Influence of mRNA Sequence on Translation

Influence of mRNA Sequence on the 3 Stages of Translation

• The formation of the initiation complex CAP depends on the Kozak sequence, and IRES sequences can allow for internal initiations.
• The 5'UTR sequence influences translation, and the scanning of the initiation sequence depends on the Kozak sequence.

Elongation Stage

• Ribosomes do not read all triplets with the same ease, and there is a bias in the use of codons, making some easier to read than others due to the abundance of tRNAs for each codon.
• Slipping sequences can change the reading pattern, and even the presence of a bulky amino acid can hinder the exit of the peptide through the ribosomal channel.

Termination Stage

• The stop triplet can sometimes be read as selenocysteine or pyrrolysine, and there are stops that allow the small subunit to remain on the mRNA and reach later reading patterns.

Regulation of Translation at the mRNA Level

5.1 Regulation with Implication of 5'UTR and ORFs

• The GCN4 gene has a long 5'UTR region and two short ORFs that code for peptides without function, which trap ribosomes that have started translation from the CAP and are located before the sequence that codes for GCN4.
• The first uORF terminates in a permissive termination, and the second uORF4 terminates in a non-permissive termination.

Regulation of GCN4 Translation

• In conditions of sufficient amino acids, eIF2 is active, allowing the ribosome to translate and encounter the first reading pattern (uORF1), which it translates, and upon reaching the stop triplet, it encounters a permissive termination, causing the large subunit to dissociate, but the small subunit remains bound to the mRNA.
• The small subunit then reaches the second reading pattern (uORF4), but since the concentration of active eIF2 is high, it has time to bind to the small subunit before it starts reading uORF4, allowing the complete ribosome to form and translate uORF4.
• In conditions of amino acid scarcity, the concentration of active eIF2 is low, and the small subunit reaches the first reading pattern (uORF1) and encounters a permissive termination, but since the concentration of active eIF2 is low, the small subunit continues scanning the mRNA and bypasses the non-permissive termination of uORF4, allowing it to reach the GCN4 sequence and translate it.

uORFs and Their Effects on Translation

• uORFs are a trap for ribosomes, making them start translating from the uORF and getting trapped, and never reaching the real coding sequence.
• The presence of a uORF is a regulatory mechanism that determines the translation and/or stability of the mRNA.

5.2 Alternative Translation Initiations

• Regulation of protein quantity in the cell can occur through alternative translation initiations, such as skipping the uORF or using alternative AUGs.
• Alternative ORFs (aORFs) can produce truncated proteins, and the cell can combine alternative translation initiations and produce proteins with variable N-termini.

5.3 5'-UTR: Autogenous Regulation

• The 5'-UTR region has various regulatory elements, such as IRES sequences, uORF sequences, and sequences that form structures recognizable by proteins related to translation regulation.
• There are autogenous regulatory mechanisms, similar to those in prokaryotes, that control the translation of certain proteins based on whether they are in excess or deficiency.

5.4 3'-UTR: Activation via PABP

• The 3'-UTR region has structures recognizable by proteins related to mRNA stability, and binding sites for miRNAs, which can regulate translation initiation.
• The polyA tail in the 3'-UTR stimulates translation by being a binding site for PABP, which is an stimulator of the formation of the cap-binding complex, making it easier to initiate translation.