Transcription in prokaryotes and eukaryotes

Transcription in prokaryotes and eukaryotes

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  1 Transcription in Prokaryotesand Eukaryotes Overview  Introduction  TranscriptioninProkaryotesandEukaryotes  Pre-Initiation  Initiation  Elongation  Termination  Post-Transcription Transcription DNA → RNA The formation or synthesis of single RNA from DNA is called Transcription. Four Stages are involved : 1. Pre-Initiation 2. Initiation 3. Elongation 4. Termination Introduction  Synthesis of single stranded RNA  5’→3’ direction  RNA Polymerase is used  No primers needed  Only a part of the genome is transcribed  First stage of gene expression and the principle conservation step. RNA Polymerase  Nuclear Polymerases
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  2  RNA PolI-ProducesrRNA(28S, 5.8S, 18S)  RNA PolII-Produces mRNA, snRNA, siRNA, miRNA  RNA PolIII-Produces tRNA, mRNA (5S), SRPRNA In Prokaryotes & Eukaryotes Prokaryotes  Occurs in cytoplasm  Coupled transcription & translation  No definite phase of occurrence  A single RNAP synthesizes mRNA, tRNA& rRNA  No initiation factors required  Polycistronic Eukaryotes  Occurs in nucleus  No coupling of transcription & translation  Occurs in the G1 & G2 phases  RNAP I, II & III synthesize rRNA, mRNA & tRNA  TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH recognize TATA box  Monocistronic Polycistronic& Monicistronic Monocistronic An mRNA molecule is said to be monocistronic if it contains genetic information to translate only a single protein. In the end only one polypeptide chain coding for one protein is obtained from a single gene with an operator & promoter region. Polycistronic PolycistronicmRNA contains information for several genes which are translated into several proteins.
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  3 mRNA contains severalORFs (open reading frames), each of which is translated in proteins. The coding is grouped and all of the genes are translated together with a common promoter & operator region (like in operons) Pre-Initiation  First step of transcription.  The Pre-Initiation Complex (PIC) includes RNA Polymerase II and six transcription factors-TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH  Other co-activators and chromatin remodeling complexes also comprise of PIC  TATA Binding Protein (TBP) is a subunit of TFIID and binds to the promoter, creating a sharp bend.  TBP-TFIIA interact; TBP-TFIIB interact; TFIIB-TFIIF interact & TFIIF recruits RNA PolII; TFIIE joins the group and recruits TFIIH  Subunits within TFIIH that haveATPaseandhelicaseactivity create negativesuperhelicaltension in the DNA. Initiation Negative superhelical tension causes approximately one turn of DNA tounwindand form thetranscription bubble. Promoter melting requires hydrolysis by ATP and is mediated by TFIIH. TFIIH pulls the double stranded DNA into the cleft of RNA Polymerase and helps in transition from closed to open state. The two strands get separated.
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  4 Abortive Initiation  Beforeentering elongation phase, The polymerase may terminate prematurely.  This produces a truncated polypeptide chain.  Many cycles of abortive initiation may occur before actually producing a growing polypeptide chain.  This helps in providing a scrunching kind of motion. Elongation  The polypeptide chain is elongated with the help of Elongation Factors.  RNA Polconveniently adds nucleotides to the 3’ end. The template strand for this is known as the sense strand and the other anti-sense strand.  There are different classes of elongation factors. Some factors can increase the overall rate of transcribing, some can help the polymerase through transient pausing sites, and some can assist the polymerase to transcribe through chromatin
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  5 Transcription Fidelity  RNApolymerases select correctnucleoside triphosphate(NTP) substrate to prevent transcription errors. Only the NTP which correctly base pairs with the coding base in the DNA is admitted to the active center.  RNA polymerase performs two known proof reading functions to detect and remove misincorporatednucleotides: pyrophosphorylyticediting and hydrolytic editing. Pausing and Backtracking  RNA polymerase does not transcribe through a gene at a constant pace. Rather it pauses periodically at certain sequences, sometimes for long periods of time before resuming transcription.  Promoter-proximal pausing during early elongation is a commonly used mechanism for regulating genes poised to be expressed rapidly or in a coordinated fashion. The blockage is released once the polymerase receives an activation signal. Pausing and Backtracking
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  6 Termination Two Types: 1. FactorDependent 2. Factor Independent o Factor dependent requires Termination Factors along with RNA PolI. o Factor Independent termination can be done by RNA PolIII. A stretch of Thyminesalong a hair pin loop causes disintegration of complexes. Post Transcriptional Modifications  5’ end Capping  A guanine nucleotide linked to the 5’ end triphosphate  Polyadenylation  Poly Adenine units added to 3’ end of the Ribonucleotidechain.