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11/6/2011 Chapter 18- Regulation of Gene Expression Overview: Conducting the Genetic Orchestra Prokaryotes and eukaryotes alter gene expression in response to their changing environment In multicellular eukaryotes, gene expression regulates development and is responsible for differences in cell types RNA molecules play many roles in regulating gene expression in eukaryotes © 2011 Pearson Education, Inc. Regulating Gene Expression Bacteria often respond to environmental change by regulating transcription 1. Control amount of mRNA that is transcribed Natural selection has favored bacteria that produce only the products needed by that cell 2. Control the rate of translation A cell can regulate the production of enzymes by feedback inhibition or by gene regulation 3. Control the activity of the protein Gene expression in bacteria is controlled by the operon model © 2011 Pearson Education, Inc. Constitutively expressed genes Constitutively expressed genes code for proteins that are always needed, therefore they are always being transcribed Other genes transcribed only when the proteins are needed Gene Expression in Prokaryotes Gene expression in bacteria was first studied in e. coli The work was done by Jacob and Monod in 1961 1 11/6/2011 Bacteria often respond to environmental change by regulating transcription Figure 18.2 Precursor Feedback inhibition Natural selection has favored bacteria that produce only the products needed by that cell A cell can regulate the production of enzymes by feedback inhibition or by gene regulation trpE gene Enzyme 1 trpD gene Enzyme 2 Regulation of gene expression trpC gene trpB gene Enzyme 3 Gene expression in bacteria is controlled by the operon model trpA gene Tryptophan (a) Regulation of enzyme activity (b) Regulation of enzyme production © 2011 Pearson Education, Inc. Operons: The Basic Concept A cluster of functionally related genes can be under coordinated control by a single “on-off switch” The regulatory “switch” is a segment of DNA called an operator usually positioned within the promoter An operon is the entire stretch of DNA that includes the operator, the promoter, and the genes that they control © 2011 Pearson Education, Inc. Corepressor The repressor can be in an active or inactive form, depending on the presence of other molecules A corepressor is a molecule that cooperates with a repressor protein to switch an operon off For example, E. coli can synthesize the amino acid tryptophan © 2011 Pearson Education, Inc. Repressor Protein The operon can be switched off by a protein repressor The repressor prevents gene transcription by binding to the operator and blocking RNA polymerase The repressor is the product of a separate regulatory gene © 2011 Pearson Education, Inc. Trp Operon By default the trp operon is on and the genes for tryptophan synthesis are transcribed When tryptophan is present, it binds to the trp repressor protein, which turns the operon off The repressor is active only in the presence of its corepressor tryptophan; thus the trp operon is turned off (repressed) if tryptophan levels are high © 2011 Pearson Education, Inc. 2 11/6/2011 Figure 18.3b-1 Repressable Operon - Trp Operon DNA trp operon Promoter Promoter Genes of operon DNA trpR Regulatory gene mRNA trpE 3 RNA polymerase Operator Start codon trpD trpC trpB trpA C B A mRNA Stop codon mRNA 5 5 E Protein Inactive repressor D Protein Polypeptide subunits that make up enzymes for tryptophan synthesis Active repressor (a) Tryptophan absent, repressor inactive, operon on Tryptophan (corepressor) (b) Tryptophan present, repressor active, operon off Figure 18.3b-2 Repressible and Inducible Operons: Two Types of Negative Gene Regulation DNA No RNA made mRNA A repressible operon is one that is usually on; binding of a repressor to the operator shuts off transcription The trp operon is a repressible operon Protein Active repressor Tryptophan (corepressor) An inducible operon is one that is usually off; a molecule called an inducer inactivates the repressor and turns on transcription (b) Tryptophan present, repressor active, operon off © 2011 Pearson Education, Inc. Inducible Operon – Lac Operon The lac operon is an inducible operon and contains genes that code for enzymes used in the hydrolysis and metabolism of lactose Lactose is an Inducer of the lac Operon When lactose is present, it is converted to allolactose Allolactose binds to the lactose repressor protein By itself, the lac repressor is active and switches the lac operon off A molecule called an inducer inactivates the repressor to turn the lac operon on Allolactose is an allosteric regulator because it binds to the repressor protein at a site other than the active site It causes the lactose repressor protein to leave its place on the operator region of the lac operon Now transcription will proceed © 2011 Pearson Education, Inc. 3 11/6/2011 Lactose Digestion Lactose is a disaccharide. The enzyme β–galactosidase splits the disaccharide into glucose and galactose The enzyme lactose permease functions to transport lactose into the bacteria The enzyme lactose transacetylase also has a role E. coli normally have low levels of these enzymes, but if they are grown in a media of lactose they will produce large quantities of these enzymes Figure 18.4a Figure 18.4b Regulatory gene Promoter Operator lac operon DNA lacI lacZ No RNA made 3 lacI DNA lacZ 5 Protein RNA polymerase Active repressor lacA 3 mRNA 5 mRNA lacY RNA polymerase mRNA 5 -Galactosidase Protein Allolactose (inducer) Permease Transacetylase Inactive repressor (b) Lactose present, repressor inactive, operon on (a) Lactose absent, repressor active, operon off Inducible vs Repressible Positive Gene Regulation Inducible enzymes usually function in catabolic pathways; their synthesis is induced by a chemical signal Some operons are also subject to positive control through a stimulatory protein, such as catabolite activator protein (CAP), an activator of transcription Repressible enzymes usually function in anabolic pathways; their synthesis is repressed by high levels of the end product When glucose (a preferred food source of E. coli) is scarce, CAP is activated by binding with cyclic AMP (cAMP) Regulation of the trp and lac operons involves negative control of genes because operons are switched off by the active form of the repressor Activated CAP attaches to the promoter of the lac operon and increases the affinity of RNA polymerase, thus accelerating transcription © 2011 Pearson Education, Inc. © 2011 Pearson Education, Inc. 4 11/6/2011 Figure 18.5a Positive Gene Regulation Promoter When glucose levels increase, CAP detaches from the lac operon, and transcription returns to a normal rate CAP helps regulate other operons that encode enzymes used in catabolic pathways lacI lacZ CAP-binding site Operator DNA Active CAP cAMP RNA polymerase binds and transcribes Inactive lac repressor Inactive CAP Allolactose (a) Lactose present, glucose scarce (cAMP level high): abundant lac mRNA synthesized © 2011 Pearson Education, Inc. Figure 18.5b Promoter lacI DNA CAP-binding site lacZ Operator RNA polymerase less likely to bind Inactive CAP Inactive lac repressor (b) Lactose present, glucose present (cAMP level low): little lac mRNA synthesized Figure 13-5 Page 262 DNA cAMP CAP dimer 5 11/6/2011 Examples of Hormone induced responses mediated by cAMP Target Tissue Adrenal Cortex Ovary Muscle Bone Heart Liver Kidney Fat Hormone Major Response ACTH Cortisol Secretion LH Progesterone secretion Adrenaline Glycogen breakdown Parathyroid Bone reabsorption hormone (PTH) Adrenaline Increase heart rate Glucagon Glycogen breakdown Vasopressin Water reabsorption Adrenaline, Triglyceride ACTH, glucagon breakdown Eukaryotic gene expression is regulated at many stages All organisms must regulate which genes are expressed at any given time In multicellular organisms regulation of gene expression is essential for cell specialization © 2011 Pearson Education, Inc. Differential Gene Expression Almost all the cells in an organism are genetically identical Figure 18.6 Signal NUCLEUS Chromatin Chromatin modification: DNA unpacking involving histone acetylation and DNA demethylation DNA Gene available for transcription Gene Transcription RNA Exon Primary transcript Differences between cell types result from differential gene expression, the expression of different genes by cells with the same genome Intron RNA processing Cap Transport to cytoplasm CYTOPLASM mRNA in cytoplasm Degradation of mRNA Abnormalities in gene expression can lead to diseases including cancer Translation Polypeptide Protein processing, such as cleavage and chemical modification Degradation of protein Gene expression is regulated at many stages Tail mRNA in nucleus Active protein Transport to cellular destination Cellular function (such as enzymatic activity, structural support) © 2011 Pearson Education, Inc. 6 11/6/2011 Figure 18.6a Figure 18.6b Signal CYTOPLASM mRNA in cytoplasm NUCLEUS Chromatin DNA Gene available for transcription Polypeptide Protein processing, such as cleavage and chemical modification Gene Transcription RNA Exon Primary transcript Intron Degradation of protein RNA processing Cap Translation Degradation of mRNA Chromatin modification: DNA unpacking involving histone acetylation and DNA demethylation Tail mRNA in nucleus Transport to cellular destination Cellular function (such as enzymatic activity, structural support) Transport to cytoplasm CYTOPLASM Regulation of Chromatin Structure Active protein Histone Modifications Genes within highly packed heterochromatin are usually not expressed In histone acetylation, acetyl groups are attached to positively charged lysines in histone tails Chemical modifications to histones and DNA of chromatin influence both chromatin structure and gene expression This loosens chromatin structure, thereby promoting the initiation of transcription The addition of methyl groups (methylation) can condense chromatin; the addition of phosphate groups (phosphorylation) next to a methylated amino acid can loosen chromatin © 2011 Pearson Education, Inc. © 2011 Pearson Education, Inc. Figure 18.7 Histone tails Amino acids available for chemical modification DNA double helix Nucleosome (end view) (a) Histone tails protrude outward from a nucleosome Animation: DNA Packing Right-click slide / select “Play” Acetylated histones Unacetylated histones (b) Acetylation of histone tails promotes loose chromatin structure that permits transcription © 2011 Pearson Education, Inc. 7 11/6/2011 Histone Modifications The histone code hypothesis proposes that specific combinations of modifications, as well as the order in which they occur, help determine chromatin configuration and influence transcription DNA Methylation DNA methylation, the addition of methyl groups to certain bases in DNA, is associated with reduced transcription in some species DNA methylation can cause long-term inactivation of genes in cellular differentiation In genomic imprinting, methylation regulates expression of either the maternal or paternal alleles of certain genes at the start of development © 2011 Pearson Education, Inc. Regulation of Transcription Initiation Chromatin-modifying enzymes provide initial control of gene expression by making a region of DNA either more or less able to bind the transcription machinery © 2011 Pearson Education, Inc. Organization of a Typical Eukaryotic Gene Associated with most eukaryotic genes are multiple control elements, segments of noncoding DNA that serve as binding sites for transcription factors that help regulate transcription Control elements and the transcription factors they bind are critical to the precise regulation of gene expression in different cell types © 2011 Pearson Education, Inc. Eukaryotic Promoter region © 2011 Pearson Education, Inc. Fig. 16.9 RNA polymerase binds to the promoter region In eukaryotes the DNA has a “TATA box” just upstream from where the RNA polymerase binds It is located upstream from the genes (25 – 35 bases upstream) 8 11/6/2011 Figure 18.8-1 Enhancer (distal control elements) Figure 18.8-2 Proximal control elements DNA Upstream Transcription start site Exon Intron Exon Poly-A signal sequence Intron Exon Transcription termination region Downstream Promoter Enhancer (distal control elements) DNA Upstream Proximal control elements Transcription start site Exon Enhancer (distal control elements) DNA Upstream Exon Intron Exon Promoter Primary RNA transcript 5 (pre-mRNA) Figure 18.8-3 Intron Intron Poly-A signal sequence Exon Transcription Exon Intron Transcription termination region Downstream Poly-A signal Exon Cleaved 3 end of primary transcript The Roles of Transcription Factors Proximal control elements Transcription start site Exon Intron Exon Intron Exon Promoter Poly-A signal sequence Intron Exon Transcription termination region Downstream Poly-A signal Exon Cleaved 3 end of primary RNA processing transcript To initiate transcription, eukaryotic RNA polymerase requires the assistance of proteins called transcription factors Transcription Exon Primary RNA transcript 5 (pre-mRNA) Intron General transcription factors are essential for the transcription of all protein-coding genes Intron RNA Coding segment mRNA G P P 5 Cap AAA AAA P 5 UTR Start codon Stop codon 3 3 UTR Poly-A tail In eukaryotes, high levels of transcription of particular genes depend on control elements interacting with specific transcription factors © 2011 Pearson Education, Inc. Enhancers and Specific Transcription Factors Transcription Factors Proximal control elements are located close to the promoter An activator is a protein that binds to an enhancer and stimulates transcription of a gene Distal control elements, groupings of which are called enhancers, may be far away from a gene or even located in an intron Activators have two domains, one that binds DNA and a second that activates transcription Bound activators facilitate a sequence of protein-protein interactions that result in transcription of a given gene © 2011 Pearson Education, Inc. © 2011 Pearson Education, Inc. 9 11/6/2011 Fig. 16.11 Enhancer Elements Eukaryotes have enhancer elements Regulatory proteins bind to the enhancer elements When the regulatory proteins are bound then the enhancer elements increase the rate of transcription by helping the RNA polymerase bind and begin transcription Fig. 16.12 Transcription Factors Proteins that regulate transcription in eukaryotes are called transcription factors. There are thousands of transcription factors, both repressors and enhancers Examples of common motifs found in these proteins: Helix-turn-helix Zinc fingers Leucine zipper proteins Turn COO- Finger 2 Finger 3 Zinc ion a-helix Finger 1 Leucine zipper region NH3+ DNA (a) DNA (b) DNA (c) Animation: Initiation of Transcription Right-click slide / select “Play” © 2011 Pearson Education, Inc. 10 11/6/2011 Figure 18.9 Transcription Factors Some transcription factors function as repressors, inhibiting expression of a particular gene by a variety of methods Activation domain DNA-binding domain DNA Some activators and repressors act indirectly by influencing chromatin structure to promote or silence transcription © 2011 Pearson Education, Inc. Figure 18.10-1 Promoter Activators DNA Enhancer Distal control element Figure 18.10-2 Gene TATA box Promoter Activators DNA Enhancer Distal control element Gene TATA box General transcription factors DNAbending protein Group of mediator proteins Figure 18.10-3 Promoter Activators DNA Enhancer Distal control element Gene TATA box General transcription factors DNAbending protein Group of mediator proteins RNA polymerase II RNA polymerase II Transcription initiation complex Coordinately Controlled Genes in Eukaryotes Unlike the genes of a prokaryotic operon, each of the co-expressed eukaryotic genes has a promoter and control elements These genes can be scattered over different chromosomes, but each has the same combination of control elements Copies of the activators recognize specific control elements and promote simultaneous transcription of the genes RNA synthesis © 2011 Pearson Education, Inc. 11 11/6/2011 Mechanisms of Post-Transcriptional Regulation Transcription alone does not account for gene expression Regulatory mechanisms can operate at various stages after transcription RNA Processing In alternative RNA splicing, different mRNA molecules are produced from the same primary transcript, depending on which RNA segments are treated as exons and which as introns Such mechanisms allow a cell to fine-tune gene expression rapidly in response to environmental changes © 2011 Pearson Education, Inc. © 2011 Pearson Education, Inc. Figure 18.13 Exons DNA 1 3 2 4 5 4 5 Troponin T gene Primary RNA transcript 3 2 1 RNA splicing Animation: RNA Processing mRNA 1 2 3 5 or 1 2 4 5 Right-click slide / select “Play” © 2011 Pearson Education, Inc. mRNA Degradation The life span of mRNA molecules in the cytoplasm is a key to determining protein synthesis Eukaryotic mRNA is more long lived than prokaryotic mRNA Nucleotide sequences that influence the lifespan of mRNA in eukaryotes reside in the untranslated region (UTR) at the 3 end of the molecule Animation: mRNA Degradation Right-click slide / select “Play” © 2011 Pearson Education, Inc. © 2011 Pearson Education, Inc. 12 11/6/2011 Initiation of Translation The initiation of translation of selected mRNAs can be blocked by regulatory proteins that bind to sequences or structures of the mRNA Alternatively, translation of all mRNAs in a cell may be regulated simultaneously For example, translation initiation factors are simultaneously activated in an egg following fertilization Animation: Blocking Translation Right-click slide / select “Play” © 2011 Pearson Education, Inc. © 2011 Pearson Education, Inc. Protein Processing and Degradation After translation, various types of protein processing, including cleavage and the addition of chemical groups, are subject to control Proteasomes are giant protein complexes that bind protein molecules and degrade them Animation: Protein Processing Right-click slide / select “Play” © 2011 Pearson Education, Inc. © 2011 Pearson Education, Inc. Figure 18.14 Proteasome and ubiquitin to be recycled Ubiquitin Proteasome Protein to be degraded Ubiquitinated protein Protein entering a proteasome Protein fragments (peptides) Animation: Protein Degradation Right-click slide / select “Play” © 2011 Pearson Education, Inc. 13 11/6/2011 Noncoding RNAs play multiple roles in controlling gene expression Effects on mRNAs by MicroRNAs and Small Interfering RNAs Only a small fraction of DNA codes for proteins, and a very small fraction of the non-protein-coding DNA consists of genes for RNA such as rRNA and tRNA MicroRNAs (miRNAs) are small single-stranded RNA molecules that can bind to mRNA These can degrade mRNA or block its translation A significant amount of the genome may be transcribed into noncoding RNAs (ncRNAs) Noncoding RNAs regulate gene expression at two points: mRNA translation and chromatin configuration © 2011 Pearson Education, Inc. © 2011 Pearson Education, Inc. Figure 18.15 MicroRNAs and Small Interfering RNAs Hairpin Hydrogen bond miRNA Dicer 5 3 (a) Primary miRNA transcript miRNA miRNAprotein complex The phenomenon of inhibition of gene expression by RNA molecules is called RNA interference (RNAi) RNAi is caused by small interfering RNAs (siRNAs) siRNAs and miRNAs are similar but form from different RNA precursors mRNA degraded Translation blocked (b) Generation and function of miRNAs © 2011 Pearson Education, Inc. A program of differential gene expression leads to the different cell types in a multicellular organism During embryonic development, a fertilized egg gives rise to many different cell types A Genetic Program for Embryonic Development The transformation from zygote to adult results from cell division, cell differentiation, and morphogenesis Cell types are organized successively into tissues, organs, organ systems, and the whole organism Gene expression orchestrates the developmental programs of animals © 2011 Pearson Education, Inc. © 2011 Pearson Education, Inc. 14 11/6/2011 Figure 18.16 Cell Differentiation and morphogenesis Cell differentiation is the process by which cells become specialized in structure and function The physical processes that give an organism its shape constitute morphogenesis Differential gene expression results from genes being regulated differently in each cell type 2 mm 1 mm (a) Fertilized eggs of a frog (b) Newly hatched tadpole Materials in the egg can set up gene regulation that is carried out as cells divide © 2011 Pearson Education, Inc. Cytoplasmic Determinants and Inductive Signals An egg’s cytoplasm contains RNA, proteins, and other substances that are distributed unevenly in the unfertilized egg Cytoplasmic determinants are maternal substances in the egg that influence early development Maternal Effect Genes The developing oocyte gets “polarized” by mRNA from the mother’s nurse cells. The mRNA is given to the oocyte prior to fertilization This is what determines anterior vs posterior As the zygote divides by mitosis, cells contain different cytoplasmic determinants, which lead to different gene expression © 2011 Pearson Education, Inc. Figure 18.17a (a) Cytoplasmic determinants in the egg Cell Signaling - Induction Unfertilized egg Sperm Fertilization Zygote (fertilized egg) Mitotic cell division Nucleus Molecules of two different cytoplasmic determinants The other important source of developmental information is the environment around the cell, especially signals from nearby embryonic cells In the process called induction, signal molecules from embryonic cells cause transcriptional changes in nearby target cells Thus, interactions between cells induce differentiation of specialized cell types Two-celled embryo © 2011 Pearson Education, Inc. 15 11/6/2011 Figure 18.17b (b) Induction by nearby cells Early embryo (32 cells) NUCLEUS Signal transduction pathway Signal receptor Animation: Cell Signaling Signaling molecule (inducer) Right-click slide / select “Play” © 2011 Pearson Education, Inc. Sequential Regulation of Gene Expression During Cellular Differentiation Determination commits a cell to its final fate Determination precedes differentiation Cell differentiation is marked by the production of tissue-specific proteins © 2011 Pearson Education, Inc. Pattern Formation: Setting Up the Body Plan Pattern formation is the development of a spatial organization of tissues and organs In animals, pattern formation begins with the establishment of the major axes Positional information, the molecular cues that control pattern formation, tells a cell its location relative to the body axes and to neighboring cells © 2011 Pearson Education, Inc. The Drosophila Fly Development in drosophila flies have stages: Egg and sperm fuse to produce a zygote The zygote undergoes embryonic development to become a larva The larva undergoes several molts and grow until it becomes a pupa A pupa has a hardened external cuticle The pupa undergoes metamorphosis and becomes a mature adult fly 96 16 11/6/2011 The Life Cycle of Drosophila Figure 18.19 Head Thorax Abdomen Follicle cell 1 Egg developing within ovarian follicle Nucleus Egg 0.5 mm Nurse cell In Drosophila, cytoplasmic determinants in the unfertilized egg determine the axes before fertilization Dorsal BODY AXES Anterior Left Right Posterior 2 Unfertilized egg Depleted nurse cells Ventral (a) Adult Egg shell Fertilization Laying of egg 3 Fertilized egg After fertilization, the embryo develops into a segmented larva with three larval stages Embryonic development 4 Segmented embryo 0.1 mm Body segments Hatching 5 Larval stage (b) Development from egg to larva © 2011 Pearson Education, Inc. Axis Establishment Maternal effect genes encode for cytoplasmic determinants that initially establish the axes of the body of Drosophila These maternal effect genes are also called eggpolarity genes because they control orientation of the egg and consequently the fly Animation: Development of Head-Tail Axis in Fruit Flies Right-click slide / select “Play” © 2011 Pearson Education, Inc. Maternal Effect Genes The developing oocyte gets “polarized” by mRNA from the mother’s nurse cells. The mRNA is given to the oocyte prior to fertilization This is what determines anterior vs posterior © 2011 Pearson Education, Inc. Establishment of the A/P axis Nurse cells secrete maternally produced bicoid and nanos mRNAs into the oocyte The two types of mRNA are transported by microtubules to opposite poles of the oocyte bicoid mRNA to the future anterior pole nanos mRNA to the future posterior pole 17 11/6/2011 Bicoid: A Morphogen Determining Head Structures Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Movement of bicoid mRNA moves maternal mRNA toward anterior end Follicle cells Nurse cells One maternal effect gene, the bicoid gene, affects the front half of the body Anterior An embryo whose mother has no functional bicoid gene lacks the front half of its body and has duplicate posterior structures at both ends Posterior Microtubules nanos mRNA moves toward posterior end a. Nucleus Anterior Posterior bicoid mRNA Establishment of the A/P axis nanos mRNA b. © 2011 Pearson Education, Inc. Fig. 19.13.a1 After fertilization, translation of the bicoid and nano mRNA will create opposing gradients of Bicoid and Nanos proteins Hunchback and Caudal Fig. 19.15.a The bicoid and nano proteins control (inhibit) the translation of two other maternal mRNAs: Hunchback and Caudal mRNAs code for transcription factors which control genes necessary for anterior and posterior (abdominal) structures 18 11/6/2011 Fig. 19.15.b Fig. 19.15.c Figure 18.21 Figure 18.22 Head Tail 100 m RESULTS Anterior end A8 T1 T2 T3 A1 A2 A3 A4 A5 A6 Wild-type larva A7 250 m Fertilization, translation of bicoid mRNA Bicoid mRNA in mature unfertilized egg Tail Bicoid protein in early embryo Tail A8 A8 A7 A6 A7 Bicoid mRNA in mature unfertilized egg Bicoid protein in early embryo Mutant larva (bicoid) Hunchback gene The bicoid research is important for three reasons – It identified a specific protein required for some early steps in pattern formation – It increased understanding of the mother’s role in embryo development – It demonstrated a key developmental principle that a gradient of molecules can determine polarity and position in the embryo Bicoid protein is also a transcription factor for the embryo’s hunchback gene. The hunchback mRNA is the first to be transcribed in the embryo after fertilization. There is both maternal and embryonic hunchback mRNA being transcribed in the embryo © 2011 Pearson Education, Inc. 19 11/6/2011 Hunchback gene Segmentation Nüsslein-Volhard and Wieschaus studied segment formation Hunchback gene is a type of gap gene, which maps out the largest subdivisions of the A/P axis in the embryo The gap genes (there are nine gap genes) code for transcription factors for pair-rule genes They created mutants, conducted breeding experiments, and looked for corresponding genes Many of the identified mutations were embryonic lethals, causing death during embryogenesis They found 120 genes essential for normal segmentation © 2011 Pearson Education, Inc. Fig. 19.13.a2 Segmentation Genes The embryo DNA produces mRNA Some of the embryonic genes code for the body segments and pattern of the organism 1. Gap genes – refine the broad regions set by maternal genes into more defined regions along A/P axis 2. Pair-rule genes - Divide the embryo into seven zones 3. Segment polarity genes - finish defining the embryonic segments Most segmentation genes code for transcription factors Fig. 19.13.b2 Fig. 19.13.c2 20 11/6/2011 Fig. 19.13.d2 Homeotic Genes After the segmentation genes have established the pattern of segments, then homeotic genes code for the development plan for the segments Like segmentation genes, these genes code mainly for transcription factors HOX genes Production of Body Plan All the homeotic genes have a highly conserved region that codes for a DNA binding domain in the transcription factors Drosophila HOM Chromosomes Antennapedia complex a. Lewis discovered the homeotic genes, which control pattern formation in late embryo, larva, and adult stages Hox 2 Ubxabd-Aabd-B Hox 3 Hox 4 Abdomen Therefore they are referred to as Hox genes since they all contain the “homeodomain” Edward B. Lewis, Christiane Nüsslein-Volhard, and Eric Wieschaus won a Nobel Prize in 1995 for decoding pattern formation in Drosophila Hox 1 Bithorax complex lab pb Dfd Scr Antp Head Thorax Genetic Analysis of Early Development: Scientific Inquiry Mouse Hox Chromosomes Drosophila HOM genes Fruit fly embryo Mouse embryo Fruit fly Mouse b. Figure 18.20 Eye Leg Antenna Wild type Mutant © 2011 Pearson Education, Inc. 21 11/6/2011 Cancer results from genetic changes that affect cell cycle control The gene regulation systems that go wrong during cancer are the very same systems involved in embryonic development © 2011 Pearson Education, Inc. Proto-oncogenes Types of Genes Associated with Cancer Cancer can be caused by mutations to genes that regulate cell growth and division Tumor viruses can cause cancer in animals including humans © 2011 Pearson Education, Inc. Figure 18.23 Oncogenes are cancer-causing genes Proto-oncogene DNA Proto-oncogenes are the corresponding normal cellular genes that are responsible for normal cell growth and division Translocation or transposition: gene moved to new locus, under new controls Gene amplification: multiple copies of the gene New promoter Conversion of a proto-oncogene to an oncogene can lead to abnormal stimulation of the cell cycle Normal growthstimulating protein in excess Point mutation: within a control within element the gene Oncogene Normal growth-stimulating protein in excess Normal growthstimulating protein in excess Oncogene Hyperactive or degradationresistant protein © 2011 Pearson Education, Inc. Oncogenes Proto-oncogenes can be converted to oncogenes by Movement of DNA within the genome: if it ends up near an active promoter, transcription may increase Amplification of a proto-oncogene: increases the number of copies of the gene Point mutations in the proto-oncogene or its control elements: cause an increase in gene expression © 2011 Pearson Education, Inc. Tumor-Suppressor Genes Tumor-suppressor genes help prevent uncontrolled cell growth Mutations 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 © 2011 Pearson Education, Inc. 22 11/6/2011 Interference with Normal Cell-Signaling Pathways Figure 18.24a MUTATION 1 Growth factor Ras 3 G protein Mutations in the ras proto-oncogene and p53 tumorsuppressor gene are common in human cancers Mutations in the ras gene can lead to production of a hyperactive Ras protein and increased cell division P P P 2 Receptor GTP Ras P P P Hyperactive Ras protein (product of oncogene) issues signals on its own. GTP 4 Protein kinases (phosphorylation cascade) NUCLEUS 5 Transcription factor (activator) DNA Gene expression Protein that stimulates the cell cycle (a) Cell cycle–stimulating pathway © 2011 Pearson Education, Inc. P53 Gene – Tumor Suppressor Gene Figure 18.24b 2 Protein kinases Suppression of the cell cycle can be important in the case of damage to a cell’s DNA; p53 prevents a cell from passing on mutations due to DNA damage 3 Active form of p53 UV light 1 DNA damage in genome Mutations in the p53 gene prevent suppression of the cell cycle MUTATION Defective or missing transcription factor, such as p53, cannot activate transcription. DNA Protein that inhibits the cell cycle (b) Cell cycle–inhibiting pathway © 2011 Pearson Education, Inc. Figure 18.24c The Multistep Model of Cancer Development EFFECTS OF MUTATIONS Protein overexpressed Cell cycle overstimulated Protein absent Increased cell division Cell cycle not inhibited Multiple mutations 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 active oncogene and the mutation of several tumor-suppressor genes (c) Effects of mutations © 2011 Pearson Education, Inc. 23 11/6/2011 Figure 18.25 Inherited Predisposition and Other Factors Contributing to Cancer Individuals can inherit oncogenes or mutant alleles of tumor-suppressor genes Colon 1 Loss of tumorsuppressor gene APC (or other) Normal colon epithelial cells Colon wall 4 Loss of tumorsuppressor gene p53 2 Activation of ras oncogene Small benign growth (polyp) 3 Loss of tumorsuppressor gene DCC Larger benign growth (adenoma) 5 Additional mutations Mutations in the BRCA1 or BRCA2 gene are found in at least half of inherited breast cancers, and tests using DNA sequencing can detect these mutations Malignant tumor (carcinoma) © 2011 Pearson Education, Inc. Post-transcriptional control How long a mRNA remains active is important Bacterial mRNA has a short half life – minutes Eukaryotic mRNA undergoes posttranscriptional modification Eukaryotic mRNA can remain active for hours Post-translational Modification Modification of the protein or chain of amino acids: 1. Protein Kinases – phosphorylate proteins, activating them Post-transcriptional modifications 1. A 5’ cap is added to the 5’ end which may protect the mRNA from being degraded. 2. A poly-A tail is added near the 3’ end which may help the mRNA to be exported from the nucleus 3. Introns must be removed 4. Small RNAs can inhibit translation of mRNA Post-translational Modification 4. Glycosylation – sugars are added to some proteins, some of these sugar chains are important to “address” the protein 5. Methionine is often removed 2. Enzyme inhibitors and activators 6. Proteases degrade the proteins 3. Proteolysis – some polypeptide chains are cut into smaller chains 7. Proteasome degrades proteins using the marker ubitiquin 24 11/6/2011 Important concepts Know the vocabulary of the lecture Know in detail how e. coli regulates the transcription of genes responsible for lactose digestion. Understand the inducible genes and the repressible genes Understand the differences between positive control and negative control of gene transcription Know how the lac operon is controlled by positive control, including role of cAMP and CAP, and Important Concepts Understand what leads to differences observed in cell types Know the stages of development in drosophila flies Understand what maternal effect genes are, include in your discussion, what the origin of the genes are, at what point in development do these genes appear, and what effect do they have on the organism. Understand what segmentation genes, Gap genes, and homeotic genes are and what they do. Important Concepts Understand how eukaryotes regulate gene transcription, including the role of TATA box, UPEs, and enhancers, the types of transcription factors Know how gene expression is controlled post transcriptionally and posttranslationally Important Concepts Know the basics of cell growth including the role of growth factors, growth factor receptors and protein kinases What are the main types of cancer genes – (oncogenes and tumor suppressor genes) and what do they do, examples of each. 25