POST TRANSCRIPTIONAL MODOFICATIONS

1. Dr. P. Suganya
Assistant Professor
Sri Kaliswari College (Autonomous), SIvakasi

2.  Prokaryotes: RNA transcribed from DNA template and
used immediately in protein synthesis
 Eukaryotes: Primary transcript (hn RNA) must
undergo certain modifications to produce mature
mRNA (active form) for protein synthesis.

3.  “Post-transcriptional modification is a set of biological
processes common to most eukaryotic cells by which an
primary RNA transcript is chemically altered following
transcription from a gene to produce a mature, functional
RNA molecule that can then leave the nucleus and perform
any of a variety of different functions in the cell.”

4. OVERVIEW
Transcription
 Genetic information from DNA is copied into
messenger RNA (mRNA).
 In this process, mRNA is synthesized from the 5’ end
to the 3’ end.
 The initial transcript is known as heterogeneous
nuclear RNA (hnRNA) or pre-mRNA.

6.  Gene expression from DNA, the genetic sequence, is
transcribed into the RNA (transcription):
Transcription of genetic information is the first step in
gene expression and is the process through which a
coding region of DNA (double-stranded structure) is
used as a template for the synthesis of messenger RNA
(mRNA). The mature mRNA is translated into amino
acids, forming proteins (translation) with the help of
ribosomal RNA and transfer RNA (tRNA). This image
shows transcription without post-transcriptional
modifications of the RNA.

7. Modifications
 Primary transcripts, or immediate products of
transcription, undergo alterations to become biologically
functional.
 mRNA:
 Prokaryotes: Most primary mRNAs have no modifications.
 Eukaryotes: Synthesized transcript of mRNA (or hnRNA)
undergoes processing before leaving the nucleus.
 Addition of 5’ cap
 Addition of 3’ poly-A tail
 Splicing

8.  Modification of hnRNA produces mature mRNA, which is
transported to the cytoplasm through nuclear pores.
 In some cases, RNA editing occurs with base changes,
creating a sequence different from that copied from the DNA.
 A different mRNA sequence produces a different protein; this
varies from the old hypothesis of “one gene–one
polypeptide.”
 Transfer RNA (tRNA) and ribosomal RNA (rRNA):
 Structural molecules that are not translated
 Both have pre-tRNAs and pre-rRNAs that undergo processing.

10.  Summary of post-transcriptional modifications of
hnRNA into a mature mRNA:
The addition of the 5’ cap and the 3’ poly-A tail and
splicing (removal of the intervening sequences or
introns)

11. Addition of the 5′ Cap and 3′ Poly-
A Tail
5′ cap
 7-Methylguanosine (methylated guanylyl residue) is added
to the 5’ end of hnRNA via:
 Removal of the leading phosphate group at the 5’ terminal
by RNA triphosphatase
 Transfer of guanosine monophosphate (GMP) from the
guanosine triphosphate group by guanylyl transferase
 Methylation of guanine by guanine-7-methyltransferase
(methyl group from S-adenosylmethionine (SAM))
 Functions:
 Prevents exonuclease degradation
 Recognition sequence for translation

12. 3′ Poly-A tail
 50 to 250 adenylyl residues (AMP) are added to the 3’
end of hnRNA via:
 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
 Function:
 Prevents degradation in the cytosol by 3′
exoribonucleases
 Stabilizes mRNA

13. Post-transcriptional modifications of RNA:
The 5’ cap (7-methylguanosine) and 3’ poly-A tail
modifications prevent degradation of the mRNA in the
cytosol.

14. Heterogeneous Nuclear RNA
Splicing
 Exons and introns
 Heterogeneous nuclear (pre-mRNA) contains:
 Coding sections called exons (expressed sequences)
 Noncoding sections called introns (intervening
sequences)
 Processing:
 hnRNA needs processing (splicing) to produce the
mRNA carrying the proper coding sequences.
 Occurs in most eukaryotic genes, most commonly on
mRNA

15. Pre-mRNA exons and introns with an overview of splicing (from top to bottom):
Pre-mRNA transcript contains exons and introns. Interactions of the transcript with
small nuclear ribonucleoproteins and other proteins form a spliceosome at certain
junctions of the transcript. Cuts are made at the splice sites, and the intron is
released. Spliced RNA now only has exons, which contain the coding sequence.

16. Splicing
 Removal of introns from the hnRNA/pre-mRNA, while
linking the exons to form the mature mRNA
 Process involves the hnRNA and additional
components:
 Small nuclear ribonucleoproteins (which are made up of
small nuclear RNAs (snRNAs) and proteins)
 Other binding proteins

17.  Junctions where splicing reaction occurs:Splice sites:
 Areas where cuts are made between the exon and intron
 Base sequences identify these sites, one at the 5’ side
(beginning of the intron) and the other at the 3’ side
(end of the intron).
 5’ site/donor splice site: invariant GU
 3’ site/acceptor splice site: invariant AG
 Branch site: located upstream from 3’ site

18. Mechanism
 Small nuclear ribonucleoproteins recognize the splice sites
and branch site owing to the base sequences on the hnRNA.
 hnRNA, small nuclear ribonucleoproteins, and other proteins
combine to form the spliceosome.
 The spliceosome complex makes a cut on the 5’ donor splice
site (occurs via a nucleophilic attack by an adenylyl residue in
the branch site).
 The now free 5’ terminus of the intron links to the branch
site, forming a loop, or lariat, structure.
 The 3’ splice site is recognized, and the second cut occurs
there. Release of the lariat follows, and the 2 exons are joined
to form the mature RNA
 Occurs simultaneously with the 5’ cap and 3’ poly-A tail
hnRNA modifications

19. Alternative splicing
 Differential splicing of one hnRNA sequence
 Mechanisms:
 Exons are selectively included or excluded.
 Alternative 5′ donor or 3′ acceptor sites are used.
 Polyadenylation sites can differ.

20.  Up to 95% of multi-exon genes undergo alternative splicing
(AS) to encode proteins with different cellular functions.
 AS is a rapidly responsive regulation step needed for fine-
tuning protein synthesis and thereby determining cell
phenotypes and proliferation rates.
 Approximately 15% of hereditary diseases and cancers are
reported to be associated with AS.
 Different combinations of exons can lead to different
related proteins being created from the same hnRNA:
 Immunoglobulin molecules (genes for heavy chains have
exons related to individual subtypes)
 Tropomyosin variants in muscle
 Dopamine receptors in the brain (D2 receptors with 2
isoforms)

21. Examples of alternative splicing:
Protein A: Exons 1–5 were joined after
splicing of introns.
Proteins B and C: An exon was selectively
excluded to form a different protein

22.  References
 https://en.wikipedia.org/wiki/Post-
transcriptional_modification#:~:text=Post%2Dtranscri
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