File - Ms. Oldendorf's AP Biology
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Control of Prokaryotic (Bacterial) Genes AP Biology 2007-2008 Why regulate genes? Maintain homeostasis 1. - gene expression is affected by changes in the external environment 2. Multicellular organisms – have specialized cells AP Biology All cells have the full set of genes, but not all genes are needed in each cell (some are turned off) Bacterial metabolism Bacteria need to respond quickly to changes in their environment AP Biology Bacterial metabolism if they have enough of a product, need to stop production why? waste of energy to produce more how? stop production of enzymes for synthesis STOP AP Biology If they find a new food/energy source… need to utilize it quickly why? metabolism, growth, reproduction how? start production of enzymes for digestion GO Natural selection favors cells that can do this well because it’s COST EFFECTIVE AP Biology Enzyme Review AP Biology Allosteric Regulation AP Biology Remember Regulating Metabolism? Feedback inhibition product acts as an allosteric inhibitor of 1st enzyme in tryptophan pathway but this is wasteful production of enzymes Oh, I remember this from our Metabolism Unit! AP Biology - = inhibition - Different way to Regulate Metabolism Gene regulation AP Biology instead of blocking enzyme function, block transcription of genes for all enzymes in tryptophan pathway Now, that’s a good idea from a lowly bacterium! - = inhibition - - so why is this good? saves energy by not wasting it on unnecessary protein synthesis (remember…it’s all about energy!) AP Biology Overview - Gene regulation in bacteria Cells vary amount of specific enzymes by regulating gene transcription AP Biology turn genes on or turn genes off On and Off. Off and On. On and Off. STOP GO AP Biology Turn genes OFF example if bacterium has enough tryptophan then it doesn’t need to make enzymes used to build tryptophan Turn genes ON example if bacterium encounters new sugar (energy source), like lactose, then it needs to start making enzymes used to digest lactose Operon – What is the structure? Operon (has all of these) genes grouped together with related functions promoter = RNA polymerase binding site AP Biology example: all enzymes in a metabolic pathway single promoter controls transcription of all genes in operon transcribed as one unit & a single mRNA is made operator = DNA binding site of repressor protein So how can these genes be turned off? Repressor protein AP Biology binds to DNA at operator site blocking RNA polymerase blocks transcription Regulatory gene – codes for a regulatory protein, at different site than the gene it is regulating AP Biology Operon model Operon: operator, promoter & genes they control serve as a model for gene regulation RNA polymerase RNA repressor TATA polymerase gene1 gene2 gene3 gene4 1 2 3 4 enzyme1 enzyme2 enzyme3 enzyme4 mRNA promoter DNA operator Repressor protein turns off gene by blocking AP BiologyRNA polymerase binding site. repressor = repressor protein Synthesis Pathway Model When excess tryptophan is present, it binds to tryp repressor protein & triggers repressor to bind to DNA blocks (represses) transcription Usually on AP Biology Repressible operon: tryptophan (excess tryptophan is present so it binds to tryp repressor protein & triggers repressor to bind to DNA) RNA polymerase blocks (represses) transcription RNA trp repressor TATA polymerase gene1 gene2 gene3 gene4 1 2 3 4 enzyme1 enzyme2 enzyme3 enzyme4 mRNA promoter DNA trp operator trp trp repressor repressor protein trp trp trp trp trp trp conformational change in AP Biologyprotein! repressor trp repressor tryptophan trp tryptophan – repressor protein complex Tryptophan operon What happens when tryptophan is present? Don’t need to make tryptophan-building enzymes Tryptophan AP Biology is allosteric regulator of repressor protein Digestive Pathway Model When lactose is present, binds to lac repressor protein & triggers repressor to release DNA induces transcription Usually off AP Biology Inducible operon: lactose lac lac RNA polymerase (lactose is present so it binds to lac repressor protein & triggers repressor to release DNA) lac lac lac lac induces transcription lac RNA lac repressor TATA polymerase gene1 gene2 gene3 gene4 1 2 3 4 enzyme1 enzyme2 enzyme3 enzyme4 mRNA promoter operator repressor lac conformational change in AP Biologyprotein! repressor lac repressor DNA repressor protein lactose lactose – repressor protein complex Lactose operon What happens when lactose is present? Need to make lactose-digesting enzymes Lactose is allosteric regulator of repressor protein AP Biology 1961 | 1965 Jacob & Monod: lac Operon Francois Jacob & Jacques Monod first to describe operon system coined the phrase “operon” AP Biology Jacques Monod Francois Jacob Operon summary Repressible operon Usually on usually functions in anabolic pathways AP Biology synthesizing end products when end product is present in excess, gene is turned off and cell allocates resources to other uses Inducible operon Usually off usually functions in catabolic pathways, produce enzymes only when nutrient is available AP Biology digesting nutrients to simpler molecules cell avoids making proteins that have nothing to do, cell allocates resources to other uses Lac operon http://vcell.ndsu.edu/animations/lacOpe ron/index.htm AP Biology There’s more to the story… we have talked about negative regulation repressor protein binds to the operator, turns off transcription There is also positive regulation AP Biology activator binds to the promoter, turns transcription up (amplifies expression) But…there are other nutrients bacteria can use besides lactose Bacteria prefer glucose over lactose Glucose is needed for cell respiration to make ATP AP Biology When ATP is used up, it turns to ADP and even AMP Cyclic AMP – when ATP levels are low AP Biology Fig. 18-5 Promoter Operator DNA lacI CAP-binding site Active CAP cAMP Inactive CAP lacZ RNA polymerase binds and transcribes Inactive lac repressor Allolactose CAP (a) Lactose present, glucose scarce (cAMP level Regulation high): abundant lac mRNA synthesized Promoter DNA lacI CAP-binding site Inactive CAP AP Biology Operator lacZ RNA polymerase less likely to bind Inactive lac repressor (b) Lactose present, glucose present (cAMP level low): little lac mRNA synthesized CAP,cAMP, and Glucose As glucose supply increases, cAMP decreases, CAP is inactive, lac expression is slow/low As glucose supply decreases, cAMP increases, CAP is active, lac expression is fast/high AP Biology Why is it regulated 2 ways? Gives the bacteria choice If a bacteria is given both glucose and lactose, it chooses glucose Lactose causes the lac operon to be transcribed, but glucose (low cAMP) slows that transcription way down AP Biology Review Negative Regulation – a repressor protein binds to the promoter and turns transcription off Positive Regulation – an activator binds to the promoter and turns transcription on Lac – inducer (repressor unbinds) Trp – corepressor (repressor binds) Lac – CAP is an activator The lac operon is regulated in 2 ways Overall themes – homeostasis and energy/nutrient conservation AP Biology Rest of Today Complete the operon worksheet AP Biology Control of Eukaryotic Genes AP Biology 2007-2008 Yesterday Review Allosteric regulation Protein structure Transcription New stuff AP Biology Look at the structure of the operon (found in bacteria) Understand postive vs. negative gene regulation Understand repressible vs. inducible regulation Understand lac vs. trp operon regulation Prokaryotic vs eukaryotic Genomes Protein synthesis Cell respiration ETC. AP Biology The BIG Questions… How are genes turned on & off in eukaryotes? How do cells with the same genes differentiate to perform completely different, specialized functions? AP Biology Evolution of gene regulation – Why? Prokaryotes single-celled evolved to grow & divide rapidly must respond quickly to changes in external environment exploit transient resources – lac and trp operons Gene regulation turn genes on & off rapidly flexibility & reversibility adjust levels of enzymes for synthesis & digestion AP Biology Evolution of gene regulation Eukaryotes multicellular evolved to maintain constant internal conditions while facing changing external conditions homeostasis regulate body as a whole growth & development long term processes specialization turn on & off large number of genes - #18 20% of genes are turned on in a typical cell - # 19 AP Biology must coordinate the body as a whole rather than serve the needs of individual cells Points of control The control of gene expression can occur at any step in the pathway from gene to functional protein 1. packing/unpacking DNA 2. Transcription (common to all organisms) 3. mRNA processing 4. mRNA transport 5. translation 6. protein processing 7. protein degradation AP Biology 1. DNA packing How do you fit all that DNA into nucleus? DNA coiling & folding double helix nucleosomes chromatin fiber looped domains chromosome from DNA double helix to AP Biology chromosome condensed Nucleosomes 8 histone molecules “Beads on a string” 1st level of DNA packing histone proteins 8 protein molecules positively charged amino acids bind tightly to negatively charged DNA AP Biology DNA packing movie DNA packing as gene control (Epigenetic inheritance) Degree of packing of DNA regulates transcription tightly wrapped around histones no transcription genes turned off heterochromatin darker DNA (H) = tightly packed euchromatin lighter DNA (E) = loosely packed H AP Biology E Histone acetylation Acetylation of histones unwinds DNA loosely wrapped around histones attachment of acetyl groups (–COCH3) to histones AP Biology enables transcription genes turned on conformational change in histone proteins transcription factors have easier access to genes DNA methylation Methylation of DNA blocks transcription factors no transcription genes turned off attachment of methyl groups (–CH3) to cytosine nearly permanent inactivation of genes AP Biology C = cytosine ex. inactivated mammalian X chromosome = Barr body Can be passed down to new cells Genomic Imprinting When there is phenotypic variation due to which parent gave a certain gene (not sex-linked, found on autosomes) 1 allele is silenced by DNA methylation Only 1 copy of certain genes is needed for normal development If 0 or 2 genes is active, then major developmental problems can occur AP Biology 2. Transcription initiation Control regions on DNA promoter enhancer AP Biology nearby control sequence on DNA binding of RNA polymerase & transcription factors “base” rate of transcription distant control sequences on DNA binding of activator proteins “enhanced” rate (high level) of transcription Classes of Transcription Factors General – produce a low rate of transcription, needed for all genes Specific – interact with specific control elements (upstream of the gene) – greatly increases transcription rate AP Biology Model for Enhancer action Enhancer DNA sequences Activator proteins distant control sequences bind to enhancer sequence & stimulates transcription Silencer proteins bind to enhancer sequence & block gene transcription AP Biology Turning on Gene movie Transcription complex Activator Proteins • regulatory proteins bind to DNA at Enhancer Sites distant enhancer sites • increase the rate of transcription regulatory sites on DNA distant from gene Enhancer Activator Activator Activator Coactivator A E F B TFIID RNA polymerase II H Core promoter and initiation complex Initiation Complex at Promoter Site binding site of RNA polymerase AP Biology Signal transduction pathways Signal molecule can: 1) Bind to cell surface 1) 2) Cause activator or repressor to be turned on/off Diffuse through the cell 1) Bind to an activator or repressor to be turned on/off AP Biology Prokaryotic vs Eukaryotic Transcriptional Control Bacteria have operons regulated by 1 promoter Eukaryotes have related genes on different chromosomes with different promoters Eukaryotes have control elements for each gene AP Biology Control elements – specific nucleotide sequences, unique combination for each gene Coordinated control of genes in eukaryotes Related genes are clustered together Chromatin structure of that area is manipulated Turns all genes in that area on or off Related genes have the same combination of control elements AP Biology When those control elements are present, all related genes are turned on 3. Post-transcriptional control Alternative RNA splicing AP Biology variable processing of exons creates a family of proteins 4. Regulation of mRNA degradation Life span of mRNA determines amount of protein synthesis Eukaryotic mRNA can last from hours to weeks AP Biology RNA processing movie 5. Control of translation Block initiation of translation stage regulatory proteins attach to 5' end of mRNA prevent attachment of ribosomal subunits & initiator tRNA block translation of mRNA to protein AP Biology Control of translation movie 6-7. Protein processing & degradation Protein processing folding, cleaving, adding sugar groups, targeting for transport Protein degradation - #33-34 ubiquitin tagging proteasome degradation Ex. Cyclins – regulate cell cycle AP Biology Protein processing movie 1980s | 2004 Ubiquitin “Death tag” mark unwanted proteins with a label 76 amino acid polypeptide, ubiquitin labeled proteins are broken down rapidly in "waste disposers" AP proteasomes Aaron Ciechanover Biology Israel Avram Hershko Israel Irwin Rose UC Riverside Proteasome Protein-degrading “machine” breaks down any proteins into 7-9 amino acid fragments cellular recycling Cancer cells have mutations in cell cycle proteins that protect them from proteasomes AP Biology play Nobel animation 6 7 Gene Regulation protein processing & degradation 1 & 2. transcription - DNA packing - transcription factors 5 4 initiation of translation mRNA processing 3 & 4. post-transcription - mRNA processing - splicing - 5’ cap & poly-A tail - breakdown by siRNA 5. translation - block start of translation 1 2 initiation of transcription AP Biology mRNA splicing 3 6 & 7. post-translation - protein processing - protein degradation 4 mRNA protection RNA Review DNA is transcribed into Coding RNA Non-coding RNA AP Biology mRNA tRNA rRNA siRNA miRNA RNA interference DNA is transcribed into mRNA, tRNA, rRNA, and these: Small interfering RNAs (siRNA) short segments of RNA (21-28 bases) bind to mRNA create sections of double-stranded mRNA “death” tag for mRNA triggers degradation of mRNA cause gene “silencing” AP Biology post-transcriptional control siRNA turns off gene = no protein produced miRNA’s AP Biology Action of siRNA dicer enzyme mRNA for translation siRNA double-stranded miRNA + siRNA breakdown enzyme (RISC) mRNA degraded AP Biology functionally turns gene off Turn your Question Genes on! AP Biology 2007-2008 Rest of Class Create a model of the regulation of the lac operon When you are done, show me AP Biology 6 7 Gene Regulation 1 & 2. _________________ - ____________________ - ____________________ 5 4 1 2 3 & 4. _________________ - ____________________ - ____________________ - ____________________ - ____________________ 5. _________________ - ____________________ ____________________ AP Biology 3 4 6 & 7. _________________ - ____________________ - ____________________
