Kuliah 8.

Transcription
Dwi Suryanto
Program Studi Biologi
Universitas Sumatera Utara
dwisuryanto@usu.ac.id
Untuk kalangan sendiri dan diambil dari banyak sumber WA: 082165131177
Transcription is the process of making an RNA copy of a
gene’s DNA sequence. This copy, called messenger RNA
(mRNA), carries the gene’s protein information encoded in
DNA. In humans and other complex organisms, mRNA
moves from the cell nucleus to the cell cytoplasm (watery
interior), where it is used for synthesizing the encoded
protein.
Transcription generates three kinds
of RNA:
• Messenger RNA (mRNA) bears the
message for protein synthesis.
In Bacteria and Archaea, the
mRNA often bears coding A Polycistronic Bacterial Messenger RNA
information transcribed from
adjacent genes. Therefore it is
said to be polygenic or
polycistronic.
• Transfer RNA (tRNA) carries
amino acids during protein
synthesis.
• Ribosomal RNA (rRNA) molecules
are components of ribosomes.
 Perbedaan Transkripsi dan Translasi pada
Sel Prokaryota dan Eukaryota
• In prokaryotes (organisms without a nuclear
membrane), DNA undergoes replication and transcription
and RNA undergoes translation in an undivided
compartment.
• All three processes can occur simultaneously.
• In eukaryotes (organisms with a nuclear
membrane), DNA undergoes replication and transcription in
the nucleus, and proteins are made in the cytoplasm.
• RNA must therefore travel across the nuclear
membrane before it undergoes translation. This means
that transcription and translation are physically
separated.
• The primary transcript, heterogeneous nuclear
RNA (hnRNA), undergoes extensive post-transcriptional
processing to make a messenger RNA (mRNA) molecule
that can pass through the nuclear membrane.
Perbedaan transkripsi dan translasi pada sel Prokaryota dan Eukaryoya
 Prokaryote Transcription
• Bacterial transcription is the process in which a segment of
bacterial DNA is copied into a newly synthesized strand of
messenger RNA with use of the enzyme RNA polymerase.
• The process occurs in three main steps: initiation,
elongation, and termination; and the end result is a strand of
mRNA that is complementary to a single strand of DNA.
• Generally, the transcribed region accounts for more than one
gene.
• In fact, many prokaryotic genes occur in operons, which
are a series of genes that work together to code for the
same protein or gene product and are controlled by a
single promoter.
• Bacterial RNA polymerase is made up of four subunits and
when a fifth subunit attaches, called the σ-factor, the
polymerase can recognize specific binding sequences in the
DNA, called promoters.
• In prokaryotes, the promoter consists of two short
sequences at -10 and -35 positions upstream from the
transcription start site.
• The sequence at -10 is called the Pribnow box, or the -10
element, and usually consists of the six nucleotides
TATAAT.
Pribnow box (TATAAT)
• Daerah 5’ dari sekuen promotor disebut sekuen hulu upstream).
• Daerah 3’ dari sekuen promotor disebut sekuen hilir
(downstream).

Struktur gene Prokaryota

a. Initiation of transcription
• Initiation of transcription requires promoter regions, which
are specific nucleotide consensus sequences that tell the σ-
factor on RNA polymerase where to bind to the DNA.
• The promoters are usually located 15 to 19 bases apart and
are most commonly found upstream of the genes they
control.
• RNA polymerase is made up of 4 subunits, which include
two alphas, a beta, and a beta prime (α, α, β, and β’).
• A fifth subunit, sigma (called the σ-factor), is only present
during initiation and detaches prior to elongation.
Fungsi masing-masing sub-unit RNA polimerase
• Each subunit plays a
role in the initiation of
transcription, and the σ-
factor must be present
for initiation to occur.
• When all σ-factor is
present, RNA polymerase
is in its active form and
is referred to as the
holoenzyme.
• When the σ-factor detaches, it is in core polymerase form.
• The σ-factor recognizes promoter sequences at -35 and -10
regions and transcription begins at the start site (+1). The
sequence of the -10 region is TATAAT and the sequence of
the -35 region is TTGACA.
• Bagian DNA promotor yang terbuka (untwisted) disebut
open promoter complex
• Bagian DNA yang ditranskripsi ke dalam molekul RNA
disebut unit transkripsi.
Initiation of transcription
b. Elongation of transcription
• The transcription elongation phase begins with the release
of the σ subunit from the polymerase.
• The dissociation of σ allows the core RNA polymerase
enzyme to proceed along the DNA template, synthesizing
mRNA in the 5′ to 3′ direction at a rate of approximately 40
nucleotides per second.
• Basically, elongation is the stage when the RNA strand gets
longer, thanks to the addition of new nucleotides.
• During elongation, RNA polymerase "walks" along one
strand of DNA, known as the template strand, in the 3' to 5'
direction.
• RNA polymerase moves along to transcribe the DNA sequence
into a single strand RNA of the coding gene.
• When transcribing, the RNA polymerase interact with DNA
sequence forming transcription bubble.
• DNA double helix is reformed as the RNA polymerase moves
forward.
Elongation of transcription in bubble
b. Termination of transcription
• Transcription
termination in
prokaryotes can
be rho-independent
(intrinsic terminators
exist in the RNA
polymerase) and rho-
dependent, i.e., the
RNA polymerase
requires the cofactor
rho for termination
of transcription.
• Rho-independent termination is controlled by specific
sequences in the DNA template strand.
• As the polymerase nears the end of the gene being
transcribed, it encounters a region rich in C–G nucleotides.
• The mRNA folds back on itself, and the complementary C–G
nucleotides bind together.
• The result is a stable hairpin that causes the polymerase to
stall as soon as it begins to transcribe a region rich in A–T
nucleotides.
• The complementary U–A region of the mRNA transcript
forms only a weak interaction with the template DNA.
• This, coupled with the stalled polymerase, induces enough
instability for the core enzyme to break away and liberate the
new mRNA transcript.
• Upon termination, the process of transcription is complete.
• By the time termination occurs, the prokaryotic transcript would
already have been used to begin synthesis of numerous copies of
the encoded protein because these processes can occur
concurrently in the cytoplasm.
• The unification of transcription, translation, and even mRNA
degradation is possible because all of these processes occur in
the same 5′ to 3′ direction and because there is no membranous
compartmentalization in the prokaryotic cell. In contrast, the
presence of a nucleus in eukaryotic cells prevents simultaneous
transcription and translation.
• Sequences with the
DNA code consisting
of inverted repeats
upstream of
termination point.
• The hairpin
structure and a
stretch of at the end
play a role to stop
transcription.
• RNA polymerase moves
pass the inverted repeats
and transcribes the
termination sequence.
• Because of the inverted
repeat arrangement, RNA
synthesized forms a
hairpin loop structure.
• Hairpin loop makes the
RNA polymerase slow
down and eventually
stops,
• RNA polymerase can not continue attached to DNA.
• RNA polymerase dissociate.
• Rho-dependent termination is controlled by the rho
protein, which tracks along behind the polymerase on the
growing mRNA chain.
• Near the end of the gene, the polymerase encounters a run
of G nucleotides on the DNA template and it stalls. As a
result, the rho protein collides with the polymerase. The
interaction with rho releases the mRNA from the
transcription bubble.
• Terminator is a
sequence rich in
C (C-rich) and
poor G (G-poor).
• Terminator does
not form a
hairpin loop.
• Rho binds to C-
rich sequence
upstream of the
termination site
• Rho, which is a
helicase, moves to the
location of the RNA
polymerase.
• Rho unwinds the
DNA/RNA hybrid and
the RNA transcript is
release.
• RNA polymerase and
Rho dissociate.
 Eukaryote Transcription
Eukaryotic transcription is the elaborate process
that eukaryotic cells use to copy genetic information stored
in DNA into units of transportable complementary RNA replica.
Unlike prokaryotic RNA polymerase that initiates the
transcription of all different types of RNA, RNA polymerase in
eukaryotes (including humans) comes in three variations, each
translating a different type of gene.
 Organization of a eukaryotic protein-coding gene region

The transcriptional unit comprises the expressed exons and intervening introns, as well as 5' non-coding
sequences required for RNA polymerase binding (Shine-Delgarno sequence) and trailing 3' non-coding
sequences. Outside the transcriptional area, the 5' flanking region ("upstream") gene region includes the 'CAT
box' and 'TATA box' promoters required for RNA polymerase recognition prior to transcription. Enhancers that
regulate occurrence, timing, and amount of transcription occur in both the upstream region and the 3' flanking
region ("downstream") region; multiple enhancers may occur many hundreds of nucleotides upstream (Figure
& text © 2018 by Steven M Carr)
A eukaryotic cell has a nucleus that separates the processes of
transcription and translation. Eukaryotic transcription occurs
within the nucleus where DNA is packaged
into nucleosomes and higher order chromatin structures. The
complexity of the eukaryotic genome necessitates a great
variety and complexity of gene expression control.
By Erinp.5000 - Own work, CC BY-SA 4.0, https://commons.wikimedia.org/w/index.php?curid=79167319
To start transcription, general transcription factors, such as
TFIID, TFIIH, and others, must first bind to the TATA box and
recruit RNA polymerase to that location. The binding of
additional regulatory transcription factors to cis-acting
elements will either increase or prevent transcription. In
addition to promoter sequences, enhancer regions help
augment transcription. Enhancers can be upstream,
downstream, within a gene itself, or on other chromosomes.
Transcription factors bind to enhancer regions to increase or
prevent transcription.
Promoter sequences define the direction of transcription and
indicate which DNA strand will be transcribed; this strand is
known as the sense strand. Many eukaryotic genes have a
conserved promoter sequence called the TATA box, located 25
to 35 base pairs upstream of the transcription start site.
Following the formation of the preinitiation complex, the
polymerase is released from the other transcription factors,
and elongation is allowed to proceed as it does in prokaryotes
with the polymerase synthesizing pre-mRNA in the 5' to 3'
direction. As discussed previously, RNA polymerase II
transcribes the major share of eukaryotic genes, so this
section will focus on how this polymerase accomplishes
elongation and termination.
Although the enzymatic process of elongation is essentially
the same in eukaryotes and prokaryotes, the DNA template is
more complex. When eukaryotic cells are not dividing, their
genes exist as a diffuse mass of DNA and proteins called
chromatin. The DNA is tightly packaged around charged
histone proteins at repeated intervals. These DNA–histone
complexes, collectively called nucleosomes, are regularly
spaced and include 146 nucleotides of DNA wound around
eight histones like thread around a spool.
For polynucleotide synthesis to occur, the transcription
machinery needs to move histones out of the way every time
it encounters a nucleosome. This is accomplished by a special
protein complex called FACT, which stands for “facilitates
chromatin transcription.” This complex pulls histones away
from the DNA template as the polymerase moves along it.
Once the pre-mRNA is synthesized, the FACT complex
replaces the histones to recreate the nucleosomes.
Elongation of transcription in eukaryote
https://www.pngitem.com/middle/mwiwJh_elongation-of-transcription-in-eukaryotes-hd-png-download/
References
https://bio.libretexts.org/Bookshelves/Introductory_and_General_Biology/Book%3A_General_Biolog
y_(OpenStax)/3%3A_Genetics/15%3A_Genes_and_Proteins/15.3%3A_Eukaryotic_Transcription
H Liang et al., “Fast evolution of core promoters in primate genomes,” Molecular Biology and
Evolution 25 (2008): 1239–44.
R. E. Halbeisen, A. Galgano, T. Scherrer & A. P. Gerber. Post-transcriptional gene regulation: From
genome-wide studies to principles. Cellular and Molecular Life Sciences volume 65,
Article number: 798 (2008)
 Diskusi
“ Jika engkau punya satu apel dan aku punya satu apel lalu kita
pertukarkan maka kita masing-masing hanya punya satu apel, jika
engkau punya satu ide dan aku punya satu ide lalu kita pertukarkan
maka kita masing-masing punya dua ide” (George B. Shaw)
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