ASIM'S WEBWORLD View: Main School Main AP Biology Chapter Guides RELATED LINKS Review Questions and Answers		AP BIOLOGY: Chapter Sixteen Outline REGULATORY REGIONS DETERMINE GENE ACTIVITY Rationale for Controls Found in Bacteria Bacteria must exploit transient resources of environment Gene control adjusts cell's activities to fit environment Changes in gene expression are generally reversible Rationale for Controls Found in Multicellular Organisms Cells need to be protected from transient environmental changes Prefer constant conditions Homeostasis Maintenance of constant internal environment Hall mark of multicellular organisms Individual cells respond to signals in immediate environment Alter their gene expression Also participate in regulating body as a whole Changes in gene expression produce variety of results Compensate for changes in body's physiological condition Ensure that correct genes are expressed in development Genes must be transcribed in careful order, for specific time fig 16.1 Many genes activated only once Example: stem cells Develop onto differentiated tissues via strict genetic program Changes in gene expression serve needs of whole, not individual cell GENERAL PRINCIPLES OF TRANSCRIPTIONAL CONTROL Gene Expression Is Regulated at Many Levels fig 16.2 Transcriptional control Most common form in bacteria and eukaryotes Transcription controlled by RNA polymerase Post-transcriptional control Less common form of control Influences mRNA produced by genes Influences activity of proteins encoded by mRNA Control Expression by Controlling RNA Polymerase Polymerase must have access to DNA helix Must be able to bind to gene's promoter Specific nucleotide sequence at one end of gene Informs polymerase where to begin transcription Other sequences on DNA affect binding of polymerase and promoter Binding sites only 10 to 15 nucleotides long Hundreds of sequences characterized Each provides binding site for specific protein Binding can block RNA polymerase and transcription Binding can stimulate transcription by facilitating binding of polymerase to promoter HOW PROTEINS BIND TO SPECIFIC DNA SEQUENCES Molecular Recognition of Proteins to Regulatory Sequences DNA helix does not need to unwind for recognition by proteins Proteins bind to outer surface, at edges of base-pairs Major groove Deeper of two helical grooves that wind around DNA molecule fig 16.4 Nucleotide chemical groups are accessible, make unique patterns Protein-DNA recognition is being actively studied Proteins are unique, less variability where it actually binds to DNA Several different groups called structural motifs The Helix-Turn-Helix Motif Most common DNA-binding motif Two alpha-helical segments linked by short nonhelical segment fig 16.5,6a Segments are at right angles to one another Recognition helix fits into major groove of DNA Other helix butts up against outside of DNA Ensures proper positioning of recognition helix Regulatory sequences recognized by this motif occur in symmetrical pairs Sequences separated by distance equal to one turn of helix fig 16.7 Two sites doubles contact zone between protein and DNA The Homeodomain Motif Special class of helix-turn-helix motif Discovered in homeotic mutants of Drosophila fig 16.8 Mutations alter how body parts are assembled Mutant genes encode regulatory proteins that initiate key stages in development by binding to developmental switch-point genes Proteins have nearly identical sequence of 60 amino acids Sequence called homeodomain fig 16.6b Center of homeodomain is helix-turn-helix motif The Zinc Finger Motif Uses atoms of zinc to coordinate binding to DNA fig 16.6c In one form zinc links alpha-helical segment to beta-sheet segment Helical segment fits into major groove Often occurs in clusters Beta-sheet spaces helical segments to each contacts major groove Binding stronger with more zinc fingers Other forms replace beta-sheet with another helical segment The Leucine Zipper Motif Two protein subunits create a single DNA-binding site Subunits interact at leucines forming Y-shaped molecule Arms of Y fit into major groove fig 16.6d Allow great flexibility in controlling gene expression Related motif replaces leucines with two helix-turn-helix motifs TRANSCRIPTIONAL CONTROL IN BACTERIA Repressors Are "OFF" Switches Only genes that are directly needed are transcribed Others held in reserve Make enzymes to degrade a type of food only when it is present Example: tryptophan-producing (trp) genes in E. coli Cluster of five genes manufacturers tryptophan Unit called an operon, produces long strand of mRNA RNA polymerase binds to promoter at first gene, transcription ensues fig 16.9 Tryptophan present, trp genes shut off by repressor Helix-turn-helix regulatory protein, binds to trp promoter fig 16.10 Repressor at promoter prevents binding of RNA polymerase Repressor can't bind unless first bound to two tryptophans Tryptophan alters orientation of helix-turn-helix in repressor Recognition helices fit into DNA major groove fig 16.11 Synthesis of tryptophan tied to absence of it in environment Without it nothing activates repressor, transcription ensues With it, it binds to repressor, blocks transcription Activators Are "ON" Switches Some gene promotors constructed to be poor RNA polymerase binding sites Transcription of these genes rarely occurs unless promotor can bind better Requires transcriptional activator Binds to DNA nearby Holds polymerase against promoter, RNA polymerase binds better Example: catabolite activator protein (CAP) of E. coli fig 16.12 Initiates transcription of genes to utilize food when glucose absent Decreasing glucose leads to increase in intracellular cyclic AMP (cAMP) cAMP binds to CAP, protein changes shape CAP's helix-turn-helix motif binds to DNA near several promoters Promoters activates, genes transcribed Combination of Switches Sophisticated systems created by combining ON and OFF switches Example: lac operon of E. coli fig 16.13 Produces three proteins that import lactose and break it into glucose and galactose lac operon has two regulatory sites CAP site adjacent to lac promoter fig 16.14 Ensures genes not transcribed when glucose is present If glucose present, levels of cAMP are low CAP prevented from binding to DNA, lac promoter not activated If glucose absent CAP binds to DNA, promoter functional Operator is second regulatory site, adjacent to promoter fig 16.15 lac repressor binds to operator, only when glucose absent Repressor covers part of promoter when bound to operator RNA polymerase can't bind, lac genes not transcribed Cell doesn't transcribe genes to make product it doesn't need If lactose present, lactose isomer binds to repressor Repressor binding motif twisted away from major groove fig 16.16 Repressor can't bind to operator, RNA polymerase can bind topromoter, transcription of lac genes ensues fig 16.17 Lactose utilizing proteins made when lactose present, glucose not present TRANSCRIPTIONAL CONTROL IN EUKARYOTES Transcription Factors Assists binding of RNA polymerase to promotor fig 16.1 Assembles on promotor Guides and stabilizes binding of polymerase fig 16.18 Assembly begins 25 nucleotides upstream from start site Binds to short TATA sequence May then phosphorylate bound polymerase Several transcription factors provides numerous points for control Enhancers Composed of two distinct modules (domains) DNA-binding domain: attaches protein to DNA at specific site Regulatory domain: interacts with regulatory proteins Activators: accelerate transcription, speed capture of polymerase Repressors: interact with activator Compete with it for promotor binding site Complex with it to prevent its binding to transcription complex Modular design uncouples regulation from DNA binding Allows binding at one site and regulation at more distant site Enhancers are distant regulatory sites Occur occasionally in bacteria, regularly in eukaryotes fig 16.19 Mechanism of distant action fig 16.20 DNA loops around to position enhancer near promotor Regulatory domain brought into direct contact with transcription complex attached to promotor THE EFFECT OF CHROMOSOME STRUCTURE ON GENE REGULATION Histones Affect Gene Transcription Nucleosomes formed by wrapping DNA around histone proteins fig 11.15 Histones over promotors block assembly of transcription factor complexes Transcription factors unable to bind to nucleosome-packaged promotor Nucleosomes may prevent continuous transcription initiation Activators and RNA polymerase not inhibited by nucleosomes Regulatory domains of activators plus enhansers displace histones Displacement of histones required for assembly of complex fig 16.21 With transcription, RNA polymerase pushes histones aside Methylation Once Thought to Regulate Gene Transcription in Vertebrates Cytosine and uracil can be methylated, doesn't affect guanine or adenine Many inactive mammalian genes are methylated Once though to be cause of inactivation Now thought to simply block transcription of "turned-off" genes Ensures that once a gene is turned off, it stays off POST-TRANSCRIPTIONAL CONTROL IN EUKARYOTES Gene Transcription Can Be Regulated at Points After Transcription fig 16.22 All serve as control points for some eukaryotic genes mRNA sequences recognized by regulatory proteins, RNA molecules Processing of the Primary Transcript Exon-intron patchwork structure of eukaryotic genes Numerous exons are short, coding sequences Introns are lengthy, intervening noncoding sequences Introns removed by enzymes during RNA processing fig 16.23 RNA splicing fig 16.24 Exons can be spliced together in many ways to control expression Allows for production of various polypeptides from single gene Such alternative splicing common in vertebrates and insects Gene expression regulated by changing splicing event during development or in different tissues Transport of the Processed Transcript Out of the Nucleus Processed mRNA transcripts transported out through nuclear pores Active process requires recognition by pore receptors Poly-A tail at 3' end plays a role in this recognition Transport doesn't occur if any splicing enzymes are still attached Ensures partially processed transcripts are not transported Little evidence of regulation at this point 10% of transcribed genes are exons, 5% reaches cytoplasm Only half of primary transcripts leave nucleus Unknown whether this is under selective control Selecting Which mRNAs Are Translated Translation of processed mRNA transcripts Involves complex of proteins called translation factors Gene expression regulated by modification of these factors Translation repressor proteins shut down translation Bind to beginning of transcript, prevent attachment to ribosome Example: ferritin shut off by aconitase repressor protein Aconitase binds to 30 nucleotide sequence of ferritin mRNA Forms stable loop to which ribosomes cannot bind Presence of iron causes aconitase to dissociate Increases ferritin production 100-fold Selectively Degrading mRNA Transcripts Most eukaryotic mRNA transcripts are very stable Regulatory protein and growth factor transcripts are less stable Instability due to specific sequences at 3' end Sequences make them targets for mRNA degrading enzymes Examples Sequence of A and U nucleotides near 3' poly-A tail Promotes removal of tail Destabilizes mRNA Sequences that are endonuclease recognition sites, cause transcripts to be digested quickly Regulatory transcript instability facilitates rapid alteration of level