What type of mutation has occurred? a. lacZ b. lacI c. lacY d. lacA

28. 7. Why is the lac operon called an operon? a. The genes are under the control of the same promoter. b. The genes are transcribed into the same molecule of mRNA. c. The genes are commonly regulated. d. All of the above. 8. The regulatory protein of the lac operon is: a. lacI b. lacZ c. lacY d. lacA 9. Lactose induces the transcription of: a. lacZ, lacY b. lacZ, lacI c. lacI, lacY d. lacX, lacA

29. How is the trp operon different? • Read p256-257 and take notes on the trp operon (also take notes of the similarities and differences between the lac and trp operons, Table 2 on p258) Homework: • Do questions #1-6 on p258