AP Biology Day 30-31 Wednesday, November 2, 2016 Friday, November 3, 2016
Do-Now 1. With your neighbors, plan out your response and jot down your ideas for the 2016 short FRQ 2
Part (a) The primary transcript in the figure is 15 kb long, but the mature mrna is 7 kb in length. Describe the modification that most likely resulted in the 8 kb difference in length of the mature mrna molecule. Identify in your response the location in the cell where the change occurs Describe (1 pt) à Discuss the removal of introns during RNA splicing/processing Identify (1 pt) à This occurs in the nucleus (transcription)
Part (b) Predict the length of the mature gene X mrna if the fulllength gene is introduced and expressed in prokaryotic cells. Justify your prediction. Predict (1 pt) à 15 kb Justify (1 pt) à mrna splicing/processing does not occur in prokaryotes
Announcements AEend tutoring Biotech labs begin next week!!! Avoid absences! No collab Monday à Even Period Day! Progress Report grades submieed next Wed AM Unit logs must be submieed
CW/HW Assignments 10. Ch. 18 Notes Outline PLANNER 1. Outline Ch. 18 Notes (use ppt or GR) 2. Study Ch. 17-18 à Quiz Monday 3. Even period schedule Monday labs next week! 4. Clear missing work/finish retakes
EssenOal knowledge standards 3.B.1: Gene regulation results in differential gene expression, leading to cell specialization 3.B.2: A variety of intercellular and intracellular signal transmissions mediate gene expression 4.A.3: Interactions between external stimuli and regulated gene expression result in specialization of cells, tissues, and organs
I will be able to: FLT explain the concept of an operon and the funcoon of the operator, repressor, and corepressor Explain how DNA methylaoon and histone acetylaoon affect chromaon structure and the regulaoon of transcripoon Explain the role of promoters, enhancers, acovators, and repressors in transcripoon control By comple1ng Ch. 18 Lecture Notes
Ch. 18: Regula.on of Gene Expression
Recall: Gene Expression 10
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Gene Expression Does every gene need to always be expressed? Human cells contain ~20,000-25,000 genes! Some will always be expressed Some can be turned on or off This is gene regulaoon J 12
Overview Prokaryotes and eukaryotes alter gene expression in response to their changing environment In mulocellular eukaryotes, gene expression regulates development and is responsible for differences in cell types RNA molecules play many roles in regulaong gene expression in eukaryotes 13
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I. Gene Expression in Prokaryotes A. Cells have two main ways of controlling metabolism B. Operon C. NegaOve Gene RegulaOon D. PosiOve Gene RegulaOon 15
A. Cells have two main ways of controlling metabolism 1. Regulate enzyme acovity 2. Regulate gene expression 16
Precursor Feedback inhibi.on trpe gene Enzyme 1 Enzyme 2 trpd gene trpc gene Regula.on of gene expression Enzyme 3 trpb gene trpa gene Tryptophan (a) Regula.on of enzyme ac.vity (b) Regula.on of enzyme produc.on
Does every gene need to always be expressed? Human cells contain ~20,000-25,000 genes! Some will always be expressed Some can be turned on or off This is gene regulaoon J 18
1. Concept B. Operon A cluster of func.onally related genes can be under coordinated control by a single on-off switch 19
2. Operator B. Operon A segment of DNA that acts as the regulatory switch. Ac.vators and repressors can bind to it. 20
3. Promoter B. Operon Segment of DNA where RNA polymerase binds for transcrip.on to begin (ex/ TATA) 21
4. Operon B. Operon The en.re stretch of DNA that includes the operator, promoter, and the genes that they control 22
5. Repressor B. Operon = a protein that can switch off the operon (blocks transcrip.on) The repressor prevents gene transcrip.on as it binds to the operator and blocks RNA polymerase The repressor is the product of a separate regulatory gene The repressor can be in an acove or inacove form, depending on the presence of other molecules 23
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B. Operon 6. Inducer = molecules that help control the operator gene 7. Regulatory gene = regulate the ac.vity of other genes by coding for repressors or ac.vators 27
B. Operon 8. Corepressor = A molecule that cooperates with a repressor protein to switch an operon off 28
Pair-Share-Respond 1. How are genes expressed? 2. What is an operon? 3. What is a promoter? 4. What is an operator and what can bind to it? 5. How does a repressor work?
Gene RegulaOon RegulaOon of genes can be negaove or posiove NegaOve control: Genes are expressed unol they are switched off by a repressor 30
Gene RegulaOon PosiOve control: Genes are expressed only when an acovator is present 31
C. NegaOve Gene RegulaOon E. coli can synthesize the amino acid tryptophan (one of the essenoal amino acids) 32
C. NegaOve Gene RegulaOon Synthesis of tryptophan can be regulated through the trp operon 33
C. NegaOve Gene RegulaOon 1. Repressible enzymes Repressible pathways à transcrip.on of the gene(s) con.nues un.l the operon is switched off by high levels of the end product (corepressor) 34
C. NegaOve Gene RegulaOon 1. Repressible enzymes Example: trp operon Normally, the trp operon is on and the genes for tryptophan synthesis are transcribed 35
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C. NegaOve Gene RegulaOon 1. Repressible enzymes When tryptophan levels are high, it binds to the trp repressor protein, which turns the operon off (stops transcrip.on) Repressible operon is usually on, but the binding of a repressor can shut off transcripoon 37
trp Operon - Repressible DNA Regulatory gene mrna Promoter Protein 5ʹ trpr trpe trpd trpc trpb Operator Start codon Stop codon 3ʹ RNA mrna 5ʹ polymerase E D C B Inac.ve repressor Promoter trp operon Genes of operon Polypep.de subunits that make up enzymes for tryptophan synthesis (a) Tryptophan absent, repressor inac4ve, operon on. trpa A DNA No RNA made mrna Protein Tryptophan (corepressor) Ac.ve repressor (b) Tryptophan present, repressor ac4ve, operon off.
Pair-Share-Respond 1. What is the difference between nega.ve and posi.ve control of gene expression? 2. Explain how the trp operon func.ons
C. NegaOve Gene RegulaOon E. Coli can also metabolize lactose! This involves several proteins 40
C. NegaOve Gene RegulaOon 2. Inducible enzymes Inducible operon = one that is usually off; a molecule called an inducer inac.vates the repressor and turns on transcrip.on Ex: lac operon 41
C. NegaOve Gene RegulaOon 2. Inducible enzymes It s wasteful to produce enzymes to metabolize lactose when it s not present If no lactose is present, the repressor on the operator prevents trancsripoon 42
C. NegaOve Gene RegulaOon 2. Inducible enzymes In the presence of lactose, an inducer binds to the repressor to inacovate it RNA polymerase can then transcribe the genes for metabolizing lactose 43
Regulatory gene Promoter Operator lac Operon - inducible DNA laci lacz mrna 5ʹ 3ʹ RNA polymerase No RNA made Protein Ac.ve repressor (a) Lactose absent, repressor ac.ve, operon off. lac operon DNA laci lacz lacy laca mrna 5ʹ 3ʹ RNA polymerase mrna 5ʹ Protein β-galactosidase Permease Transacetylase Allolactose inducer Inac.ve repressor (b) Lactose present, repressor inac.ve, operon on.
Lactose present 45
Enzymes made & repressor is released à lac operon also produces enzymes that break down lactose and allolactose, and repressor can bidn to operator to stop addioonal transcripoon 46
C. NegaOve Gene RegulaOon 2. Inducible enzymes Note: this is not the complete explanaoon for the lac operon 47
C. NegaOve Gene RegulaOon 2. Inducible enzymes Inducible enzymes usually funcoon in catabolic pathways; their synthesis is induced by a chemical signal Repressible enzymes usually funcoon in anabolic pathways; their synthesis is repressed by high levels of the end product Regula.on of the trp and lac operons involves nega.ve control of genes because operons are switched off by the ac.ve form of the repressor 48
Gene RegulaOon PosiOve control: Genes are expressed only when an acovator is present 49
D. PosiOve Gene RegulaOon The lac operon is controlled by mul.ple switches 1. Catabolite AcOvator Protein (CAP) CAP is an acovator of transcripoon 50
Whenever glucose is present, E. Coli will metabolize it before using alteraove sources of energy (such as lactose) If both glucose and lactose are available, the genes for lactose metabolism will only be transcribed at low levels When glucose starts to run low, then the lac genes will be transcribed at a greater rate 51
E. Coli: Glucose > Lactose 52
The presence or absence of glucose affects the lac operon by affecong the concentraoon of cyclic AMP camp is derived from ATP 53
camp can help facilitate the transcrip.on of the lac operon églucose, then êcamp êglucose, then écamp 54
In the presence of lactose and absence of glucose, camp joins with a CAP that binds to the lac promoter and facilitates the transcripoon of the lac operon églucose, then êcamp êglucose, then écamp 55
D. PosiOve Gene RegulaOon 1. Catabolite AcOvator Protein (CAP) CAP is an acovator of transcripoon a. Lactose present/glucose scarce In the presence of lactose and absence of glucose, camp joins with a CAP (ac.va.ng it) that binds to the lac promoter and facilitates the transcrip.on of the lac operon AcOvated CAP aeaches to the promoter of the lac operon and increases the affinity of RNA polymerase, acceleraong transcripoon Maximal transcripoon of the lac operon occurs only when glucose is absent and lactose is present. 56
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D. PosiOve Gene RegulaOon 1. Catabolite AcOvator Protein (CAP) CAP is an acovator of transcripoon b. Lactose present/glucose present When glucose levels increase, CAP detaches from the lac operon, and transcrip.on returns to a normal rate Genes for lactose metabolism transcribed to a small extent (low levels) 58
Posi.ve Gene Regula.on Promoter Operator DNA laci CAP-binding site camp Ac.ve CAP lacz RNA polymerase binds and transcribes Inac.ve CAP Allolactose Inac.ve lac repressor (a) Lactose present, glucose scarce (camp level high): abundant lac mrna synthesized Promoter Operator DNA CAP-binding site Inac.ve CAP laci lacz RNA polymerase less likely to bind Inac.ve lac repressor (b) Lactose present, glucose present (camp level low): liale lac mrna synthesized
Pair-Share-Respond 1. Explain the lac operon in terms of nega.ve control 2. The CAP can help accelerate transcrip.on of the lac operon. When will it do this? 3. What ac.vates the CAP?
III. Control of Gene Expression in Eukaryotes A. DifferenOal gene expression B. Stages in gene expression that can be regulated 61
A. DifferenOal gene expression Almost all the cells in an organism are geneocally idenocal Differen.al gene expression = when cells express different genes (making them different cell types) Remember: this is from the same DNA 62
A. DifferenOal gene expression Errors in gene expression can lead to diseases including cancer Gene expression is regulated at many stages 63
Signal Eukaryo.c Differen.al Gene Expression NUCLEUS Chroma.n Chroma.n modifica.on DNA Gene Gene available for transcrip.on Transcrip.on RNA Exon Intron Primary transcript RNA processing Tail Cap mrna in nucleus Transport to cytoplasm mrna in cytoplasm CYTOPLASM Degrada.on of mrna Transla.on Polypep.de Protein processing Degrada.on of protein Ac.ve protein Transport to cellular des.na.on Cellular func.on
B. Stages in gene expression that can be regulated 1. RegulaOon of chromaon structure 2. TranscripOonal regulaoon 3. Post-TranscripOonal RegulaOon 4. Noncoding RNAs 65
1. RegulaOon of ChromaOn Structure Chemical modificaoons to histones and DNA of chromaon influence both chromaon structure and gene expression 66
1. RegulaOon of ChromaOn Structure a. Histone acetyla.on Acetyl groups are aaached to posi.vely charged lysines in histone tails This loosens chroma.n structure, thereby promo.ng the ini.a.on of transcrip.on 67
Histone Modifica.ons Acetyla.on Promotes Transcrip.on DNA double helix Histone tails Amino acids available for chemical modifica.on (a) Histone tails protrude outward from a nucleosome. Unacetylated histones Acetylated histones (b) Acetylation of histone tails promotes loose chromatin structure that permits transcription.
1. RegulaOon of ChromaOn Structure b. DNA methyla.on Addi.on of methyl groups Methyla.on tends to condense chroma.n thereby restric.ng transcrip.on DNA methylaoon can cause long-term inacovaoon of genes in cellular differenoaoon In genomic imprinong, methylaoon regulates expression of either the maternal or paternal alleles of certain genes at the start of development 69
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1. RegulaOon of ChromaOn Structure Although the chromaon modificaoons just discussed do not alter DNA sequence, they may be passed to future generaoons of cells EpigeneOc inheritance = Process by which heritable modificaoons in gene funcoon occur but are not due to changes in the base sequence of the DNA of the organism 73
2. TranscripOonal regulaoon ChromaOn-modifying enzymes provide inioal control of gene expression by making a region of DNA either more or less able to bind the transcripoon machinery Associated with most eukaryooc genes are control elements a. Control elements = segments of noncoding DNA that help regulate transcripoon by binding certain proteins Enhancers = control rate of gene transcripoon AcOvators = will bind to the enhancer to help increase the rate 74
Control Elements = Transcrip.on Factors Enhancer distal control elements DNA Upstream Proximal control elements Exon Promoter Intron Exon Poly-A signal sequence Termination region Intron Exon Transcription Downstream Primary RNA transcript 5ʹ Exon Intron Exon Intron Exon RNA processing Cleaved 3ʹ end of primary transcript Intron RNA Poly-A signal Coding segment mrna 5ʹ Cap 5ʹ UTR Start codon Stop codon 3ʹ UTR Poly-A tail 3ʹ
2. TranscripOonal regulaoon a. Control elements Proximal control elements are located close to the promoter Enhancers = Distal control elements (may be far away from a gene or even located in an intron) AcOvators = A protein that binds to an enhancer and somulates transcripoon of a gene Bound acovators cause mediator proteins to interact with proteins at the promoter 76
DNA Enhancer Activators Distal control element DNA-bending protein Promoter TATA box Gene General transcription factors Group of mediator proteins RNA polymerase II Transcription initiation complex RNA polymerase II RNA synthesis
2. TranscripOonal regulaoon b. Combinatorial control of gene acovaoon Unlike the genes of a prokaryooc operon, each of the coordinately controlled eukaryooc genes has a promoter and control elements These genes can be scaeered over different chromosomes, but each has the same combinaoon of control elements Copies of the acovators recognize specific control elements and promote simultaneous transcripoon of the genes 78
3. Post-TranscripOonal RegulaOon TranscripOon alone does not account for gene expression. Regulatory mechanisms can operate at various stages aper transcripoon. These mechanisms allow a cell to rapidly respond to environmental changes by using: a. RNA processing b. mrna degradaoon c. IniOaOon of translaoon d. Protein processing and degradaoon 79
Alterna4ve RNA Processing Exons DNA Troponin T gene Primary RNA transcript RNA splicing mrna or
Proteasome - Protein Degrada4on Ubiquitin Proteasome Proteasome and ubiquitin to be recycled Protein to be degraded Ubiquitinated protein Protein entering a proteasome Protein fragments (peptides)
Modifiers of Transla4on Hairpin mirna Hydrogen bond Dicer 5ʹ 3ʹ (a) Primary mirna transcript mirna mirnaprotein complex mrna degraded Translation blocked (b) Generation and function of mirnas
4. Noncoding RNAs The inhibi.on of gene expression by RNA molecules is called RNA interference (RNAi) RNAi is caused by small interfering RNAs (sirnas), which act as silencers a. micrornas (mirnas) b. Small interfering RNAs (sirnas) 83
4. Noncoding RNAs The inhibioon of gene expression by RNA molecules is called RNA interference (RNAi) RNAi is caused by small interfering RNAs (sirnas) a. micrornas (mirnas) b. Small interfering RNAs (sirnas) a. sirnas play a role in heterochroma.n forma.on and can block large regions of the chromosome b. Small RNAs may also block transcrip.on of specific genes 84
IV. DifferenOal Gene Expression and SpecializaOon A program of differenoal gene expression leads to the different cell types in a mulocellular organism During embryonic development, a fer.lized egg gives rise to many different cell types (stem cells and differenoaoon) 85
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IV. DifferenOal Gene Expression and SpecializaOon Cell types are organized successively into.ssues, organs, organ systems, and the whole organism 87
IV. DifferenOal Gene Expression and SpecializaOon Gene expression orchestrates the developmental programs of animals Embryonic Development: The transformaoon from zygote to adult results from cell division, cell differenoaoon, and morphogenesis 88
Fig. 18-14 (a) Fertilized eggs of a frog (b) Newly hatched tadpole
IV. DifferenOal Gene Expression and SpecializaOon A. Cell DifferenOaOon B. Morphogenesis C. Cytoplasmic Determinants D. InducOon E. DeterminaOon F. PaEern FormaOon 90
A. Cell DifferenOaOon 91
B. Morphogenesis = Biological process in determining the shapes of Ossues, organs, and the whole organism 92
C. Cytoplasmic Determinants A. An egg s cytoplasm contains RNA, proteins, and other substances that are distributed unevenly in the unferolized egg B. Cytoplasmic determinants = maternal substances in the egg that influence early development C. As the zygote divides by mitosis, cells contain different cytoplasmic determinants, which lead to different gene expression 93
Differen4al Gene Expression Unfertilized egg cell Sperm Nucleus Fertilization Two different cytoplasmic determinants Early embryo (32 cells) NUCLEUS Zygote Mitotic cell division Signal transduction pathway Signal receptor Two-celled embryo Signal molecule (inducer) (a) Cytoplasmic determinants in the egg (b) Induction Signals by nearby cells
Nucleus MyoD Protein - Muscle Cell Differen4a4on Master regulatory gene myod Other muscle-specific genes Embryonic precursor cell DNA OFF OFF mrna OFF Myoblast (determined) MyoD protein (transcription factor) mrna mrna mrna mrna Part of a muscle fiber (fully differentiated cell) MyoD Another transcription factor Myosin, other muscle proteins, and cell cycle blocking proteins
Extra The gene regula1on systems that go wrong during cancer are the very same systems involved in embryonic development. Cancer can be caused by muta1ons to genes that regulate cell growth and division. Tumor viruses can cause cancer in animals including humans 96
Extra Oncogenes are cancer-causing genes. Proto-oncogenes are the corresponding normal cellular genes that are responsible for normal cell growth and division. Conversion of a proto-oncogene to an oncogene can lead to abnormal somulaoon of the cell cycle. 97
Proto-ongogene to Ongogene DNA Proto-oncogene Translocation or transposition: Gene amplification: Point mutation: within a control element within the gene New promoter Oncogene Oncogene Normal growthstimulating protein in excess Normal growth-stimulating protein in excess Normal growthstimulating protein in excess Hyperactive or degradationresistant protein
Extra Tumor-suppressor genes help prevent uncontrolled cell growth. Muta1ons that decrease protein products of tumorsuppressor genes may contribute to cancer onset. Tumor-suppressor proteins: Repair damaged DNA & control cell adhesion. Inhibit the cell cycle in the cell-signaling pathway. 99
Extra MulOple mutaoons are generally needed for fullfledged cancer; thus the incidence increases with age. At the DNA level, a cancerous cell is usually characterized by at least one acove oncogene and the mutaoon of several tumor-suppressor genes. 100
Mul.-Step Model of Cancer Development Colon EFFECTS OF MUTATIONS Colon wall 1 Loss of tumorsuppressor gene APC (or other) 2 Activation of ras oncogene 4 Loss of tumor-suppressor gene p53 Normal colon epithelial cells Small benign growth (polyp) 3 Loss of tumor-suppressor gene DCC Larger benign growth (adenoma) 5 Additional mutations Malignant tumor (carcinoma)
Review Genes not expressed Promoter Genes expressed Operator Genes Fig. 18-UN2 Active repressor: no inducer present Inactive repressor: inducer bound
Regula.on of Gene Expression Chromatin modification Genes in highly compacted chromatin are generally not transcribed. Histone acetylation seems to loosen chromatin structure, enhancing transcription. Transcription Regulation of transcription initiation: DNA control elements bind specific transcription factors. DNA methylation generally reduces transcription. Bending of the DNA enables activators to contact proteins at the promoter, initiating transcription. Coordinate regulation: Enhancer for liver-specific genes Enhancer for lens-specific genes Chromatin modification Transcription RNA processing Primary RNA transcript RNA processing Alternative RNA splicing: mrna degradation Translation mrna or Protein processing and degradation Translation mrna degradation Each mrna has a characteristic life span, determined in part by sequences in the 5ʹ and 3ʹ UTRs. Initiation of translation can be controlled via regulation of initiation factors. Protein processing and degradation Protein processing and degradation by proteasomes are subject to regulation.