Part 1: Pre-mRNA Processing

Pre-mRNA processing is critical for producing mature mRNA capable of being translated into
proteins. Three main steps are involved: capping, polyadenylation, and splicing.

1. Capping the 5’ End: The 5' cap of pre-mRNA is added immediately after transcription
initiation, where a 7-methylguanosine molecule is attached. The basic machinery involved
includes the enzyme guanylyltransferase, which adds the guanine residue, and methyltransferase,
which methylates the guanine. The 5' cap plays a crucial role in protecting mRNA from
degradation, promoting ribosome binding, and facilitating nuclear export (Ahern et al., 2018).

Chemistry: The capping process involves forming a unique 5' to 5' triphosphate linkage between
the 7-methylguanosine cap and the RNA. This unusual linkage makes the mRNA more resistant
to exonucleases.

Aberrancies: Failure in the capping process could lead to mRNA instability, rapid degradation,
and a subsequent decrease in protein synthesis, affecting gene expression.

2. Polyadenylation of the 3’ End: The polyadenylation process involves two steps: cleavage of
the RNA at the polyadenylation site (5’-AAUAAA-3’) and the addition of around 200 adenine
residues. Polyadenylation factors like CPSF (cleavage and polyadenylation specificity factor)
and PAP (poly(A) polymerase) facilitate this process (Unit 4: RNA Processing, n.d.).

Chemistry: CPSF recognizes the polyadenylation signal, and PAP catalyzes the addition of
adenine molecules. The poly(A) tail aids in mRNA stability, nuclear export, and translation
initiation.

Aberrancies: Defects in polyadenylation can lead to improper mRNA export or translation,

potentially decreasing protein production. Mutations in polyadenylation sites can contribute to
diseases such as cancer due to altered mRNA stability (Unit 4: Gene Expression, n.d.).

3. Splicing: Splicing involves the removal of noncoding introns from pre-mRNA. The
spliceosome, a complex of small nuclear ribonucleoproteins (snRNPs), catalyzes this process.
Alternative splicing can generate different mRNA variants from a single gene, allowing for
protein diversity (Ahern et al., 2018).

Chemistry: The spliceosome recognizes intron-exon junctions, cleaves the introns, and ligates
the exons. This process involves phosphodiester bond cleavage and formation.

Aberrancies: Incorrect splicing can result in the inclusion of introns or exclusion of exons,
leading to nonfunctional or harmful proteins. Aberrant splicing is linked to diseases like spinal
muscular atrophy (Unit 4: RNA Processing, n.d.).
Part 2: Small Noncoding RNAs and mRNA Silencing

Small noncoding RNAs, like miRNAs and siRNAs, regulate gene expression post-
transcriptionally by inhibiting mRNA translation or promoting its degradation.

1. Nuclear Export and Conversion to dsRNA: Small hairpin RNAs are transported out of the
nucleus by exportin-5. Once in the cytoplasm, they are converted into double-stranded RNA
(dsRNA) by proteins like Drosha and Dicer (Unit 4: RNA Processing, n.d.).

Chemistry: This step involves cleavage by Dicer, an RNase III endoribonuclease, which
processes hairpin structures into 22-28 nucleotide-long dsRNAs (Ahern et al., 2018).

2. dsRNA Cleavage by Dicer: Dicer further processes dsRNA into miRNAs or siRNAs. It
cleaves the RNA into small fragments, ready for silencing activities (Unit 4: RNA Processing,
n.d.).

3. Separation of Strands: The dsRNA is unwound into a sense and antisense strand. The
antisense strand, which is complementary to target mRNA, is incorporated into the RNA-induced
silencing complex (RISC) (Unit 4: Gene Expression, n.d.).

Chemistry: ATP hydrolysis helps unwind the RNA strands.

4. RISC and Argonaute Action: RISC uses the antisense strand to locate complementary
mRNA. If there is perfect complementarity, the Argonaute protein, a catalytic subunit, cleaves
the mRNA, leading to degradation (Unit 4: Translation, n.d.).

Aberrancies: Aberrant miRNA levels can disrupt normal gene silencing, leading to diseases like
cancer. Overactive silencing could degrade essential mRNAs, while insufficient silencing may
allow oncogene expression (Ahern et al., 2018).

Part 3: Translation Process

Translation converts mature mRNA into a functional protein through initiation, elongation, and
termination steps.

1. Initiation: The ribosome assembles at the start codon (AUG), and an amino-acyl-tRNA binds
to the mRNA at the ribosome’s A-site. The initiation factors (IFs) help position the mRNA and
tRNA (Unit 4: Translation, n.d.).

Chemistry: GTP hydrolysis powers ribosomal assembly. The ribosome consists of two subunits,
40S and 60S, in eukaryotes.

Aberrancies: Defective initiation could result from mutations in the start codon or IFs, leading
to failure in protein synthesis. This can be linked to developmental disorders.
2. Elongation: The ribosome moves along the mRNA, with tRNAs bringing amino acids to the
growing polypeptide chain. The ribosomal P-site holds the peptidyl-tRNA, while the A-site
accepts new tRNAs (Unit 4: Translation, n.d.).

Chemistry: Peptide bond formation occurs between the amino group of the incoming amino
acid and the carboxyl group of the existing polypeptide.

Aberrancies: Mutations in tRNAs or elongation factors can slow translation, leading to protein
insufficiency. This is observed in diseases like muscular dystrophy.

3. Termination: When a stop codon is reached, release factors (RFs) bind to the ribosome,
prompting the release of the completed polypeptide (Ahern et al., 2018).

Chemistry: Hydrolysis of the bond between the polypeptide and tRNA facilitates release.

Aberrancies: Premature termination caused by nonsense mutations leads to truncated proteins,

which are often nonfunctional or harmful, contributing to genetic diseases (Unit 4: Translation,
n.d.).

References

Ahern, K., Rajagopal, I., & Tan, T. (2018). Biochemistry free for all (version 1.3). Oregon State
University. https://biochem.oregonstate.edu/content/biochemistry-free-and-easy
Unit 4: Gene Expression. (n.d.). University of the People.
https://my.uopeople.edu/pluginfile.php/1892132/mod_book/chapter/525108/Unit%204%20Gene
%20expression%20.pdf
Unit 4: RNA Processing. (n.d.). University of the People.
https://my.uopeople.edu/pluginfile.php/1892132/mod_book/chapter/525108/Unit%204%20RNA
%20Processing.pdf
Unit 4: Translation. (n.d.). University of the People.
https://my.uopeople.edu/pluginfile.php/1892132/mod_book/chapter/525108/Unit
%204%20Translation.pdf