Chapter 28 Regulation of Gene Expression The cellular concentration of a protein is determined by a delicate balance of at least seven processes o Synthesis of the primary RNA transcript o Posttranscriptional modification of mRNA o Messenger RNA degradation o Protein synthesis translation o Posttranslational modification of proteins o Protein targeting and transport o Protein degradation 28 1 Principles of Gene Regulation Housekeeping genes Regulated gene expression genes for products that are required at all times and are expressed at a more or less constant level in virtually every cell of a species or organism Constitutive gene expression fall in response to molecular signals Inducible molecular circumstances gene products that increase in concentration under particular when cellular levels of gene products rise and unvarying expression of a gene o Induction the process of increasing the expression of these genes gene products that decrease in concentration in response to a Repressible molecular signal o Repression Transcription is regulated by protein DNA interactions the process of decreasing the expression of these genes RNA Polymerase Binds to DNA at Promoters The regulation of transcription initiation often entails changes in how RNA polymerase interacts with a promoter Differences in the promoter sequence may affect the frequency of transcription initiation o Mutations that result in a shift away from the consensus sequence usually decrease promoter function o Mutations towards consensus enhance promoter function The RNA polymerase promoter interaction strongly influences the rate of transcription initiation for housekeeping genes Transcription Initiation Is Regulated by Proteins That Bind to or near Promoters Three types of proteins regulate transcription initiation by RNA polymerase alter the specificity of RNA polymerase for a given o Specificity factors promoter or set of promoters o Repressors o Activators impede access of RNA polymerase to the promoter enhance the RNA polymerase promoter interaction The subunit of the E coli RNA polymerase holoenzyme is a specificity factor that mediates promoter recognition and binding o 70 is the most common o 32 is used during heat stress Repressors bind to specific sites on the DNA o Operators a promoter such binding sites in bacterial cells that are generally near o Blocks binding or movement of RNA polymerase o Negative regulation blocks transcription o Effector a molecular signal that regulates repressor binding regulation by means of a repressor protein that Usually a small molecule or protein that binds to the repressor causing a conformational change activators bind to DNA and enhance the activity of RNA Positive regulation polymerase at a promoter o Particularly common in eukaryotes Many Bacterial Genes Are Clustered and Regulated in Operons Many bacterial mRNAs are polycistronic meaning they have multiple genes on a single transcript and require one promoter which is the site for regulation of all the genes in the cluster o Operon that function together in regulation the gene cluster and promoter plus additional sequences The lac Operon Is Subject to Negative Regulation The lac operon contains genes for o galactosidase Z o Galactoside permease Y o Thiogalactoside transacetylase A In the absence of lactose the lac operon genes are repressed In higher eukaryotes almost all protein encoding genes are transcribed separately Regulatory Proteins Have Discrete DNA Binding Domains To bind specifically to DNA sequences regulatory proteins must recognize surface features on the DNA Most of the protein DNA contacts that impart specificity are hydrogen bonds DNA binding sites for regulatory proteins are often inverted repeats of a short DNA sequence Helix turn helix o Crucial to the interaction of many bacterial regulatory proteins with o Comprises about 20 amino acids in two short helical segments DNA separated by a turn One of the two helices is called the recognition helix because it usually contains many of the amino acids that interact with the DNA in a sequence specific way The recognition helix is positioned in or nearly in the major groove o The Lac repressor has this motif Zinc Finger o 30 amino acids form an elongated loop held together at the base by a Zn2 ion which stabilizes the motif o Weak interaction with DNA for a single Zn finger o Usually in eukaryotes Homeodomain o A domain of 60 amino acids that is seen as a DNA binding domain in transcriptional regulators Homeobox the DNA sequence that encodes this domain Regulatory Proteins Also Have Protein Protein Interaction Domains Leucine Zipper o An amphipathic helix with a series of hydrophobic amino acid residues concentrated on one side o The hydrophobic surface forms the area of contact between the two polypeptides of a dimer Basic Helix Loop Helix o A conserved region of about 50 amino acids that can form two short amphipathic helices linked by a loop of variable length o The helix loop helix motifs of two polypeptides interact to form dimers Subunit Mixing in Eukaryotic Regulatory Proteins o Three types of additional domains for protein protein interaction Glutamine rich domains Proline rich domains Acidic domains 28 2 Regulation of Gene Expression in Bacteria The lac Operon Undergoes Positive Regulation The availability of glucose also affects the expression of the lac genes Catabolite repression a regulatory mechanism that restricts expression of the genes required for catabolism of lactose arabinose and other sugars in the presence of glucose even when these secondary sugars are also present cAMP receptor protein CRP a homodimer with binding sites for DNA and cAMP o Mediates the effect of glucose in the lac operon o Has a helix turn helix motif in its DNA binding domain o A positive regulatory element responsive to glucose levels As cAMP declines CRP binding to DNA declines thereby decreasing the expression of the lac operon o In the presence of glucose synthesis of cAMP is inhibited o Strong induction of the lac operon therefore requires both lactose to inactivate the lac repressor and a lowered concentration of glucose a network of operons with a common regulator Regulon Many Genes for Amino Acid Biosynthetic Enzymes Are Regulated by Transcription Attenuation Genes for the enzymes needed to synthesize a given amino acid are generally clustered in an operon and are expressed whenever existing levels of that amino acid are inadequate o When the amino acid is
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