17.5 Eukaryotic Transcription Initiation Is Regulated by Transcription Factors That Bind to Cis-Acting Sites

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1 17.5 Eukaryotic Transcription Initiation Is Regulated by Transcription Factors That Bind to Cis-Acting Sites 1

2 Section 17.5 Transcription regulatory proteins, transcription factors, target cis-acting sites of genes regulating expression Activators increase transcription initiation Repressors decrease transcription initiation The effects regulatory proteins can be finely turned to the appropriate cell type in response to environmental cues or during development The human metallothionein IIA gene (hmtiia) provides an example of how a gene can be transcriptionally regulated through the interplay of multiple promoter and enhancer elements and the transcription factors that bind to them 2

3 The presence of multiple regulatory elements and transcription factors allow the hmtiia gene to be transcriptionally activated or repressed in response to subtle changes in both extracellular and intracellular conditions 3

4 Section 17.5 Proteins that serve as transcription factors have two functional domains (clusters of amino acids with a specific function) A DNA-binding domain Binds to specific DNA sequences in the cis-acting regulatory site A trans-activating domain Activates or represses transcription by binding to other transcription factors or RNA polymerase Characteristic domains of DNA-binding proteins include helix-turn-helix (HTH) zinc finger basic leucine zipper (bzip) 4

5 17.6 Activators and Repressors Interact with General Transcription Factors at the Promoter 5

6 Section 17.6 Basal (general) transcription factors are required for the binding of RNA polymerase II to the promoter Pre-initiation complex (PIC) TFIID, the first general transcription factor to bind the promoter, binds to the TATA box through the TATA binding protein (TBP) 6

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8 Section 17.6 Transcription activators and repressors may increase or decrease the rate of transcription initiation in several ways Activators/repressors may bind to chromatin near the promoter and open regions of promoter for interaction with transcription machinery, or produce repressive structure and inhibit transcription initiation Activators/repressors may bind directly to transcription factors to enhance or inhibit initiation Activators bind to enhancers and form the enhanceosome, which interacts with the transcription complex Repressor proteins bind at silencer DNA elements to repress transcription 8

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10 17.7 Gene Regulation in a Model Organism: Transcription of the GAL Genes of Yeast 10

11 Section 17.7 The GAL system in yeast is made up of four structural genes and three regulatory genes The products of the structural genes transport galactose into the cell for metabolism The products of the regulatory genes positively /negatively control transcription of structural genes The GAL genes are inducible (transcribed) by the presence of galactose, but only if the concentration of glucose is low This indicates that the GAL genes are also subject to catabolite repression 11

12 Section 17.7 The GAL1 and GAL10 genes are controlled by a central control region, UAS G, that contains four binding sites for the Gal4 protein (Gal4p) UAS are functionally similar to enhancers in eukaryotes The chromatin structure of a UAS is constitutively open, or DNase hypersensitive, meaning that it is free of nucleosomes 12

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14 Section 17.7 A mutation in the regulator of the GAL genes, GAL4, prevents activation, indicating that transcription is under positive control the regulator must be present to turn on gene transcription Gal4p is negatively regulated by Gal80p, which covers the Gal4p activation domain Binding of phosphorylated galactose to Gal80p and/or Gal4p exposes the activation domain of Gal4p 14

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16 17.8 Posttranscriptional Gene Regulation Occurs at All the Steps from RNA Processing to Protein Modification 16

17 Section 17.8 Although transcriptional control is perhaps the major type of regulation in eukaryotes, posttranscriptional regulation plays an equal if not more significant role This includes: removal of introns and splicing together of exons addition of a cap and poly-a tail translation stability 17

18 Section 17.8 Alternative splicing can generate different forms of mrna from identical pre-mrna, giving rise to a number of proteins from one gene As a result, the number of proteins that a cell can make (its proteome) is not directly related to the number of genes in the genome Approximately two-thirds of human genes undergo alternative splicing Humans produce several hundred thousand different proteins from approximately 25,000 genes in the haploid genome Mutations that affect regulation of splicing (spliceopathies) contribute to several genetic disorders Myotonic dystrophy, fragile-x, Huntington 18

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20 Alternative splicing of the Dscam gene mrna 20

21 Section 17.8 The Sex lethal (Sxl), transformer (tra), and doublesex (dsx) genes are part of a hierarchy of gene regulation for sex determination in Drosophila The Sxl gene acts as a switch that selects the pathway of sexual development by controlling splicing of the dsx transcript in a female-specific fashion 21

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23 Section 17.8 The steady-state level of an mrna is its amount in the cell. This is determined by a combination of the transcription rate and rate of mrna degradation This determines the amount of mrna available for translation and is regulated in response to cellular needs The quantity and activity of a gene product can be regulated in many ways Translation can be regulated to produce the correct quantity of a protein Posttranslational stability of the protein can be modulated: P53 protein A protein can be modified after translation to change its structure and hence its activity 23

24 17.9 RNA Silencing Controls Gene Expression in Several Ways 24

25 Section 17.9 Short RNA molecules regulate gene expression in the cytoplasm of plants, animals, and fungi by repressing translation and triggering mrna degradation This form of sequence-specific posttranscriptional regulation is known as RNA interference (RNAi) Together, these phenomena are known as RNA-induced gene silencing RNA interference uses a protein called Dicer to cleave doublestranded RNA molecules into small interfering RNAs (sirnas) and micro RNAs (mirnas) sirnas and mirnas repress mrna translation and trigger degradation Inhibit transcription of specific genes by associating with RNAinduced initiation of transcription (RITS) and RNA-induced silencing complex (RISC) 25

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27 Section 17.9 RNAi technology has been studied in the lab, and RNAi is being developed as a pharmaceutical agent Any disease caused by overexpression of a specific gene or normal expression of an abnormal gene product could be attacked by therapeutic RNAi Scientists have had promising results to reduce the severity of HIV, influenza, polio Animal models have shown successful RNAi treatment of viral infection eye diseases cancers inflammatory bowel disease RNAi holds powerful promise for molecular medicine, with the expectation of the first RNAi drugs to be available within the next decade 27