Eukaryotic Transcription
In all species, transcription begins with the binding of the RNA polymerase
complex (or holoenzyme) to a special DNA sequence at the beginning of the gene
known as the promoter. Activation of the RNA polymerase complex enables
transcription initiation, and this is followed by elongation of the transcript. In
turn, transcript elongation leads to clearing of the promoter, and the transcription
process can begin yet again. Transcription can thus be regulated at two levels: the
promoter level and the polymerase level. These elements differ among bacteria
and eukaryotes.





Process of creating an equivalent RNA copy of sequences of DNA.
DNA strand is read by RNA polymerase and synthesize complementary,
antiparallel strand.
Transcription is different than in case of prokaryotes (Complex).
RNA polymerase in bacteria is less complex than RNA polymerase in eukaryotes.
Some of the increased complexity of RNA polymerase in eukaryotes reflects
differences between DNA in eukaryotes and DNA in bacteria. Eukaryotes
organize their DNA into nucleosomes and have more complex mechanisms for
regulation of gene transcription. In order for transcription to occur, DNA must be
released from being tightly coiled in nucleosomes in case of eukaryotes. Another
complication of eukaryotic gene expression regulation is that gene sequences
controlling transcription are often distant from the DNA site where transcription
starts.
RNA polymerase in eukaryotes (including humans) comes in three variations,
each encoding a different type of gene.
RNA polymerase I is responsible for transcribing RNA that codes for genes that
become structural components of the ribosome, a protein responsible for the
translation of RNA into proteins.
2. RNA polymerase II transcribes protein-encoding genes, or messenger RNAs,
which are the RNAs that get translated into proteins.
3. RNA polymerase III transcribes a different structural region of the ribosome,
transfer RNAs, which are also involved the translation process, as well as nonprotein encoding RNAs.
1.

The promoter regions for RNA polymerases I and II are located upstream of the
start site, but the promoter for polymerase III is oddly located downstream.

One key difference between prokaryotic and eukaryotic transcription is that
eukaryotic polymerases are unable to recognize promoter regions. They have no
direct parallel to the sigma subunit of their prokaryotic counterpart. Instead,
eukaryotic polymerases depend on other proteins that bind to the promoter
regions and then recruit the RNA polymerases to the correct spots.
Pre-initiation:








Before initiation, initiation factors require promoter sequences on DNA
strand.
Promoter regions are present -30, -75 and -90 bp upstream.
Core promoter sequences (Sequences within promoters).
RNA polymerases bind at core promoter regions.
Most of the TATA box present at -30 bp.
TATA binding proteins (TBP), TATA is a binding site for transcriptional
factors.
Pre-initiation complex is composed of activators + repressors + transcriptional
factors + RNA polymerases + core promoter sequences + DNA helicase.
Upstream control elements (UCFs)
Initiation:



Attachment of polymerase enzyme (core enzyme).
RNAP (RNA polymerase) does not directly recognize promoter site rather
transcription factors mediate.
Initiation complex consists of transcriptional factors + RNAP.
Promoter Clearance:




After first bond is synthesized RNAP must release promoter.
Abortive initiation (tendency to release the RNA transcript & product
truncated transcript, it continues untill -factor rearranges, resulting in the
transcription elongation complex (35 bp moving front).
-factor releases before 80 nucleotides of mRNA synthesized.
ATP dependent process.
Elongation:






One strand of the DNA, the template strand (or noncoding strand), is used as a
template for RNA synthesis.
As transcription proceeds, RNA polymerase traverses the template strand and
uses base pairing complementarity with the DNA template to create an RNA
copy.
RNA polymerase traverses the template strand from 3' → 5', the coding (nontemplate) strand and newly-formed RNA can also be used as reference points, so
transcription can be described as occurring 5' → 3'.
This produces an RNA molecule from 5' → 3', an exact copy of the coding strand
(except that thymines are replaced with uracils, and the nucleotides are composed
of a ribose (5-carbon) sugar where DNA has deoxyribose (one less oxygen atom)
in its sugar-phosphate backbone).
Unlike DNA replication, mRNA transcription can involve multiple RNA
polymerases on a single DNA template and multiple rounds of transcription
(amplification of particular mRNA), so many mRNA molecules can be rapidly
produced from a single copy of a gene.
Elongation also involves a proofreading mechanism that can replace incorrectly
incorporated bases. In eukaryotes, this may correspond with short pauses during
transcription that allow appropriate RNA editing factors to bind. These pauses
may be intrinsic to the RNA polymerase or due to chromatin structure.
Termination:
Eukaryotic protein genes contain a poly-A signal located downstream of the last exon.
This signal is used to add a series of adenylate residues during RNA processing.
Transcription often terminates at 0.5 - 2 kb downstream of the poly-A signal, but the
mechanism is unclear.
Two termination mechanisms are well known:
Intrinsic termination (Rho-independent transcription termination)
This type of termination involves terminator sequences within the RNA that
signal the RNA polymerase to stop. The terminator sequence is usually a palindromic
sequence that forms a stem-loop hairpin structure that leads to the dissociation of the
RNAP from the DNA template.
Rho-dependent termination:
This type of termination uses a termination factor called rho (ρ-factor) factor
which is a protein to stop RNA synthesis at specific sites. This protein binds at a rho
utilization site on the nascent RNA strand and runs along the mRNA towards the RNAP.
A stem loop structure upstream of the terminator region pauses the RNAP, when ρ-factor
reaches the RNAP; it causes RNAP to dissociate from the DNA, terminating
transcription.
The role of regulatory transcription factors
In early 1990s, when the mystery of transcriptional regulation in prokaryotes have
been largely unveiled, scientists still knew very little about the regulation mechanism
in eukaryotes. The breakthrough came in 1996 when a number of research groups
discovered that certain transcriptional coactivators are histone acetyltransferases
(HATs). It has been known for some time that binding of transcriptional activators to
the enhancer region, in most cases, is not sufficient to stimulate transcription. Certain
co-activators are also required. Similarly, transcriptional repression often requires both
repressor binding on the silencer element and the participation of co-repressor
proteins. The precise role of these co-activators and co-repressors was not clear until
1996.
In eukaryotes, the association between DNA and histones prevents access of the
polymerase and general transcription factors to the promoter. Histone acetylation
catalyzed by HATs can relieve the binding between DNA and histones
Binding of activators to the enhancer element recruits HATs to relieve association
between histones and DNA, thereby enhancing transcription.
Binding of repressors to the silencer element recruits histone deacetylases (denoted by
HDs or HDACs) to tighten association between histones and DNA.