CONCEPT OF GENE

EXPRESSION 1:
TRANSCRIPTION
ACHIEVERS ACADEMY
 INTRODUCTION
• Gene expression is the process by which information from a gene is used in the
synthesis of a functional gene product. Gene expression is the process by which the
instructions in our DNA are converted into a functional product, such as a protein.
• Gene expression is a tightly regulated process that allows a cell to respond to its changing
environment. It acts as both an on/off switch to control when proteins are made and also
a volume control that increases or decreases the number of proteins made.
• NB: There are two stages of Gene expression: Transcription & Translation. In essence, we
understand that enzymes and hormones drive metabolic processes. Where does the info
about the synthesis of these molecules come from?? The Genes!! This info follows two
stages in order to completely synthesize a functional product.
• In the first stage called Transcription, the information (nucleotide sequence) is converted
from DNA to RNA. Transcription is necessary because the language of the ribosomes
(where protein synthesis occurs) is understood in RNA codes.
 TRANSCRIPTION
• DNA transcription is the process by which the genetic information
contained within DNA is re-written into messenger RNA (mRNA)
by RNA polymerase.
• NB: Transcription takes place in the nucleus. It begins when
the cell wants a gene (segment of a DNA) to be expressed. It
uses DNA as a template to make an RNA (mRNA) molecule.
• This mRNA then exits the nucleus, where it acts as the basis for the
translation of DNA. By controlling the production of mRNA within
the nucleus, the cell regulates the rate of gene expression.
 TERMINOLOGIES USED IN TRANSCRIPTION
• 1. Promoter: A promoter is a region of DNA upstream of a gene
where relevant proteins (such as RNA polymerase and transcription factors)
bind to initiate transcription of that gene
• NB: Promoters are located near the transcription start sites of
genes, upstream on the DNA (towards the 5' region of the
sense strand). Promoters can be about 100–1000 base pairs long.
• 2. Transcription bubble: A transcription bubble is a molecular
structure formed during DNA transcription when a limited portion of
the DNA double helix is unwound.
• NB: A transcription bubble is formed when the RNA
polymerase enzyme binds to a promoter and causes two DNA
strands to detach.
• 3. Coding & Template strand: The template strand can be described as that
particular polynucleotide strand that the RNA polymerase uses to build the
new mRNA transcript. It reads from the 3’-5’ direction.
• NB: The template strand is also called the anti-sense or non-coding strand.
The cell uses a non-coding/antisense DNA sequence as a template to
produce mRNA
• NB: During transcription, the RNA pol binds to the promoter sequence on
the template strand, reads it in the 3’-5’ direction & synthesize the mRNA
transcript in the 5’-3’ direction.
• NB: the coding strand is also called the non-template (sense) strand. This
strand is similar to the mRNA transcript. It is also the DNA strand
whose base sequence is identical to the base sequence of
the RNA transcript produced (although with thymine replaced
by uracil).
• 4. Transcriptional factors: Transcription factor (TF) is a protein that
controls the rate of transcription of genetic information from DNA
messenger RNA, by binding to a specific DNA sequence.
• NB: In eukaryotes, transcriptional factors works with specialised
proteins to alter the rate of transcription. The include: co-
activators (to upregulate the rate of transcription) & compressors
(to down-regulate the rate of transcription).
• NB: In eukaryotes, an important class of transcription factors
called general transcription factors (GTFs) are necessary for
transcription to occur.
• The most common GTFs are TFIIA, TFIIB, TFIID (also TATA
binding protein), TFIIE, TFIIF, and TFIIH.
• 5. RNA POLYMERASE: This is an enzyme that synthesises RNA from a DNA
template. It is also called DNA-dependent RNA polymerase.

• NB: Bacterial RNA polymerase is a Holoenzyme, and consists of the core

enzyme & sigma factor. The core enzyme has five subunits designated α, α,
β´, β, and ω. The sigma factor, particularly (sigma-70) acts as a
transcription factor.
• NB: All eukaryotes have three different RNA polymerases (RNAPs) which
transcribe different types of genes. RNA polymerase I transcribes rRNA
genes, RNA polymerase II transcribes mRNA, miRNA, snRNA, and snoRNA
genes, and RNA polymerase III transcribes tRNA and 5S rRNA genes
• ENHANCER: This is a short region of DNA that can be bound
by proteins (activators) to increase the likelihood that transcription of
a particular gene will occur. Without enhancers, eukaryotic
transcription will be a slow process due to the length of the genome;
hence RNA polymerase will not locate the promoter on time
 MECHANISM OF TRANSCRIPTION
• DNA Transcription is divided into 3 stages: Initiation, Elongation & Termination.
INITIATION
• IN PROKARYOTES: To begin transcribing a gene, RNA polymerase holoenzyme
uses its sigma factor to bind to the DNA, which ultimately slides to the
promoter sequence which is located upstream.
• Once bound to the promoter sequence, RNA polymerase unwinds a
portion of the DNA double helix, exposing the bases on each of the two
DNA strands, thus creating what is known as a transcription bubble.
• NB: There is a conserved promoter sequence in bacteria called -35 and
-10 region. The -10 sequence is referred to as the Pribnow box. It
interacts with the sigma factor of RNA polymerase. The -35 sequence -
TTGACA is essential for the unwinding of DNA
• IN EUKARYOTES: Unlike the prokaryotic RNA polymerase that can bind to a
DNA template on its own, eukaryotes require several other proteins, called
transcription factors, to first bind to the promoter region and then help
recruit the appropriate polymerase.
• NB: The completed assembly of transcription factors and RNA polymerase
bind to the promoter, forming a transcription pre-initiation complex (PIC).
The consensus promoter in eukaryotes is the TATA box
• NB: The first TF to interact with promoter is the TFIID. This makes use of the
TATA-binding protein (TBP) portion to interact with the promoter. Once TFIID
is bound to promoter via the TBP, other transcription factors can then
interact to facilitate the unwinding of the double-stranded DNA.
• NB: The complete interactions on the promoter (TATA box) consisting of
transcription factors and RNA polymerase forms the pre-initiation complex
 ELONGATION
• NB: In transcription only one strand of DNA [called template strand or non-
coding strand] takes part. The template strand reads in the 3’-5’ direction.
• During elongation, RNA polymerase attaches to the template strand, reads
it in the 3’-5’ direction and synthesizes the mRNA transcript in the 5’-3’
direction.
• RNA polymerase makes use of ribonucleotides (ATP, GTP, CTP & UTP) to
elongate the chain
 TERMINATION
• IN PROKARYOTES: There are two mechanisms of transcriptional termination.
They include: Rho-dependent & Rho-independent termination .
RHO-DEPENDENT TERMINATION
• In the “Rho-dependent” type of termination, a protein factor called “Rho” is
used to stop RNA synthesis at specific sites. This protein binds at a Rho
utilisation site on the nascent RNA strand and runs along the mRNA towards
the RNA polymerase.
• When rho reaches the RNAP, it causes RNAP to dissociate from the DNA,
terminating transcription. In other words, it destabilizes the interaction
between the template and the mRNA, thus releasing the newly synthesized
mRNA from the elongation complex.
• NB: In preparation for post transcriptional modification the mRNA in
eukaryotic cells is called pre-mRNA.
 RHO-INDEPENDENT TERMINATION
• It is also known as intrinsic transcription termination. It 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.
• In the Rho-independent transcription termination, RNA transcription stops
when the newly synthesized RNA molecule forms a G-C rich hairpin loop,
followed by a run of U’s, which makes it detached the DNA template.
 POST-TRANSCRIPTIONAL MODIFICATION
• RNA is transcribed, but must be processed into a mature
form before translation can begin. Post-transcriptional
modifications (PTMs) are processes that facilitate the
generation of mature, functional RNA
• NB: Notable examples of PTM includes: Splicing, Alternate
splicing, Capping, Tailing & RNA editing
• 1. SPLICING: The is the process by which introns (the noncoding regions)
of genes, are excised out of the primary messenger RNA transcript, and
the exons (i.e., coding regions) are joined together to generate mature
messenger RNA
• For those eukaryotic genes that contain introns, splicing is usually
needed to create an mRNA molecule that can be translated into
protein. Splicing occurs in a series of reactions which are catalyzed
by the spliceosome.
• NB: Introns are sequences of DNA that don’t code for proteins.
They are also called Intragenic regions. In recent times, introns
have better been described as regulatory sequences within the
DNA.
• 2. ALTERNATIVE SPLICING: Alternative splicing is a cellular process in
which exons from the same gene are joined in different combinations,
leading to different, but related, mRNA transcripts
• NB: In essence, a single gene can contain numerous exons and introns, and
the exons can be spliced together in different ways. For example, if a gene
contains 10 exons, one version of the mRNA transcribed from that gene
might contain exons 1-9. Another version of the mRNA might contain
exons 1-8, and exon 10.
• NB: The different forms of the mRNA produced from alternate splicing are
called transcript variants or splice variants, thewhthe proteins made from
them are called isoforms.
• NB:
• 3: CAPPING: Capping is the first modification made to RNA polymerase II-
transcribed RNA and takes place co-transcriptionally in the nucleus. The 5′
m7G cap is an evolutionarily conserved modification of eukaryotic mRNA.
• NB: Three enzymatic activities are required to generate the cap structure,
namely, RNA triphosphatase (TPase), RNA guanylyltransferase (GTase)
and guanine-N7 methyltransferase (guanine-N7 MTase)
• NB: In capping, 7-Methylguanosine is added to the 5’ end of
hnRNA via: STEP 1: Removal of the leading phosphate
group at the 5’ terminal by RNA triphosphatase
• STEP 2: Transfer of guanosine monophosphate (GMP) from
the guanosine triphosphate group by guanylyl transferase
• STEP 3: Methylation of guanine by guanine-7-
methyltransferase (methyl group from S-
adenosylmethionine (SAM)
 SIGNIFICANCE OF CAPPING
• Capping acts as the anchor for the recruitment of initiation factors that
initiate protein synthesis. In essence, the 5’ capping indicates the initiation
site for protein synthesis
• It is also crucial to prevent mRNA degradation by bacteria or other
extrinsic factors.
• 4. TAILING: This process is also called Polyadenylation.
Polyadenylation is the addition of a poly(A) tail to an RNA
transcript, typically a messenger RNA (mRNA). The poly(A) tail
consists of multiple adenosine monophosphates.
• In this process, 50 to 250 adenylyl residues (AMP) are added to
the 3’ end of hnRNA. The process involves Cleaving of about
20 nucleotides downstream from an AAUAA recognition
sequence; Addition (and extension up to 250 nucleotides)
of poly-A tail (generated from ATP) by poly-A polymerase.
• SIGNIFICANCE OF POLYADENYLATION
• The poly-A tail makes the RNA molecule more stable and prevents its
degradation. Additionally, the poly-A tail allows the mature messenger
RNA molecule to be exported from the nucleus and translated into a
protein by ribosomes in the cytoplasm.
• 5. RNA EDITING:
RNA editing is a post-transcriptional process in which nucleotide
changes are introduced into a RNA sequence, many of which can thus
contribute to protein sequence variation.
• NB: There are basically two classes of RNA editing : Substitution
Editing & Insertion/Deletion editing.
• A. SUBSTITUTION EDITING: This involves the chemical alteration of
individual nucleotides (the equivalent of point mutations). These alterations
are catalyzed by enzymes that recognize a specific target sequence of
nucleotides.
• NB: A good example of substitution editing is the cytidine
deaminases that convert a C in the RNA to uracil (U).
• NB: There are two generic classes of RNA editing in nuclei, involving
enzymatic deamination of either C-to-U or A-to-I nucleotides.
• NB: In the Intestine
• In the cells of the intestine, an additional step of pre-mRNA processing
occurs: the chemical modification of the C nucleotide in Codon 2153
(CAA) into a U.

• This RNA editing changes the codon from one encoding the amino acid
glutamine (Gln) to a STOP codon (UAA). The modification is catalyzed by
the enzyme cytidine deaminase that recognizes the sequence of the RNA
at that one place in the molecule and catalyzes the deamination of C thus
forming U.

• Translation of the mRNA stops at codon #2153 forming apolipoprotein B-

48 — a protein containing 2152 amino acids — that aids in the
absorption of dietary lipids from the contents of the intestine.