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BIOTECHNOLOGY UNIT:

BIOTECHNOLOGY UNIT:. “The Immortal Life of Henrietta Lacks” Dr. Wayne Grody- Guest Speaker “GATTACA” Ch18&19 Pro&Eukaryotic control of gene expression Ch11 Cell Signaling Cell Communication Ch20&21 Biotechnology…manipulating microorganisms to do our bidding!!!! Tools/techniques

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BIOTECHNOLOGY UNIT:

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  1. BIOTECHNOLOGY UNIT: • “The Immortal Life of Henrietta Lacks” • Dr. Wayne Grody- Guest Speaker • “GATTACA” • Ch18&19 Pro&Eukaryotic control of gene expression • Ch11 Cell Signaling Cell Communication • Ch20&21 Biotechnology…manipulating microorganisms to do our bidding!!!! Tools/techniques • LAB 6 MOLECULAR BIOLOGY • Bacterial transformation • Restriction Enzymes/DNA Digests • Gel Electrophoresis (DNA fingerprinting) • BIOTECH RESEARCH PAPER & SYMPOSIUM

  2. Control of Gene Expression Chapter 18

  3. VOCABULARY VOCABULARY QUESTIONS TO ANSWER

  4. Q & A • Q:What is gene expression? • A: Activating a gene to produce a protein. • Q: What is an operon? • A: The genes for creating an enzyme and the genes that regulate their transcription. • Q: What makes up an operon? • Answer: 1) Promoter region 2) Operator region 3) Structural genes

  5. OPERONPromoter, Operator, Structural Genes

  6. Control of gene expression • The expression of genetic material controls cell products, and these products determine the metabolism and nature of the cell. • Gene expression is regulated by both environmental signals and developmental cascades or stages. • Cell signaling mechanisms can also modulate and control gene expression. • Thus, structure and function in biology involve two interacting aspects: the presence of necessary genetic information and the correct and timely expression of this information.

  7. Gene expression can be under: • Negative regulation: When the operon is turned off by chemicals. repressible or inducible or • Positive regulation:When gene expression is stimulated by chemicals.

  8. 2 Types of Negative Regulation Part 1: The trp operon(repressible operon) • What does repressible mean? • This operon is located in E. coli bacteria. • The purpose of the operon is to create the enzymes that synthesize the amino acid tryptophan. • When the operon is “ON:

  9. RNA polymerase binds to the promoter region. • RNA polymerase crosses over the operator region because • the repressor is inactive and therefore does not bind to the operator. • The structural genes are transcribed and tryptophan is synthesized. OPERON “ON”

  10. Switching the trp operon “off” • Done by a repressor: a protein that binds to the operator region. • It blocks the attachment of RNA polymerase to the promoter region. • A gene called a regulatory gene (located away from the operon) produces the trp operon repressor. • The repressor protein is allosterichaving an active and inactive state/shape.

  11. 3) At first the repressor protein is in its inactive form. 4) When the amino acid tryptophan binds to the repressor the repressor becomes its active form and binds to the operator region… blocking transcription. 5) Since tryptophan assists in turning the operon off it is called a co-repressor. 6) When the levels of tryptophan drop the repressor loses its tryptophan, changes shape, and the repressor is released from the operator , initiating transcription again.

  12. TURNING THE OPERON OFF

  13. regulatory gene operator Regulatory gene codes for the repressor which may bind to the operator

  14. Types of Negative Regulation Part 2: The lac operon(inducible operon) • What does inducible mean? • This operon is also located in E. coli bacteria. • It was first discovered by: Francois Jacob and Jacques Monod (1961) called the “Jacob and Monod model” • The purpose of the operon is to produce the enzyme B-galactosidase that splits (via hydrolysis) lactose into glucose and galactose. • This operon is normally off because the repressor protein is formed in its active shape, thus binds to the operator region blocking RNA polymerase.

  15. If the RNA polymerase is being blocked how does transcription ever occur???

  16. Done by an inducer: a molecule that binds to and inactivates the repressor. • The molecule allolactose (an isomer of lactose) binds to the repressor, inducing an allosteric change. • The repressor is released from the operator region. • The RNA polymerase can move along the template strand catalyzing the synthesis of mRNA. • When the lactose levels decrease the repressor binds to the operator region and transcription is shut down.

  17. Figure 18.21a The lac operon: regulated synthesis of inducible enzymes

  18. Figure 18.21b The lac operon: regulated synthesis of inducible enzymes

  19. Compare and contrast: • The enzymes produced by the trp operon are called repressible enzymes and are involved in anabolic pathways. • The enzymes produced by the lac operon are called inducible enzymes and are involved in catabolic pathways.

  20. Types of Positive Regulation:A closer look at the lac operon • In order for the lac operon to produce enzymes in large quantities, a second factor must exist… • a low concentration of glucose.

  21. How does E. coli sense the low levels of glucose and how is this relayed to the lac operon? By the molecule cyclic AMP or cAMP. cAMP is present in large quantities when the glucose levels are low. • The cAMP binds to an allosteric protein called cAMP receptor protein or CRP • The activated CRP binds to a site within the lac promoter adjacent to the TATA box. • The attachment of CRP makes it easier for RNA polymerase to bind to the promoter region.

  22. Figure 18.22a Positive control: cAMP receptor protein

  23. Figure 18.22b Positive control: cAMP receptor protein CRP is known as an activator proteinbecause it activates transcription. If the levels of glucose increase the levels of cAMP decrease and the CRP is released from its binding site.

  24. Figure 18-22x cAMP

  25. Figure 18.20b The trp operon: regulated synthesis of repressible enzymes (Layer 2)

  26. Figure 18.21b The lac operon: regulated synthesis of inducible enzymes

  27. Figure 18.22a Positive control: cAMP receptor protein

  28. Figure 18.22b Positive control: cAMP receptor protein

  29. The Organization and Control of Eukaryotic Genomes Chapter 19

  30. How is eukaryotic gene expression different from prokaryotic gene expression? 1. Importance of cell specialization in multicelluar Euk’s. 2. Greater size of genome of Euk’s and chromatin structure: -single, circular, chromosome in PROKARYOTES -double, linear, protein enhanced in EUKARYOTES* Histones/Nucleosomes = DNA is coiled around bundles of 8 or 9 histone proteins to form DNA-histone complexes called nucleosomes. 1. Euchromatin = regions where DNA is loosely bound to nucleosomes and is actively transcribed. 2. Heterochromatin = regions where nucleosomes are more tightly compacted and DNA is inactive. (stains darker) * Compactly organized as chromosomes during cell division.

  31. Figure 19.1 Levels of chromatin packing

  32. DNA Packing is the first level of control of eukaryotic gene expression

  33. Figure 19.0 Chromatin in a developing salamander ovum

  34. Only _3_% of eukaryotic DNA is translated into protein products, compared to almost _100_% of prokaryotic DNA. Sizewise…several million nucleotide pairsvs.2 x 10 8 pairs per chromosome(that’s x 46 in humans!)

  35. Figure 19.x1a Chromatin HETEROCHROMATIN EUCHROMATIN

  36. Repetitive DNA = noncoding segments not transcribed within a gene • CENTROMERE- center • TELOMERE- ends (telomerase) • PSEUDOGENES = not transcribed, almost identical to a coding gene. May represent evolutionary precursor—mutated over the years. • TRANSPOSONS or “jumping genes”= can move to a new location on the same chromosome or to a different chromosome. Discovered by Barbara McKlintock (maise) Have the effect of a mutation… can change the expression of a gene • Turn on or off its expression • Have no effect at all

  37. Enduring understanding 3.B: Expression of genetic information involves cellular and molecular mechanisms. • Essential knowledge 3.B.1: • Gene regulation results in differential gene expression, leading to cell specialization.

  38. Enduring understanding 3.B: Expression of genetic information involves cellular and molecular mechanisms. • Essential knowledge 3.B.1: • Gene regulation results in differential gene expression, leading to cell specialization.

  39. 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. (ex. promoter, terminator, enhancer) • A regulatory gene is a sequence of DNA encoding a regulatory protein (repressor & activator) or RNA (miRNA & siRNA) blocks translation on a transcribed mRNA by binding to it. • Micro RNA • Small Interfering RNA

  40. RNA interference (RNAi) is a biological process in which RNA molecules inhibit gene expression, typically by causing the destruction of specific mRNA molecules. • Two types of small ribonucleic acid (RNA) molecules – microRNA (miRNA) and small interfering RNA (siRNA) – are central to RNA interference. RNAs are the direct products of genes, and these small RNAs can bind to other specific messenger RNA (mRNA) molecules and either increase or decrease their activity, for example by preventing an mRNA from producing a protein. RNA interference has an important role in defending cells against parasitic nucleotide sequences – viruses and transposons – but also in directing development as well as gene expression in general.

  41. In eukaryotes, gene expression is complex and control involves regulatory genes, regulatory elements and transcription factors that act in concert. • 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.

  42. Controlling Eukaryotic Gene Expression • Under positive control • Transcription will not take place without the assembly of the transcription complex • Transcription Factors are regulatory proteins that bond to the enhancer region, the promoter (TATA box) and to each other. • Once the transcription factors • have assembled around the • promoter, they are called a • transcription complex.

  43. Areas that regulate eukaryotic transcription: • The Enhancer Region: causes the chromosome to loop and make contact with the Promoter regions. • Located thousands of nucleotides away from the promoter. • Activator proteins bind to the enhancer regions and then to the transcription complex after the DNA loops. • When the activator proteins (special transcription factors) bind to the transcription complex, RNA polymerase is positioned over the promoter region and the rate of transcription increases.

  44. The Silencer Region: a repressor region. • Located close to the enhancer region. • Repressor Proteins bind to the silencer sites prevent the activator proteins from binding to the enhancer region.

  45. Turning On A Gene

  46. Turning On A Eukaryotic Gene

  47. Figure 19.8 A eukaryotic gene and its transcript

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