5' cap

5' cap

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The 5' cap is a specially altered nucleotide
Nucleotide
Nucleotides are molecules that, when joined together, make up the structural units of RNA and DNA. In addition, nucleotides participate in cellular signaling , and are incorporated into important cofactors of enzymatic reactions...

 on the 5' end of precursor messenger RNA and some other primary RNA transcripts as found in eukaryotes. The process of 5' capping is vital to creating mature
Mature messenger RNA
Mature messenger RNA, often abbreviated as mature mRNA is a eukaryotic RNA transcript that has been spliced and processed and is ready for translation in the course of protein synthesis...

 messenger RNA
Messenger RNA
Messenger RNA is a molecule of RNA encoding a chemical "blueprint" for a protein product. mRNA is transcribed from a DNA template, and carries coding information to the sites of protein synthesis: the ribosomes. Here, the nucleic acid polymer is translated into a polymer of amino acids: a protein...

, which is then able to undergo translation
Translation (genetics)
In molecular biology and genetics, translation is the third stage of protein biosynthesis . In translation, messenger RNA produced by transcription is decoded by the ribosome to produce a specific amino acid chain, or polypeptide, that will later fold into an active protein...

. Capping ensures the messenger RNA's stability while it undergoes translation in the process of protein synthesis, and is a highly regulated process that occurs in the cell nucleus
Cell nucleus
In cell biology, the nucleus is a membrane-enclosed organelle found in eukaryotic cells. It contains most of the cell's genetic material, organized as multiple long linear DNA molecules in complex with a large variety of proteins, such as histones, to form chromosomes. The genes within these...

.

5' cap structure




The 5' cap is found on the 5' end of an mRNA molecule and consists of a guanine nucleotide connected to the mRNA via an unusual 5' to 5' triphosphate linkage. This guanosine is methylated on the 7 position directly after capping in vivo by a methylated transferase. It is referred to as a 7-methylguanylate cap, abbreviated m7G.

Further modifications include the possible methylation of the 2' hydroxy-groups of the first 2 ribose sugars of the 5' end of the mRNA. The methylation of both 2' hydroxy-groups is shown on the diagram.

The 5' cap looks like the 3' end of an RNA molecule (the 5' carbon of the cap ribose is bonded, and the 3' unbonded). This provides significant resistance to 5' exonucleases.

Capping process


The starting point is the unaltered 5' end of an RNA molecule. This features a final nucleotide followed by three phosphate groups attached to the 5' carbon.
  1. One of the terminal phosphate groups is removed (by RNA terminal phosphatase), leaving two terminal phosphates.
  2. GTP is added to the terminal phosphates (by a guanylyl transferase
    Guanylyl transferase
    Guanylyl transferases are enzymes that transfer a guanosine monophosphate group, usually from GTP to another molecule, releasing pyrophosphate. Many eukaryotic guanylyl transferases are capping enzymes that catalyze the formation of the 5' cap in the co-transcriptional modification of messenger RNA...

    ), losing two phosphate groups (from the GTP) in the process. This results in the 5' to 5' triphosphate linkage.
  3. The 7-nitrogen of guanine is methylated (by a methyl transferase).
  4. Other methyltransferases are optionally used to carry out methylation of 5' proximal nucleotides.

5' capping targeting


The Capping Enzyme Complex (CEC) required for capping is found bound to the RNA polymerase II
RNA polymerase II
RNA polymerase II is an enzyme found in eukaryotic cells. It catalyzes the transcription of DNA to synthesize precursors of mRNA and most snRNA and microRNA. A 550 kDa complex of 12 subunits, RNAP II is the most studied type of RNA polymerase...

 before transcription starts. As soon as the 5' end of the new transcript emerges the enzymes transfer to it and begin the capping process (this is a similar kind of mechanism to ensure capping as for polyadenylation
Polyadenylation
Polyadenylation is the addition of a poly tail to an RNA molecule. The poly tail consists of multiple adenosine monophosphates; in other words, it is a stretch of RNA that has only adenine bases. In eukaryotes, polyadenylation is part of the process that produces mature messenger RNA for translation...

).

The enzymes for capping can only bind to RNA polymerase II
RNA polymerase II
RNA polymerase II is an enzyme found in eukaryotic cells. It catalyzes the transcription of DNA to synthesize precursors of mRNA and most snRNA and microRNA. A 550 kDa complex of 12 subunits, RNAP II is the most studied type of RNA polymerase...

 ensuring specificity to only these transcripts, which are almost entirely mRNA.

5' cap function


The 5' cap has 4 main functions:
  1. Regulation of nuclear export.
  2. Prevention of degradation by exonucleases.
  3. Promotion of translation (see ribosome
    Ribosome
    A ribosome is a component of cells that assembles the twenty specific amino acid molecules to form the particular protein molecule determined by the nucleotide sequence of an RNA molecule....

     and translation).
  4. Promotion of 5' proximal intron excision.


Nuclear export of RNA is regulated by the Cap binding complex
Cap binding complex
The 5' cap of eukaryotic messenger RNA is bound at all times by various Cap-binding complexes.-Nuclear cap-binding complex:In the nucleus freshly transcribed mRNA molecules are bound on the 5' cap by the nuclear cap-binding complex of Cbc1/Cbc2 in yeast or CBC20/CBC80 in metazoans. These aid in...

 (CBC), which binds exclusively to capped RNA. The CBC is then recognized by the nuclear pore complex and exported. Once in the cytoplasm after the pioneer round of translation, the CBC is replaced by the translation factors eIF-4E and eIF-4G. This complex is then recognized by other translation initiation machinery including the ribosome.

Cap prevents 5' degradation in two ways. First, degradation of the mRNA by 5' exonucleases is prevented (as mentioned above) by functionally looking like a 3' end. Second, the CBC complex and the eIF-4E/eIF-4G block the access of decapping enzymes to the cap. This increases the half-life
Half-life
Half-life, abbreviated t½, is the period of time it takes for the amount of a substance undergoing decay to decrease by half. The name was originally used to describe a characteristic of unstable atoms , but it may apply to any quantity which follows a set-rate decay.The original term, dating to...

 of the mRNA, essential in eukaryotes as the export process takes significant time.

Decapping of an mRNA is catalyzed by the decapping complex made up of at least Dcp1 and Dcp2, which must compete with eIF-4E to bind the cap. Thus the 5' cap is a marker of an actively translating mRNA and is used by cells to regulate mRNA half-lives in response to new stimuli. Undesirable mRNAs are sent to P-bodies
P-bodies
Processing bodies were first described in the scientific literature by Bashkirov et. al. in 1997, in which they describe "small granules… discrete, prominent foci" as the cytoplasmic location of the mouse exoribonuclease mXrn1p. It wasn’t until 2002 that a glimpse into the nature and importance of...

 for temporary storage or decapping, the details of which are still being resolved.

The mechanism of 5' proximal intron excision promotion is not well understood, but the 5' cap appears to loop around and interact with the spliceosome
Spliceosome
A spliceosome is a complex of snRNA and protein subunits that removes introns from a transcribed pre-mRNA segment. This process is generally referred to as splicing.-Composition:...

in the splicing process, promoting intron excision.