22. Transcription Flashcards
Define DNA transcription
DNA transcription: mRNA synthesis from a DNA gene
- in interphase (S) - only in this phase chromosomes despiralise - can be read
Sequence of transcription + enzymes
- INITIATION: RNA polymerase binds to promoter sequence - unwinds DNA - NTPs (E) used for mRNA synthesis from template strand (antisense)
- ELONGATION: synthesis of mRNA in 5’ to 3’ end
- TERMINATION: gene synthesised until terminator sequence - RNA polymerase and mRNA detach from template strand (antisense)
RNA transcript is identical to sense strand (5’-3’) and complementary to antisense strand (3’-5’)
Sections of a gene
- Promoter: non-coding, initiates trasncription, binding site for RNA polymerase (controlled by transcription factors in euk.)
- Coding sequence: transcribed by RNA polymerase into mRNA
- Terminator: non-coding, stops transcription, RNA polymerase detaches
Sense strand vs antisense strand
SENSE strand: coding strand, not transcribed, identical to mRNA (T→U)
ANTISENSE strand: template strand, transcribed, complementary to mRNA
Post-transcriptional modifications
- Capping: addition of methyl group to 5’ end, protection against exonucleases, can be recognised by cell’s translation machinery
- Polyadenylation: addition of poly-A tail to 3’ end, improves stability, facilitates export out of nucleus
- Splicing: introns are removed - mature mRNA
- Alternative splicing: removal of some exons - different proteins from same gene
Exons vs introns
INTRONS - intruding sequence
EXONS - expressing sequence
What regulates gene expression?
TRANSCRIPTION FACTORS: from a complex with RNA polymerase - transcription cannot be started without - levels regulate expression
REGULATORY PROTEINS: bind to DNA sequences (control elements) and interact with transcription factors, enhancer (activator, increases rate of transcription) - silencer (repressor, decreases rate of transcription)
Types of control elements in transcription
Distal control elements: distant from promoter, regulatory proteins bind
Proximal control elements: close to promoter, transcription factors bind
Examples of environmental influences on gene expression
- Hydrangeas: pH of soil determines - pink/blue flowers
- Himalayan rabit produces different fur colour depending on the temperature
- Humans produce different amounts of melanin depending on light exposure
- Certain fish/reptiles change sex depending on social cues
Histone tail modifications and their influence on gene expression
ACTIVATION - DEACTIVATION of genes by nucleosome packing
- Acetylation: neutralises the + charge of histone tails - DNA less tightly coiled - increases transcription of those genes - EUCHROMATIN
- Methylation: maintains + charge of histone tails - DNA more coiled - transcription of those genes reduced - HETEROCHROMATIN
- Phosphorylation: phosphate group addition
DNA methylation
DNA methylation decreases gene expression - prevents transcription factor binding
Define epigenetics
EPIGENETICS: study of changes in phenotype due to variation in gene expression levels
Shows DNA methylation patterns - change over life
Methylation influenced by:
- heritability but not genetically pre-determined (identical twins different methylation patterns)
- environmental effects (diet, exercise, smoking)
Reverse transcription
In retroviruses - by reverse transcriptase - cDNA (complementary) transcribed from mRNA - cDNA represent sequences which are actively transcribed
- used to determined gene expression in healthy - diseased
- cDNA used in gene transfer (don’t possess introns)
Explain codons how many of them
- codons code for specific amino acids, three bases translated into one amino acid (reading frame)
- 64 codons in total: 61 code for the 20 possible amino acids, 3 STOP codons
- Several codons can code for the same amino acid, but one codon corresponds only to one amino acid
- AUG (Met) start codon
What are the three RNA polymerases and what to they synthesise?
Explain how transcription is initiated: promoters, transcription factors, RNA polymerase
- Promoter regions in DNA - upstream (earlier) the gene
- Transcription factors bind to the promoter regions
- RNA polymerase II can bind to the DNA strand when transcription factors indicates where - transcription initiation complex
- TATA box - DNA sequences crucial for forming transcription initiation complexes in eukaryotes
Explain how distant elements promote transcription during initiation stage
- Away from the gene region activator proteins bind to specific regions - act as enhancers in transcription
- Distant elements (activator proteins) come in contact with transcription via a mediator complex to which both of them bind → initiation of transcription (once activator proteins come in contact RNA polymerase II is released to transcribe)
Termination in eukaryotes vs prokaryotes
- in bacteria transcription ends at the terminator region - mRNA can be translated
- in eukaryotes polyadenylation sequence must be transcribed and termination happens when 10-35 more nucleotides are added post adenylation transcript in mRNA
What are the post transcriptional modifications of mRNA?
- when only transcribed - pre-mRNA - RNA processing modifies pre-mRNA in nucleus before sending off to cytoplam: both ends of pre-mRNA are changed: 5’ end 5’ cap, 3’ end poly-A tail
- Splicing occurs to form mature mRNA
What are the functions of modifying mRNA ends? Poly-A tail and 5’ cap
- facilitate transport out of the nucleus
- protect mRNA from hydrolytics enzymes
- help ribosomes attach to 5’ end
Structure of a mature mRNA ready for transcription
the start and stop codons in the sequence are for translation
Explain split genes and RNA splicing
What is the evidence for mRNA splicing?
What are the signals on mRNA for splicing?
Always in eukaryotes: AG-GU (A) AG-G signal splicing
What is the mechanism for splicing?
- occurs in splicosomes
- 2 nucleophilic attacks to get rid of an intron in a lariet structure (the loop) - small nuclear ribonuclear proteins (snRNP) form the splicosome
What allows RNA to function as an enzyme?
- can form 3D structures with itself due to base pairing
- some RNA abses functional groups that may act in catalysis
- RNA can form hydrogen bonds with other nucelic acid molecules
- Examples: snRNAs, ribozymes are catalytic RNAS
What are self splicing RNA introns?
- tetrahymena, some mitochondrial and chloroplast genes contain these introns
- don’t require additional molecules
- rely on extensive base pairing with intron sequence
- 2 possible groups of self splicing
- 2 group mechanism resembles mammalian splicing in splicosomes (A performs nucleophilic attack, repeated nucleophilic attack on another part in intron - lariet structure)
- splicosomal splicing may have originated from group 2 self splicing (same mechanism)
What are the uses of introns in transcription?
- regulate gene expression
- alternative RNA splicing may result in different mRNAS from the same DNA sequence (depends which sections are treated as exons)
