Lecture 10 - Chapter 6 PDF

RNA Synthesis … aka Transcription Sections 6.1 – 6.2 (and a little 7.2) Transcription Learning Objectives Compare and contrast DNA replication (Chapter 5) and transcription (Chapter 6) Explain how E. coli RNA polymerase initiates transcription Diagram a bacterial promoter sequence....

RNA Synthesis … aka Transcription Sections 6.1 – 6.2 (and a little 7.2) Transcription Learning Objectives Compare and contrast DNA replication (Chapter 5) and transcription (Chapter 6) Explain how E. coli RNA polymerase initiates transcription Diagram a bacterial promoter sequence. Describe the processes of transcriptional elongation and termination. Summarize the roles of different eukaryotic RNA polymerases. Distinguish between the binding of bacterial and eukaryotic RNA polymerases to promoters. Describe the functions of the general transcription factors for RNA polymerase II. From 7.2: Compare and contrast promoters and enhancers. From 7.2: Describe how activators and repressors affect transcription. Transcription Key Concepts Transcription begins at a promoter sequence located upstream of the gene sequence and ends at a terminator sequence located downstream of the gene sequence on the DNA template strand. Transcription occurs in 3 stages: initiation, elongation, and termination. Prokaryotic initiation involves a sigma factor and RNApol holoenzyme. Eukaryotic initiation involves general transcription factors, RNApol II, and a mediator complex. Additional gene specific transcription factors that bind to enhancer sequences in the DNA may also be required. Gene specific transcription factors may activate or repress gene transcription. Replication vs. Transcription DNA Replication makes a _______ copy of the ____________… During the process of transcription, only _________ of DNA sequence are transcribed into _______ BOTH DNA and RNA are synthesized _________ Coding Strand vs. Template Strand Transcription produces RNA that is ________________ to one strand of DNA Non-template/coding strand Template strand Both DNA strands contain gene sequences The DNA strand that serves as the template strand is different for different genes. Practice… Shown below is a section of double stranded DNA. The template strand for three genes are found at this particular location (blue, orange, purple). Recall that RNA is synthesized from 5’ to 3’ complementary and antiparallel to the DNA template. Indicate the locations of the promoter for each gene (blue, orange, purple) Above or below each gene box, draw the resulting mRNA including 5’ and 3’ directionality and an arrow pointing in the direction that the mRNA was synthesized. 3’ 5’ 5’ 3’ RNA polymerase carries out transcription RNA Polymerase – Synthesizes single stranded ________ using __________ as a template. – New RNA strand synthesized ______________ E. coli RNA polymerase Sequences in DNA tell RNA pol where to start and stop transcription Note: this is not the same as a start codon or stop codon E. coli promoters contain ____ and ____ ___________ sequences NOTE CHECK The promoter sequence is transcribed first, followed by the DNA of the gene. a. True b. False NOTE CHECK Join.nearpod.com pin = Bacterial transcription stops at the ______________ sequence NOTE CHECK E. coli RNA polymerase differs from E. coli DNA polymerase in that A. it reads the template DNA strand in the 3′ to 5′ direction. B. it does not need a primer sequence to initiate reading. C. the double-stranded DNA gene does not need to be unwound for reading. NOTE CHECK Join.nearpod.com pin = Eukaryotic RNA Polymerases Recognize different promoters Transcribe different classes of genes All consist of 12-17 different subunits 9 conserved subunits 5 are related to bacterial subunits Quick Review Every somatic cell in a multicellular organism contains the same set of DNA. a. True b. False Every somatic cell in a multicellular organism contains the same set of mRNA transcripts. a. True b. False _____________________ are required for eukaryotic RNA pol 1979 – Robert Roeder discovered that RNA pol-II can only initiate transcription if additional proteins are present. General Transcription Factors (GTFs) Additional gene-specific transcription factors Eukaryotic promoters contain Promoters of different genes contain different several sequence elements combinations of core promoter elements. near transcription initiation sites. 5 general transcription factors are minimally required for transcription by RNA pol-II 1. Binding of TFIID (large protein complex) 2. Recruitment of TFIIB 3. Recruitment of RNA pol-II and TFIIF 4. Binding of TFIIE and TFIIH complete preinitiation complex – TFIIH contains a kinase subunit that phosphorylates the C-terminal domain of RNA pol-II RNA polymerase II/Mediator complexes and transcription initiation Large (>20 protein) complex called _____________ also required Plays a key role in linking general transcription factors to gene-specific transcription factors NOTE CHECK Which would not be true if a yeast cell could not make TFIIF? a. TFIID would not be recruited to the initiation complex. b. TFIIE would not be recruited to the initiation complex. c. TFIIH would not be recruited to the initiation complex. d. Transcription could not proceed. Join.nearpod.com pin = Chapter 7.2 What additional gene-specific transcription factors interact with Mediator for robust eukaryotic transcription? Eukaryotic Promoters …and Enhancers An individual gene may have multiple cell- or time-specific enhancers Image source: HHMI BioInteractive Enhancers are required for anything beyond “basal” transcription Distant regulatory elements are united by DNA looping and Mediator Gene-specific TFs can be ______________ or _______________ Mini-Case Study: Sickle Cell Anemia Image source: HHMI BioInteractive Mini-Case Study: Sickle Cell Anemia Fetal hemoglobin Adult hemoglobin Image source: Pritišanac et al. 2021 Image source: Protein Data Bank 101 Mini-Case Study: Sickle Cell Anemia BCL11A is a gene that is not expressed in fetuses, but is expressed in adults. When BCL11A function is removed from an adult mouse, fetal hemoglobin is now produced in the blood cells of the adult mouse. What type of protein does the BCL11A gene encode? a) A transcriptional activator of adult hemoglobin b) A transcriptional repressor of adult hemoglobin c) A transcriptional activator of fetal hemoglobin d) A transcriptional repressor of fetal hemoglobin Join.nearpod.com pin = Mini-Case Study: Sickle Cell Anemia Scientists have generated BCL11A-deficient hematopoietic stem cells (bone marrow!) that can be transplanted into mice, resulting in red blood cells (RBC) that lack BCL11A and produce fetal hemoglobin. Which of the following genomic editing strategies could NOT give this result? a) Deleting an adult-RBC-specific activator-binding sequence in an enhancer for BCL11A b) Deleting the genomic DNA coding sequence for BCL11A c) Deleting the genomic DNA coding sequence for LIN28B, a protein that represses BCL11A expression Join.nearpod.com pin =