Chapter 10 Outline 10 2 Transcription is the synthesis of an RNA molecule from a DNA template Transcription is a highly selective process o Genes are transcribed only as their products are needed Three major components of transcription 1 DNA template 2 Substrates needed to build a new RNA molecule 3 Transcription apparatus Transcription Template Christmas tree like structures o Trunk DNA removed by deoyribonuclease o Branches RNA removed by ribonuclease o At the top of the tree little DNA is transcribed into RNA but as the transcription apparatus moved down the DNA more DNA is transcribed Template one strand of DNA RNA molecule is complementary and antiparallel to the DNA template and has the same base sequence as replication would except U instead of T Transcription unit stretch of DNA that encodes an RNA molecule Moves downstream toward the terminator 1 Promoter DNA sequence the transcription apparatus recognizes and binds to Indicates which of the two strands to be read as a template and the direction of transcription Determines start site which nucleotide will be transcribed first Located NEXT to the start site but is not transcribed itself upstream 2 RNA coding sequence 3 Terminator Sequence of DNA nucleotides copied into a RNA molecule Sequence of nucleotides that signals where transcription should end Usually part of the coding sequence Substrate for Transcription RNA synthesized from ribonucleoside triphospahtes Nucleotides are added one at a time to the 3 OH group Two phosphates are cleaved allowing the remaining two to create a phosphodiester bond Synthesized in 5 to 3 direction Does NOT require a primer Transcription Apparatus RNA Polymerase carries out all required steps of transcription Requires an array of other proteins o Core enzyme allows transcription to occur Catalyzes elongation of RNA molecule o Sigma factor Controls binding of RNA polymerase to the promoter Tells WHERE transcription should start o Core enzyme Sigma factor Holoenzyme Once formed and bonded to promoter transcription starts Eukaryotic RNA Polymerases RNA Poly I o Transcribes rRNA RNA Poly II o Transcribed pre mRNA RNA Poly III Process of Bacterial Transcription Initiation o Transcirbes tRNA small rRNA miRNA o Promoter Recognition formation of transcription bubble creation of first bonds between rNTPs and escape of transcription apparatus from promoter o Consensus nucleotides short stretches of common nucleotides Implies that the sequence is associated with an important function 10 consensus sequence Prinbows box TATAAT box 35 consensus sequence TTGACA o Holoenzyme binds to consensus sequences and binds loosely at first then tightens bond unwinding the DNA Elongation downstream o Polymerase escapes from promoter and begins transcribing Termination o Takes place within the transcription bubble about 18 nucleotides o RNA polymerase unwinds the DNA downstream joins nucleotides to RNA molecule and rewinds DNA upstream o RNA polymerase must stop synthesizing RNA molecule must be released from RNA polymerase and RNA must dissociate from DNA o Rho dependent terminator only able to terminate if rho factor is o Rho independent terminator able to terminate even if rho factor is o Polycistronic RNA when a group of genes is transcribed into a single present absent RNA molecule 10 3 many genes have complex structures Gene Organization Not all genes are collinear with the proteins they encode Noncoding regions of DNA are seen in loops Introns noncoding regions o Removed by splicing o Common in Eukaryotes but not in bacteria o Tend to be longer than exons 10 4 Many RNA molecules are modified after transcription in Eukaryotes Messenger RNA processing Carries genetic information from DNA to a ribosome and helps assemble amino acids in their correct order In Bacteria transcribed directly from DNA In Eukaryotes premRNA is first transcribed and then processed to form mRNA Structure of mRNA Contains three regions 1 5 untranslated region Does not encode any amino acids of a protein In bacteria contains a consensus sequence called Shine Dalgarno serves as ribosome binding site Found 7 nucleotides upstream the start codon 2 Protein coding region Comprises the codons that specify the amino acids Begins with start codon and end with stop codon 3 3 untranslated region Not translated Affects stability and translation of mRNA 5 cap In bacteria transcription and translation occur at the same time so there is not much time for modifications o 3 undergoes transcription while 5 undergoes translation In eukaryotes separate o Transcription takes place in the nucleus o Translation takes place in the cytoplasm o Altered after transcription The addition of the 5 cap Consists of a modified nucleotide and several methyl groups Functions in the initiation of translation facilitates the binding of the ribosome increases stability of mRNA and might affect the removal of introns The addition of the poly A tail Addition of 50 250 adenine nucleotides at the 3 polyadenylation Added AFTER transcription not encoded in the DNA Many Eukaryotic genes are transcribed beyond the end of the coding sequence so the extra material is removed and the poly a tail is added The consensus sequence AAUAAA usually 11 30 nucleotides upstream from cleavage site determines where cleavage will take place U rich sequence downstream from cleavage site and help find the cleavage site Facilitates attachment of the ribosome to the mRNA RNA splicing Takes place in the nucleus Requires three sequences 5 splice site Possess short consensus sequences Possess short consensus sequences Most introns begin with GU and end with AG 3 splice site Branch point Adenine nucleotide that lies 18 to 40 nucleotides upstream Deletion or mutation of adenine nucleotide at branch point the 3 splice site prevents splicing Spliceosome one of the largest and most complex of all molecular complexes carries out the splicing reactions o Consists of 5 RNA molecules and 300 proteins o SnRNAs associate with proteins to form snRNPs o Each snRNP contains a single snRNA molecule and multiple o Spliceosome consists of 5 snRNPs names for the snRNA they contain U1 U2 U4 U5 U6 Pre mRNA is spliced in two steps o 1 Pre mRNA is cut at the 5 splice site which frees exon 1 from proteins the intron 5 of intron attaches to branch point and forms a lariat G nucleotide in consensus sequence at 5 bonds with A nucleotide at branch point and results in the 5 phosphate group of the G attached to the 2 OH group of
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