This region is important for the regulation of translation of a transcript by differing mechanisms in viruses, prokaryotes and eukaryotes.

In many organisms, however, the 5′ UTR is completely untranslated, instead forming a complex secondary structure to regulate translation.

[3] The 5′ UTR begins at the transcription start site and ends one nucleotide (nt) before the initiation sequence (usually AUG) of the coding region.

The prokaryotic 5′ UTR contains a ribosome binding site (RBS), also known as the Shine–Dalgarno sequence (AGGAGGU), which is usually 3–10 base pairs upstream from the initiation codon.

[6] In contrast, the eukaryotic 5′ UTR contains the Kozak consensus sequence (ACCAUGG), which contains the initiation codon.

[6] The eukaryotic 5′ UTR also contains cis-acting regulatory elements called upstream open reading frames (uORFs) and upstream AUGs (uAUGs) and termination codons, which have a great impact on the regulation of translation (see below).

[10] In bacteria, the initiation of translation occurs when IF-3, along with the 30S ribosomal subunit, bind to the Shine–Dalgarno (SD) sequence of the 5′ UTR.

However, it has also been noted that SXL can also repress translation of RNAs that do not contain a poly(A) tail, or more generally, 3′ UTR.

The 5′ UTR has the ability to form a hairpin loop secondary structure (known as the iron response element or IRE) that is recognized by iron-regulatory proteins (IRP1 and IRP2).

In low levels of iron, the ORF of the target mRNA is blocked as a result of steric hindrance from the binding of IRP1 and IRP2 to the IRE.

This function has gained some interest after it was revealed that the translation of amyloid precursor protein may be disrupted due to a single-nucleotide polymorphism to the IRE found in the 5′ UTR of its mRNA, leading to a spontaneous increased risk of Alzheimer's disease.

[17] Another form of translational regulation in eukaryotes comes from unique elements on the 5′ UTR called upstream open reading frames (uORF).

Control of protein regulation is determined by the distance between the uORF and the first codon in the main ORF.

[20] A uORF has been found to increase reinitiation with the longer distance between its uAUG and the start codon of the main ORF, which indicates that the ribosome needs to reacquire translation factors before it can carry out translation of the main protein.

[21] The IRES enables the viral transcript to translate more efficiently due to the lack of needing a preinitation complex, allowing the virus to replicate quickly.