5' cap

The 5' cap is added co-transcriptionally in the nucleus once the RNA polymerase II transcript reaches ~25-30 nucleotides in length. The process involves three enzymatic steps: (1) RNA triphosphatase removes a phosphate from the 5' end, (2) guanylyltransferase adds GMP in an unusual 5'-5' triphosphate linkage, and (3) methyltransferase adds a methyl group to the guanine (creating 7-methylguanosine) and often to the 2'-OH of the first one or two ribose sugars. This structure is denoted m7GpppN.

Functionally, the cap: (1) protects mRNA from 5' exonuclease degradation, increasing stability and half-life, (2) facilitates export of mature mRNA from nucleus to cytoplasm via cap-binding complex, (3) is recognized by eIF4E (eukaryotic initiation factor 4E) as part of the eIF4F complex, which recruits the 40S ribosomal subunit for cap-dependent translation initiation, and (4) enhances pre-mRNA splicing efficiency, particularly of the first intron.

Clinical/molecular biology relevance: Cap-dependent translation is a target of dysregulation in cancer—overexpression of eIF4E is oncogenic and seen in several malignancies, making it a therapeutic target. Some viruses (e.g., influenza) perform 'cap-snatching,' stealing 5' caps from host mRNAs to prime their own viral mRNA synthesis. This is a target for antiviral drugs like baloxavir marboxil, which inhibits the cap-dependent endonuclease of influenza polymerase. Additionally, understanding the cap structure is essential for mRNA vaccine technology (e.g., COVID-19 mRNA vaccines), which incorporate synthetic cap analogs to ensure stability and efficient translation of the vaccine mRNA in host cells.

Contrast with poly-A tail (3' end modification) which also protects mRNA and aids export/translation, added post-transcriptionally by cleavage and polyadenylation of the pre-mRNA.