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Transcription (What is transcription?
One strand of DNA molecules is…
Transcription
What is transcription?
- One strand of DNA molecules is used as a template to yield RNA
- The gene is the region of the DNA that is transcribed
- RNA product may be mRNA, tRNA, snRNA or another type of RNA
What is a gene?
- Hereditary unit occupying a specific position within the genome.
- Information content of genes is in the form of specific sequences of nucleotides along strands of DNA
- DNA sequence that codes for a specific polypeptide chain
- Set of segments of nucleic acid that contains the information necessary to produce a functional RNA product in a controlled manner.
Transcription V DNA replicationSimilarities:
- Chemical mechanism
- Polarity - direction of synthesis - addition of nucleotides to 3' OH
- Use of a template
- Initiation, elongation and termination
Differences:
- Transcription uses RNA polymerase so does not require a primer
- Generally involves a limited section of the DNA
- Within a transcribed segment only one DNA strand serves as a template
Basic structure of a gene:
- Promoter is a base pair - sequence that specifies where transcription beings
- Coding sequence is a base pair. Coding information polypeptide chain is specified if this is a gene for mRNA
- Terminator is a sequence that specifies the end of RNA transcript.
- Flanking regions
- Starts 5' flanking region
- Proceeds to 3' flanking region
- Leader length is variable
- Trailer length may be 50-200 nucleotides. Some mRNA have non and others have 500
- 3' trailer aids stability. Long sequence of Us cause instability
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DNA - 2 strands, the coding strand and the template strand
- Template strand - what is used as a template in the synthesis on mRNA - also called antisense strand
- Coding strand - identical in sequence to the mRNA excep DNA all Us & Ts - also called sense strand
Promotoer sequence
- Stretch of DNA that lies upstream
- Required to direct the initiation of transcription by an RNA polymerase
- Critical in regulating when and how gene is turned on
-Varies between prokaryotes and eukaryotes and different genes
RNA polymerase
- Complex enzymes consisting of a number of subunits
- Bacteria have a single type of RNA polymerase that catalyses synthesis of mRNAs, rRNAs and tRNAs encoded in the bacterial chromosomes
3 stages of RNA transcript
- Initiation
- Elongation
- Termination
Eukaryotic Cells - RNA polymerase II binding and initiation of transcription
- Promoters signal the initiation of RNA synthesis
- Transcription factors mediate the binding of RNA polymerase and the initiation of transcription
- Complete assembly of transcription factors and RNA polymerase II bound to a promoter is called transcription initiation complex
- Promoter called a TATA box is crucial in forming the initiation complex in eukaryotes
Elongation of RNA
- RNA polymerase moves along the DNA
- Untwists the double helix
-10-20 bases at a time
- Transcription progresses at a rate of 40 nucleotides per seconds in eukaryotes
- Gene can be transcribed simultaneously by several RNA polymerases
- Elongation needs to be from 3' -OH
Termination
- Requires for the RNA polymerase enzyme to be released and also the RNA product to be released
- Several mechanisms of transcription termination have been discovered in both bacteria and eukaryotes
- Prokaryote there are 2 possible mechanisms - Rho dependant and Rho independent
- Eukaryotes - 3 RNA polymerases use different termination mechanisms
Prokaryote termination of transcription - Rho dependent
- 70-80 base segment of the growing RNA transcripts wraps around Rho.
- Rho is a hexametric protein
- This interaction activate an ATPase activity of Rho that is associated with its movement along the RNA in the 3'direction until it eventually unwinds the RNA-DNA hybrid at he active site of RNA polymerase
- Transcription is terminated or not depends on whether Rho moves sufficiently fast to catch up with the polymerase
Termination in eukaryotes
- RNA pol I genes transcription is stopped using a termination facto, through a mechanism similar to rho-dependent termination in bacteria
- Transcription of RNA pol III genes ends after transcribing a termination sequence that include a polyuracil stretch (er UUUUU) by a mechanis resembling rho-independent prokaryotic termination
Termination with RNA polymerase II
- Connected mechanism of transcription termination and RNA 3' end processing
requires protein complexes to bind to the specific sequences in the newly formed RNA
- Enzyme poly-A-polymerase then adds a poly A tail. This process follows cleavage and also coordinated with termination
- 5'exonulease will attack the 5' end of the RNA strand still associated with the RNA polymerase and will aid in displacing the RNA polymerase II
- Enzymes in the eukaryotic nucleus modify pre-mRNA before the genetic messages are dispatched to the cytoplasm
- During RNA processing both ends of the primary transcript are usually altered
- Usually some interior part of the molecule are cut out and the other parts spliced together
Alteration of mRNA Ends
- The 3ʹ end gets a poly-A tail
- The 5ʹ end receives a modified nucleotide 5ʹ cap
Modification share several function:
- Facilitate the export of mRNA
- Protect mRNA from hydrolytic enzymes
- Help ribosomes attach to 5' end
- Non coding regions are called intervening sequences or intron
- Exons are eventually expressed, usually translated into amino acid sequences
- RNA splicing removed intron and joins exons - this create an mRNA molecule with a continuous coding sequence
Spliceosomes consist of a variety of proteins and several small nuclear ribonuleoproteins (snRNPs) that recognise the splice sites