4 5 13 Lecture 29 Chapter 28 Transcription Factors Transcription Factors protein that binds to DNA sequences thereby controlling flow or transcription of genetic info from DNA to mRNA o protein needed for initiation of transcription but is not part of RNA polymerase Does this by either promoting as an activator or blocking as a repressor recruitment of RNA pol to specific genes The Problem with Eukaryotic Transcription o Prokaryotes o Eukaryotes DNA is found in the nucleoid Packaging is dependent on supercoiling and proteins that stabilize the structure DNA is highly compacted into chromatin in the nucleus Chromatin Functions packages DNA into smaller volume to fit in cell strengthens DNA to allow mitosis prevents DNA damage and controls gene expression and DNA replication DNA is wrapped around histones Makes it harder for transcription factors to access it Regulation of Gene Expression o Gene expression can be regulated at multiple levels 1 DNA accessibility through chromatin structure harder to access due to DNA being wrapped around proteins 2 Initiation and elongation of transcription 3 Transcription processing addition of 5 Cap and 3 Poly A tail 4 RNA transport to cytoplasm 5 Translational control 6 mRNA degradation and turnover Role of Transcription Factors o Transcription Factor protein needed for initiation of transcription but is not part of RNA polymerase Bacteria Eukaryotes Bacterial RNA polymerase binds to DNA and recognizes the promoters o RNA polymerase can read DNA Eukaryotic RNA polymerase cannot recognize promoters alone o Requires transcription factors to direct the polymerase RNA polymerase doesn t know where to go Eukaryotic Transcription Control o Contains a core promoter region and an enhancer region Enhancer located either upstream or downstream Binds the transcription factors o Recruits the basal cell machinery RNA polymerase Increases utilization of promoter Usually tissue or temporal specific Core Promoter contains binding side for the basal transcription complex RNA polymerase II 100 bp long once transcription factors recruit basal cell machinery they move along DNA until they find the core promoter o then transcription starts Architectural Alteration DNA o Regulation of transcription involves both activators and repressors o Proteins will bind to DNA and change the structure of the DNA to either activate transcription or repress it o Two groups that do the same thing but cause difference results in transcription Activators or Repressors Binds to DNA and either brings activator proteins together or moves them farther apart Either activates transcription or inhibits transcription Transcription Factor Mode Transcription factors either act on DNA directly or indirectly o Direct EX TFIID Transcription factors like Activators control frequency of transcription by interacting with basal transcription factors by protein protein interactions Bind directly and specifically to a DNA site located in or near a promoter and making protein protein interactions with general transcription machinery Activators have independent DNA Activators are proteins that increase transcription binding and activation domains Transcription factors like Coactivators bind to proteins that bind to the promoter or response elements thus indirectly affecting transcription Coactivator is a protein that increases gene expression by binding to an activator or transcription o Indirect factor which contains a DNA binding domain Does not bind to DNA by itself Functions Stabilizes complex enabling faster clearance of promoter Recruits other transcription factors Assists in basal apparatus RNA pol position Promotes CTD phosphorylation o CTD C terminal domain of RNA polymerase o enhances transcription initiation by stabilizing formation of RNA pol holoenzyme Activators Repressors o Activators increase the rate of transcription in two ways 1 Direct protein protein interaction with basal factors at promoter 2 Indirect interaction via coactivators o Repressors decrease the rate of transcription Binds to DNA modifies chromatin and inhibits transcription o RNA Polymerase Binding Steps Activator binds to enhancer changes DNA confirmation by bending DNA bringing enhancer promoter together then recruits basal cell machinery RNA polymerase holoenzyme then binds and begins transcription Yeast RNA polymerase 2 is typically associated with mediator complex Mediator complex functions as a coactivator binds to C terminal domain of RNA Pol 2 Typical Structure of Transcription Factors o Transcription Factors contain DNA binding Domain Binds the specific DNA sequence recognized by the trasncripton factor Activation Domain in repressor Repressor Domain binds other transcription factors may have Dimerization Domain o transcription factors can generally be activators or repressors o have modular construction composed of independent units that can be joined in any order to form a one larger unit Can vary the order or number of each domain They function independently o Modify or mutate the domains changes the action they take o Action Domain Action domain can recruit general transcription factors to the promoter STEPS Activator binds to the enhancer o Changes confirmation of DNA Recruits mediator aka coactivator which is bound to RNA Pol 2 o Also recruits transcription factor TFIID TATA box binding protein TBP bind to TFIID Mediator RNA Pol2 and TFIID TBP bind to TATA box Transcription begins DNA binding Domains o DNA binding proteins transcription factors have various types of DNA binding domains Helix loop Helix Homeodomain Proteins Zinc Finger Leucine Zipper Review Helix turn Helix consist of two alpha helices common in prokaryotes and eukaryotes consists of a Recognition Helix which binds into the major groove of DNA followed by a turn and then another alpha helix Ex LacI and Lamda repressors o 1 Helix loop Helix contains helices that are responsible for dimer formation consist of two alpha helices a short and a long joined by a loop one helix is smaller allowing dimerization by folding and packing against another helix each helix contains basic amino acids bHLH dimerization motif COOH these help facilitate DNA binding generally transcription factors with helix loop helix domain are dimeric Prefer heterodimer 2 different kinds of helix loop helix bind together o Increases diversity and number of sequences it can bind to Homodimers binding of similar units bind DNA poorly Regulation bind on the major groove of DNA