RNA synthesis Flashcards
What subunit directs the E.coli RNA polymerase to the correct transcription initiation sites?
sigma
transcription
conversion of DNA into RNA
difference of RNA
use ribose sugar
no thymine but uracil
single stranded but can fold
mRNA
messenger
template for protein synthesis
heterogeneous length - average
rRNA
ribosomal
major component of ribosome
tRNA
transfer
carries amino acid in activated form to ribosome
snRNA
small nuclear - in RNA splicing
miRNA
micro
around 21 bases
bind to mRNA promoting degrade/inhibit their translation
how RNA is produced
copying strand of DNA as template - direct transcription 5’ to 3’ direction
producing other side
coding strand - 5’ to 3’ has same sequence as the RNA
catalyst for RNA synthesis
enzyme RNA polymerase
using rNTPs, release pyrophosphate
nucleotides
attached to 3’ -OH from 5’ to 3’ direction
DNA Duplex
around 17 base pairs
unwound
length of RNA-DNA hybrid duplex
8bp long
contain RNA polymerase
alpha*2, 2beta and omega - core enzyme
sigma - direct enzyme to start at initiation site
omega
stabilises within 5 subunits
sigma
initiation factor
how transcriptional units starts
marked by promoters
holoenzyme
6 subunit 450kDa
RNA polyermase
Pribnow box
a consensus sequence found at -10 on the non-template strand in bacterial promoter
consensus sequence
TTGACA
at -35
optimal sequence
best promoter site for RNA polymerase
If Pribnow box had G or C in it
harder for RNA polymerase to open up the transcription bubble
direction of RNA synthesis
5’ to 3’
antiparallel
What is formed from RNA synthesis
Transient RNA/DNA duplex
RNA polymerase size (how many bp)
able to bind to around 30bp of DNA
termination signals
palindromic GC rich regions followed by AT rich region
what the new RNA transcript forms
stem and loop
hair pin structure
when RNA transcript forms
RNA polymerase is kicked off
protein factor p(pho)
signals for termination are in newly synthesised RNA rather than in DNA template
transcription and translation in prokaryotes
occurs at the same time
after synthesis in prokaryotes
some RNAs are modified
tRNA after synthesis in prokayotes
some bases and sugar are modified
CCA is added at the 3’ end
rRNA after synthesis in prokayotes
made as one long molecule - broken up into individual subunits by endo/exonucleases post transcriptionally
synthesis in Eukaryotes
more complex due to transcriptional regulation
has nuclear membrane - separate transcription and translation
3 types of RNA polymerase
RNA polymerase I
RNA polymerase II
RNA polymerase III
RNA polymerase I role
rRNA - 18s, 5.8s and 28s
RNA polymerase II role
mRNA
RNA polymerase III role
tRNA and 5s rRNA
eukaryotes pre-mRNA processing
addition of poly A tail to 3’ end of RNA transcript
addition of cap to 5’ end - 5’-5’ triphosphate linked to 7-methyl-G
increase in stability and specifies export to cytoplasm
acts as protector role
increase half life
interact binding protein
eukaryote Pre-mRNA
contains introns and exons
introns
non-coding section
exons
coding region
order of introns and exons
can not change
proteins produced
may contain some exons and not others
splicing site
coded by RNA sequence
spliceosomes
splices the pre-mRNA
importance of spliceosome
allows synthesis of different proteins from one gene - may be cell type specific
example fibronectin
component of extracellular matrix
produced by fibroblast containing exons EIIIB and EIIIA
liver cell in example for fibronectin
don’t encode the exons which allows fibronectin to be secreted and move freely in blood stream and help clot formation
beta-thalassemia number of causes
has 3 ways of this causation
beta-thalassemia first causation - change in nucleotide
single nucleotide change - destroying normal splice site - exonskipping
beta-thalassemia second causation - extension
activation cryptic splice site - extended exon
beta-thalassemia third causation - addition of new
causing of new exons to be incorporated - have extra exons
control of transcription
gene expression need to be controlled to form proteins appropriate for tissue and its environment
cause of enhancement by activators or blocked by repressors
the interaction of RNA polymerase and a promoter
operon
in bacteria
promoter controls genes in common pathways
lac operon - in bacteria
involved n metabolism of lactose
has lac Z, lac Y and lac A
how to stop enzyme production
repressor gene binds to operator sequence - prevents transcription
when is gene expressed
when RNA is transcribed into protein
constitutive expression
genes expressed all the time
regulated expression
genes expressed under certain conditions or time
when lactose is present
allolactose binds to repressor protein which prevents binding to operator
therefore genes switched on and protein produced
allolactose
inducer of operon
can other operons be switched off
yes by other compounds
example of other operons
trp operon
trp operon
trp repressor binds to operator
in presence of compressors - switch off synthesis
in high presence of tryptophan
regulatory gene
transcribed and translated to repressor protein or repressor inducer
repressor protein
stop transcription
repressor inducer
promote transcription
control in eukaryotes complexity
tissue specificity
cell type specificity
development regulation
transcription control
DNA packed - due to condensed chromatin
modifying histones in transcription control
causes relaxation of region - exposing promoter-proximal element and promoter
exposure of ppe and promoter
allows RNA polymerase to bind and transcribe sequence
enhancers
found further up stream
binds to different transcription factor - bind to regulatory sequence close to start site
DNA being able to fold back allows
formation of different forms to regulate gene expression of particular genes down stream
advantage of DNA folding back to form different loops
help provide tissue selection and cell type
Basal transcription factor
common in most genes
function of Basal transcription factor
bind to promoter
such as TATA and CCAAT box
Regulatory transcription factor
specific for particular gene - unique
cell type/ developmentally regulated
Regulatory transcription factor function
bind to ppe and enhancer
ppe
promoter-proximal element
miRNA function
regulate gene expression
characteristics of miRNA
small non-coding DNA - 21-25 nucleotide
how miRNA is synthesised
as a precursor from nuclear DNA and processed into miRNA in nucleus and cytoplasm
human genome encode for miRNA
> 1000 miRNA
function of miRNA
base pairing complementary sequences in mRNA
silencing specific mRNA and repressing protein translation
miRNA expression
in element close by such as introns
miRNA processing
- form pri-miRNA using RNA pol II forming stem loop structure
- processed by Drosha and Pasha - trim different components to form Pre-miRNA
miRNA processing out of nucleus
- using Dicer to form miRNA/ antisense miRNA duplex
which then binds to coding mRNA = rapid degradation
after processing
- interfering RNA induced in silencing complex
- targeting mRNA by relying on complementary base pairing
product of targeting mRNA
cleave off the target RNA
translation repression and then destruction of RNA
formation of heterochromatin on DNA (target RNA being transcribed)
