Regulation of Gene Expression

1. Regulation of Gene Expression Chapter 18

2. Regulation of Gene Expression Overview of Gene Expression • The control of gene expression is vital to the proper and efficient functioning of an organism. • Cells control metabolism by either regulating enzyme activity –or- regulating the expression of genes coding for enzymes.

3. Figure 18.2

4. 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.

5. 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.

6. Prokaryotic Gene Regulation The Operon Model

7. 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 form of 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.

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

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

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

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

12. 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

13. Figure 18.5a – Positive Control

14. Figure 18.5b – Positive Control

15. 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

16. Prokaryotic Gene Regulation Review: 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

17. Chromosome Structure The Structure of the Chromosome • In Prokaryotes: • The bacterial chromosome is a double-stranded, circular DNA molecule associated with a small amount of protein • In a bacterium, the DNA is “supercoiled” and found in a region of the cell called the nucleoid • In Eukaryotes: • Eukaryotic chromosomes have linear DNA molecules associated with a large amount of protein • Chromatin is a complex of DNA and protein, and is found in the nucleus of eukaryotic cells • Histones are proteins that are responsible for the first level of DNA packing in chromatin

18. 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.

19. Eukaryotic Chromosomes

20. Eukaryotic Gene Regulation Control of Gene Expression in Eukaryotes • Eukaryotic gene expression can be regulated at any stage. • Because gene expression in eukaryotes involves more steps, there are more places where gene control can occur. • Opportunities for the control of gene expression in eukaryotes include: • Chromatin Packing, modification • Assembling of Transcription Factors • RNA Processing • Regulation of mRNA degradation and Control of Translation • Protein Processing and Degradation

21. 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.

22. Eukaryotic Gene Regulation Expression of Genes in Eukaryotes • Eukaryotic cells face the same challenges as prokaryotic cells in expressing their genes, but with two main differences: • The much greater size of the typical eukaryotic genome; • importance of cell specialization in multicellular eukaryotes. • In both prokaryotes and eukaryotes, DNA associates with proteins to form chromatin, but in the eukaryotic cell, the chromatin is ordered into higher structural levels.

23. 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.

24. Eukaryotic Gene Regulation The Eukaryotic Genome • In prokaryotes, most of the DNA in a genome codes for protein, with a small amount of noncoding DNA that consists mainly of regulatory sequences such as promoters. • In eukaryotic genomes, most of the DNA (97% in humans) does NOT encode protein or RNA. • This DNA includes introns and repetitive DNA: • Repetitive DNA are nucleotide sequences that are present in many copies in a genome, usually not within genes.

25. Eukaryotic Gene Regulation Chromatin Modifications • Chromatin modifications affect the availability of genes for transcription: • The physical state of DNA in or near a gene is important in helping control whether the gene is available for transcription. • Genes of heterochromatin (highly condensed) are usually not expressed because transcription proteins cannot reach the DNA. • DNA methylation seems to diminish transcription of that DNA. • Histone acetylation seems to loosen nucleosome structure and thereby enhance transcription.

26. Eukaryotic Gene Regulation DNA Methylation • DNA methylation is the attachment of methyl groups (-CH3) to DNA bases after DNA is synthesized. • Methylation renders DNA inactive. • Inactive DNA, such as that of inactivated mammalian X chromosomes (Barr bodies), is generally highly methylated compared to DNA that is actively transcribed. • Comparison of the same genes in different types of tissues shows that the genes are usually more heavily methylated in cells where they are not expressed. • In addition, de-methylating certain inactive genes (removing their extra methyl groups) turns them on. • At least in some species, DNA methylation seems to be essential for the long-term inactivation of genes that occurs during cellular differentiation in the embryo.

27. Eukaryotic Gene Regulation Histone Acetylation • Histone acetylation is the attachment of acetyl groups (-COOH3) to certain amino acids of histone proteins; de-acetylation is the removal of acetyl groups. • When the histones of nucleosome are acetylated, they change shape so that they grip the DNA less tightly. • As a result, transcription proteins have easier access to genes in the acetylated region.

28. Eukaryotic Gene Regulation Transcription Initiation • Transcription is controlled by the presence or absence of particular transcription factors, which bind to the DNA and affect the rate of transcription. • Thus…transcription initiation is controlled by proteins that interact with DNA and with each other. • Once a gene is “unpacked”, the initiation of transcription is the most important and universally used control point in gene expression.

29. Figure 18.8 Eukaryotic Gene and its Transcript

30. 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).

31. Eukaryotic Gene Regulation Post-Transcriptional Factors • Transcription alone DOES NOT constitute gene expression! • Post-transcriptional mechanisms play supporting roles in the control of gene expression: • Alternative RNA splicing – where different mRNA molecules are produced from the same primary transcript, depending on which RNA segments are treated as exons and which as introns. • Regulatory proteins specific to a cell type control intron-exon choices by binding to regulatory sequences within the primary transcript. • http://highered.mcgraw-hill.com/olc/dl/120080/bio31.swf

32. 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.

33. 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, and protein processing and degradation. • The life span of mRNA molecules in the cytoplasm is an important factor in determining the pattern of protein synthesis in a cell. • Most translational control mechanisms block the initiation stage of polypeptide synthesis, when ribosomal subunits and the initiator tRNA attach to an mRNA.

34. Eukaryotic Gene Regulation Protein Processing and Degradation • The final opportunities for controlling gene expression occur after translation: • Protein processing – cleavage and the addition of chemical groups required for function. • Transport of the polypeptide to targeted destinations in the cell. • Cells can also limit the lifetimes of normal proteins by selective degradation – chopped up by proteasomes.

35. 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.

36. The Biology of Cancer The Molecular 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

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

38. The Biology of Cancer Tumor-Suppressor Genes • In addition to mutations affecting growth-stimulating proteins, changes in genes whose normal products INHIBIT cell division also contribute to cancer: • Such genes are called tumor-suppressor genes because the proteins they encode normally help prevent uncontrolled cell growth.

39. The Biology of Cancer p53 Tumor Suppressor and ras Proto-Oncogeneshttp://www.learner.org/courses/biology/units/cancer/images.html • Mutations in the p53 tumor-suppressor gene and the ras proto-oncogene are very common in human cancers. • Both are components of signal-transduction pathways that convey external signal to the DNA in the cell’s nucleus. • Product of ras gene is G Protein (relays a growth signal and stimulates cell cycle). • An oncogene protein that is a hyperactive version of this protein in the pathway can increase cell division. • P53 protein – “guardian angel of the genome” • DNA damage (UV, toxins) signals expression of p53 and p53 protein acts as transcription factor for gene p21 • p21 halts cell cycle, allowing DNA repair • P53 also can cause ‘cell suicide’ if damage is too great • Many cancer patients p53 gene product does not function properly!

40. Figure 18.21 Signaling pathways that regulate cell growth (Layer 2) RAS and P53 contribute to uninhibited cell stimulation and growth- Tumor Formation

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

42. Review: Structure/Function of Eukaryotic Chromosomes • 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

43. 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

44. 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

45. 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

46. 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

47. 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