Regulation of Gene Expression

1. Regulation of Gene Expression Chapter 18

2. Figure 18.2

3. Overview of Gene Expression Regulation of Gene Expression • Both DNA regulatory sequences, regulatory genes, and small regulatory RNAs are involved in gene expression. • Regulatory sequences are stretches of DNA that interact with regulatory proteins to control transcription (promoters, terminators, enhancers). • A regulatory gene is a sequence of DNA encoding a regulatory protein or RNA. • Regulatory proteins are proteins that regulate or are involved in gene expression. • Gene regulation accounts for some of the phenotypic differences between organisms with similar genes.

4. In Viruses & Bacteria… Regulation of Gene Expression • Both positive and negative control mechanisms regulate gene expression in viruses & bacteria. • The expression of certain genes can be turned ON by the presence of an inducer. • The expression of certain genes can be turned OFF by the presence of a repressor. • Inducers and repressors are small molecules that interact with regulatory proteins and/or regulatory sequences. • Regulatory proteins INHIBIT gene expression by binding to DNA and blocking transcription (negative control). • Regulatory proteins STIMULATE gene expression by binding to DNA and stimulating transcription (positive control) or binding to repressors to inactivate them. • Certain genes are continuously expressed; always turned on.

5. In Eukaryotes… Regulation of Gene Expression • Gene expression is complex and involves regulatory genes, regulatory elements, and transcription factors that act in tandem. • Transcription factors bind to specific DNA sequences and/or other regulatory proteins. • Some of these transcription factors are activators (increase expression), while others are repressors (decrease expression). • The combination of transcription factors binding to the regulatory regions at any one time determines how much, if any, of the gene product will be produced.

6. Prokaryotic Gene Regulation Control of Gene Expression in Bacteria • Bacteria often respond to environmental change by regulating transcription. • In bacteria, genes are often clustered into operons, with one promoter serving several adjacent genes. • An operator site on the DNA switches the operon on or off, resulting in coordinate regulation of the genes.

7. Prokaryotic Gene Regulation Operons: The Basic Concept • An operon is essentially a set of genes and the switches that control the expression of those genes. • An operon consists of: • operator • promotor • and genes that they control • All together, the operator, the promoter, and the genes they control – the entire stretch of DNA required for enzyme production for the pathway – is called an operon.

8. Prokaryotic Gene Regulation The Operon Model

9. Prokaryotic Gene Regulation Repressible & Inducible Operons • There are basically two types of operons found in prokaryotes: repressible operons and inducible operons. • Both the repressible and inducible operon are types of NEGATIVE gene regulation because both are turned OFF by the active formof the repressor protein. • In either type of operon, binding of a specific repressor protein to the operator shuts off transcription. • Trp operon – repressible operon is always in the on position until it is not needed and becomes repressed or switched off. • Lac operon – inducible operon is always off until it is induced to turn on.

10. Figure 18.3a – The trp Operon http://bcs.whfreeman.com/thelifewire/content/chp13/1302002.html

11. Figure 18.3b – The trp Operon http://highered.mcgraw-hill.com/olc/dl/120080/bio26.swf

12. Figure 18.4a – The lac Operon http://www.sumanasinc.com/webcontent/animations/content/lacoperon.html

13. Figure 18.4b – The lac Operon http://highered.mcgraw-hill.com/sites/dl/free/0072835125/126997/animation27.html

14. Prokaryotic Gene Regulation Positive Gene Regulation • When glucose and lactose are both present in its environment, E. coli prefer to use glucose. • Only when lactose is present AND glucose is in short supply does E. coli use lactose as an energy source, and only then does it synthesize appreciable quantities of the enzymes for lactose breakdown. • How does the E. coli cell sense the glucose concentration and relay this information to its genome? • http://highered.mcgraw-hill.com/olc/dl/120080/bio27.swf

15. Figure 18.5a – Positive Control

16. Figure 18.5b – Positive Control

17. Factors Affecting Ability of Repressor to Bind to Operator Co-Repressor : Activates a Repressor Seen in the trp Operon Co-Repressor is tryptophan Turns normally “on” Operon “off” Inducer: Inactivates a Repressor, Induces the Gene to be Transcribed Seen in the lac Operon Inducer is allolactose Turns normally “off” Operon “on” Prokaryotic Gene Regulation

18. Prokaryotic Gene Regulation Structure/Function of Prokaryotic Chromosomes • shape (circular/nonlinear/loop) • less complex than eukaryotes (no histones/less elaborate structure/folding) • size (smaller size/less genetic information/fewer genes) • replication method (single origin of replication/rolling circle replication) • transcription/translation may be coupled • generally few or no introns (noncoding segments) • majority of genome expressed • operons are used for gene regulation and control • NOTE: plasmids – more common but not unique to prokaryotes/not part of prokaryote chromosome

19. Overview Figure 18.6 • THIS FIGURE IS HIGHLIGHTING KEY STAGES IN THE EXPRESSION OF A PROTEIN-CODING GENE. • The expression of a given gene will not necessarily involve every stage shown. • MAIN LESSON: each stage is a potential control point where gene expression can be turned on or off, sped up, or slowed down.

20. Eukaryotic Chromosomes Chromosome Structure of Eukaryotes Eukaryotic chromosomes contain DNA wrapped around proteins called histones. The strands of nucleosomes are tightly coiled and supercoiled to form chromosomes.

21. Eukaryotic Gene Regulation The Eukaryotic Genome The difference between cell types in eukaryotes are NOT due to different genes being present, but to differential geneexpression. This is the expression of DIFFERENT genes by cells with the SAME genome.

22. Eukaryotic Chromosome Structure Eukaryotic Gene Regulation Chromatin structure is based on successive levels of DNA packing. Eukaryotic chromatin is composed mostly of DNA and histone proteins that bind to the DNA to form nucleosomes, the most basic units of DNA packing. Additional folding leads ultimately to highly compacted heterochromatin, the form of chromatin in a metaphase chromosome. In interphase cells, most chromatin is in a highly extended form, called euchromatin.

23. Eukaryotic Gene Regulation Chromatin Modifications • The relationship between DNA and its histones is governed by two chemical interactions: • DNA methylation: (-CH3) the addition of methyl groups to DNA – causes DNA to be more TIGHTLY packaged, thus REDUCING gene expression. • Histone acetylation: (-COCH3) the addition of acetyl groups to amino acids of histone proteins – makes chromatin LESS TIGHTLY packed and STIMULATES transcription. • Methylation occurs primarily on DNA and reduces gene expression. • Acetylation occurs on histones and increases gene expression.

24. Figure 18.8 Eukaryotic Gene and its Transcript

25. Assembling of Transcription Factors • Activator proteins bind to enhancer sequences in the DNA and help position the initiation complex on the promoter. • DNA bending brings the bound activators closer to the promoter. Other transcription factors and RNA polymerase are nearby. • Protein-binding domains on the activators attach to certain transcription factors and help them form an active transcription initiation complex on the promoter. • http://highered.mcgraw-hill.com/olc/dl/120080/bio28.swf Control elements are simply segments of noncoding DNA that help regulate transcription of a gene by binding proteins (transcription factors).

26. Alternative Splicing Offers New Combinations of Exons = New Proteins Eukaryotic Gene Regulation The RNA transcripts of some genes can be spliced in more than one way, generating different mRNA molecules. With alternative splicing, an organism can get more than one type of polypeptide from a single gene.

27. Eukaryotic Gene Regulation Further Control of Gene Expression • After RNA processing, other stages of gene expression that the cell may regulate are: • mRNA degradation; • translation initiation, • protein processing & degradation

28. Eukaryotic Gene Regulation mRNA Degradation • The life span of mRNA molecules in the cytoplasm is important in determining the pattern of protein synthesis in a cell. • In bacteria, mRNA are typically degraded within a few minutes of their synthesis – enables bacteria to change quickly in response to environmental changes. • In eukaryotes, mRNA typically survive for days or weeks. Breakdown begins by shortening the poly-A tail and removing the 5’ cap.

29. Eukaryotic Gene Regulation Translation Initiation • Translation presents another opportunity for regulating gene expression in eukaryotes – particularly at the translation initiation stage. • This type of regulation typically occurs at the 5’ cap or poly-A tail. • If regulatory protein binds to 5’ region, ribosome cannot attach to mRNA – thus no translation occurs. • If mRNA lacks a poly-A tail of sufficient length, translation initiation will not occur because poly-A tail facilitates attachment of rRNA to mRNA during translation.

30. Eukaryotic Gene Regulation Protein Processing & Degradation • Often, eukaryotic polypeptides must be processed to yield functional proteins, and regulation can occur at any stage of protein processing: • Proper folding is required • Chemical modification is required • Protein must be transported to proper location within or outside the cell • Selective degradation regulates the length of time a particular protein functions in a cell. • Proteins to be degraded are tagged with ubiquitin, and proteasomes recognize these and chop them apart.

31. Eukaryotic Gene Regulation Noncoding RNAs & Gene Expression • A significant amount of the eukaryotic genome may be transcribed into small non-protein-coding RNAs. • These play crucial roles in regulating gene expression – generally during mRNA translation and chromatin configuration. • MicroRNAs (miRNAs): bind to mRNA sequences and degrade the mRNA before translation or block its translation. • Small Interfering RNAs (siRNAs): can be crucial for the formation of heterochromatin at the centromeres of chromosomes.

32. Structure & Function of Eukaryotic Chromosome • Chromatids • 2/sister/pari/identical DNA/ genetic information • distribution of one copy to each new cell • Centromere • noncoding/uncoiled/narrow/constricted region • joins/holds/attaches chromatids together • Nucelosome • histones/DNA wrapped arround special proteins • packaging compacting • Chromatin Form (heterochromatin/euchromatin) • heterochromatin is condensed/supercoiled • proper distribution in cell division (not during replication) • euchromatin is loosely coiled • gene expression during interphase/replication occurs when loosely packed • Kinetochores • disc-shaped proteins • spindle attachment/alignment • Genes or DNA • brief DNA description • codes for proteins or for RNA • Telomeres • tips, ends, noncoding repetitive sequences • protection against degradation/ aging, limits number of cell divisions

34. Cell Differentiation Differential Gene Expression & Cell Differentiation • A program of differential gene expression leads to the different cell types in a multicellular organism. • A zygote typically undergoes transformation in three interrelated processes: • Cell Division: the series of mitotic divisions that increases the number of cells. • Cell Differentiation: the process by which cells become specialized in structure & function. • Morphogenesis: the organization of cells into tissues and organs.

35. Control of Cell Differentiation & Morphogenesis Cell Differentiation Cytoplasmic determinants are maternal substances in the egg that influence the course of early development – they are distributed unevenly in the early cells of the embryo and this has a profound impact on early development. Cell-cell signals result from molecules, such as growth factors, produced by one cell influencing neighboring cells in a process called induction, which causes cells to differentiate. Determination is the series of events that lead to observable differentiation of a cell – caused by cell-cell signals and is irreversible. Pattern formation sets up the body plan and is a result of cytoplasmic determinants and inductive signals – determines head/tail, left/right, and back/front.

36. Early Development & Homeotic Genes Cell Differentiation • Homeotic genes are any of the master regulatory genes that control placement and spatial organization of body parts in eukaryotes by controlling the developmental fate of a group of cells. • Mutations in certain regulatory/homeotic genes can cause a misplacement of structures in a eukaryotic organism. • As it relates to homeotic genes: • You should be familiar with the “Scientific Inquiry” on page 370 (read text) and Figure 18.17 & 18.18 in text. • You should also be familiar with Inquiry/Figure 18.19 in text.

37. The Molecular Biology of Cancerhttp://science.education.nih.gov/supplements/nih1/cancer/activities/activity2_animations.htm The Biology of Cancer • Certain genes normally regulate growth and division – the cell cycle – and mutations that alter those genes in somatic cells can lead to cancer. • Proto-Oncogenes are normal genes that code for proteins which stimulate normal cell growth and division. • Oncogenes – cancer causing genes; lead to abnormal stimulation of cell cycle. • Oncogenes arise from genetic changes in proto-oncogenes: • Amplification of proto-oncogenes • Point mutation in proto-oncogene • Movement of DNA within genome

38. Genetic Changes Can Turn Proto-oncogenes into Oncogeneshttp://www.learner.org/courses/biology/units/cancer/images.html The Biology of Cancer

39. The Biology of Cancer Tumor-Suppressor Genes In addition to genes whose products normally promote cell division, cells contain genes whose normal products inhibit cell division. These genes are referred to as tumor-suppressor genes because the proteins they encode help prevent uncontrolled cell growth. Cancer can be caused by a mutation in a tumor-suppressor gene if the mutation causes the gene to fail to prevent uncontrolled division.

40. The Biology of Cancer p53: Guardian Angel of the Genome • The p53 gene is an important tumor-suppressor gene. This gene may suppress cancer in three ways: • The p53 gene halts the cell cycle by binding to cylcin-dependent kinases – allows time for DNA to be repaired before the resumption of cell division. • The p53 genes turns on genes directly involved in DNA repair. • When DNA damage is too severe to repair, the p53 gene activates suicide genes whose products cause apoptosis (cell death). • In many cancer patients, the p53 gene product does not function properly.

41. The Biology of Cancer ras Proto-oncogene • The Ras protein, encoded by the ras proto-oncogene, is a G protein that relays a signal from a growth factor receptor on the plasma membrane to a cascade of protein kinases. • The cellular response at the end of the pathway is the synthesis of a protein that stimulates the cell cycle. • Normally – this pathway will not operate unless triggered by the appropriate growth factor. • Certain mutations in the ras gene (conversion to an oncogene) can lead to production of a hyperactive Ras protein. • This hyperactive Ras protein triggers the transduction pathway in the absence of growth factors. • This hyperactivity of the Ras protein leads to excessive cell division (cancer).

42. Figure 18.21 Signaling pathways that regulate cell growth (Layer 2) RAS and p53 contribute to uninhibited cell stimulation and growth- Tumor Formation

43. Figure 18.22 A multi-step model for the development of colorectal cancer The Biology of Cancer

44. The Biology of Cancer Genetic Predispositions to Cancer • The fact that multiple genetic changes are required to produce a cancer cell helps explain the observation that cancers can run in families. • About 15% of colorectal cancers involve inherited mutations – many affecting the APC tumor-supressor gene. The APC gene is mutated in 60% of colorectal cancer patients. • Mutations in the BRCA1 or BRCA2 gene are found in at least half of all inherited breast cancers. • A woman who inherits one mutant BRCA1 allele has a 60% probability of developing breast cancer before the age of 50 (compared to 2% for an individual without the mutant allele). • Both the BRCA1 and BRCA2 are tumor-supressor genes, because the wild-type allele protects agains breast cancer.

45. The Biology of Cancer Viruses & Cancer • Viruses can contribute to cancer development in several ways if they integrate their genetic material into the DNA of infected cells. • Viral integration may donate an oncogene to the cell, disrupt a tumor-suppressor gene, or convert a proto-oncogene to an oncogene. • Some viruses produce proteins that inactivate p53 and other tumor-suppressor genes, thus making the cell more prone to becoming cancerous.

46. USEFUL ANIMATIONS http://highered.mcgraw-hill.com/olc/dl/120080/bio31.swf http://highered.mcgraw-hill.com/olc/dl/120077/bio25.swf http://highered.mcgraw-hill.com/olc/dl/120080/bio28.swf http://highered.mcgraw-hill.com/olc/dl/120082/bio34b.swf http://www.learner.org/courses/biology/units/cancer/images.html

47. NEED TO KNOW You should now be able to: • Explain the concept of an operon and the function of the operator, repressor, and corepressor • Explain the adaptive advantage of grouping bacterial genes into an operon • Explain how repressible and inducible operons differ and how those differences reflect differences in the pathways they control

48. NEED TO KNOW • Explain how DNA methylation and histone acetylation affect chromatin structure and the regulation of transcription • Define control elements and explain how they influence transcription • Explain the role of promoters, enhancers, activators, and repressors in transcription control

49. NEED TO KNOW • Explain how eukaryotic genes can be coordinately expressed • Describe the roles played by small RNAs on gene expression • Explain why determination precedes differentiation • Describe two sources of information that instruct a cell to express genes at the appropriate time

50. NEED TO KNOW • Explain how mutations in tumor-suppressor genes can contribute to cancer • Describe the effects of mutations to the p53 and ras genes