RNA Transcription Ch 13 and 14 BSCI222 Lecture 4 9 12 13 o U instead of T hydroxyl group on the 2 Carbon of the sugar whereas DNA has a Hydrogen there RNA is more reactive than DNA o RNA can take on many shapes because the single strands folds up on itself to satisfy its Hydrogen requirements o DNA is thought to have become the genetic material through evolution because it s much more stable than RNA because RNA is single stranded and its chemical structure o DNA s secondary structure is usually the Beta helix RNA s primary structure is linear secondary is folding with base pairing and some twisting stems and loops Many kinds of RNA o Of course mRNA rRNA ribosomal structural and functional components of the ribosome and tRNA but also many tiny ones that we re still learning their functions RNA is transcribed from the DNA NOT translated transcription is a copy of base pairs translation is turning it into protein language instead of nucleic acid language RNA is always synthesized 5 to 3 too and antiparallel to the template strand Always report DNA 5 to 3 But that s not the strand that is transcribed into RNA RNA is transcribed from the template strand the complimentary strand 3 to 5 RNA is made from the bottom strand the 3 to 5 therefore the RNA is made 5 to 3 Either strand of the DNA could encode the gene o Promoter upstream RNA coding region Terminator downstream stop the RNA from transcribing so the polymerase doesn t go all the way around to the end of the chromosome and make an enormous molecule Then there d be no control over the expression of genes they d all be transcribed RNA transcription synthesis o Helix has to be opened up to give us access to the bases and Hydrogen bonds in order to specify which nucleotides should be synthesized o Don t need a primer RNA polymerase can start making RNA on a single nucleotide strand o Transcription bubble moves along the helix closes up behind the polymerase o PROKARYOTIC To know a sequence is functional have to find sequences that are always found upstream or downstream of a gene Consensus sequences compare the actual sequences find common patterns almost all have a T first consensus sequence starts with T Equal proportion of T or C both pyrimidines next letter is Y N no pattern Typically find a consensus sequence 10 sequences before the transcription start site called the minus ten box TATAAT and 35 sequences before called the minus thirty five box TTGACA If every gene had exactly the same sequences upstream no control over gene expression Variation closer to consensus sequence or farther away from it it is perfect and the strongest sequence for a protein to bind to fitting the protein when binding perfectly will lower or raise protein binding strength of gene expression A subunit of the RNA polymerase binds to the minus ten box and minus thirty five box called the Sigma factor the other subunits bind anywhere no specificity The Sigma factor is what gives the polymerase specificity so it only binds to promoters and increases the rate of transcription by helping it bind more tightly Essentially the Sigma factor controls which genes will be expressed Core RNA polymerase Sigma bind to promoter creating a closed complex of six different proteins Holds all those subunits tightly to the promoter The holoenzyme RNA polymerase binds to the promoter and unwinds the helix open promoter complex beginning the process o RNA polymerase has to be stopped once it gets to the end of the gene Two mechanisms Rho protein independent mechanism A particular sequence at the end of the gene of repeats of A U which folds up into a hairpin loop causing the RNA polymerase to pause clogs it up Stall the polymerase and then the base pairs that are A U only 2 hydrogen bonds instead of 3 are weak big chance that thermal motion will cause the hairpin loop to completely fall off Rho protein dependent Still a stem loop that makes RNA polymerase stall but then Rho protein binds to the transcriptor and starts chasing the polymerase catches up when it pauses then has a gyrase activity that separates that transcript from the helix Always key to slow down the polymerase long enough for the transcript to be disassociated o DNA being transcribed into RNA looks like a Christmas tree gene start is at the top where the RNA strands are short Trunk is DNA As polymerase moves down the trunk RNA branches get longer more has been transcribed Little ornament balls on the end of the branches might be ribosomes meaning prokaryotic only place you can have simultaneous transcription and translation Each branch has individual polymerase o EUKARYOTES RNA polymerases are specialized Pol II is doing most of the messenger RNAs and some of the smaller ones Pol III does the tRNAs and some of the smaller ones Pol I only does the large ribosomal RNAs As eukaryotic genome got more complex had to figure out how to regulate transcription on a much larger scale Pol II is huge DNA goes through it RNA exits out as a single strand Consensus sequences not the same as prokaryotic 25 box called TATA box TATAAA 1 downstream of start site initiator element 30 box downstream core promoter element Prokaryotes specificity is more consensus sequences and binding strengths Eukaryotes have core promoter but also much larger regulatory promoter site upstream of the start site Will find recognizable promoter sites in different arrangements and different numbers in front of different genes Will affect how much and when the gene is expressed Their exact position doesn t matter You d think the binding site would have to be in an exact place but it s actually very flexible get essentially the same gene expression It s the Presence and number of these boxes that matters Prokaryotic transcription complex core sigma eukaryotes is much bigger Have TBP TATA binding protein bind to TATA box transcription factor 2D is recruited other proteins begin to assemble into the core transcription complex or basal transcriptional apparatus Giant conglomeration of proteins all interacting with each other to localize the polymerase to the exact spot that the transcription should start increasing its binding begin to open up the helix in that location so that the polymerase can have access to the template Core promoter plus proteins binding in the regulatory promoter A lot of interactions and H bonding between the amino acids in the protein and the DNA structure Can have sequences even further upstream called enhancers or silencers