The

mRNA In molecular biology, messenger ribonucleic acid (mRNA) is a single-stranded molecule of RNA that corresponds to the genetic sequence of a gene, and is read by a ribosome in the process of synthesizing a protein. mRNA is created during the ...

decapping complex is a protein complex in eukaryotic cells responsible for removal of the

5' cap In molecular biology, the five-prime cap (5′ cap) is a specially altered nucleotide on the 5′ end of some primary transcripts such as precursor messenger RNA. This process, known as mRNA capping, is highly regulated and vital in the creation o ...

. The active enzyme of the decapping complex is the bilobed Nudix family enzyme

Dcp2 mRNA-decapping enzyme 2 is a protein that in humans is encoded by the ''DCP2'' gene In biology, the word gene (from , ; "... Wilhelm Johannsen coined the word gene to describe the Mendelian units of heredity..." meaning ''generation'' or ...

, which hydrolyzes

and releases 7mGDP and a 5'-monophosphorylated mRNA. This decapped mRNA is inhibited for translation and will be degraded by

exonuclease Exonucleases are enzymes that work by cleaving nucleotides one at a time from the end (exo) of a polynucleotide chain. A hydrolyzing reaction that breaks phosphodiester bonds at either the 3′ or the 5′ end occurs. Its close relative is t ...

s. The core decapping complex is conserved in eukaryotes. Dcp2 is activated by Decapping Protein 1 (Dcp1) and in higher

eukaryote Eukaryotes () are organisms whose cells have a nucleus. All animals, plants, fungi, and many unicellular organisms, are Eukaryotes. They belong to the group of organisms Eukaryota or Eukarya, which is one of the three domains of life. Bact ...

s joined by the scaffold protein VCS. Together with many other accessory proteins, the decapping complex assembles in

P-bodies P-bodies, or processing bodies are distinct foci formed by phase separation within the cytoplasm of the eukaryotic cell consisting of many enzymes involved in mRNA turnover. P-bodies are highly conserved structures and have been observed in s ...

in the

cytoplasm In cell biology, the cytoplasm is all of the material within a eukaryotic cell, enclosed by the cell membrane, except for the cell nucleus. The material inside the nucleus and contained within the nuclear membrane is termed the nucleoplasm. ...

Purpose of the decapping complex

needs to be degraded, or else it will keep floating around the cell and create unwanted proteins at random. The mRNA 5' cap is specifically designed to keep mRNA from being degraded before it can be used, and so needs to be removed so the mRNA decay pathway can take care of it.

Decapping mechanism

is the protein that actually decaps mRNA, and the rest of proteins in the complex enhance its function and allow it to

hydrolyze Hydrolysis (; ) is any chemical reaction in which a molecule of water breaks one or more chemical bonds. The term is used broadly for substitution, elimination, and solvation reactions in which water is the nucleophile. Biological hydrolysis ...

the chemical bond attaching the mRNA to the

. The Nudix domain in Dcp2

hydrolyzes Hydrolysis (; ) is any chemical reaction in which a molecule of water breaks one or more chemical bonds. The term is used broadly for substitution, elimination, and solvation reactions in which water is the nucleophile. Biological hydrolysis ...

one of the bonds on the triphosphate bridge that hooks the

and the

together, causing the

7-methylguanosine 7-Methylguanosine (m7G) is a modified purine nucleoside. It is a methylated version of guanosine and when found in human urine, it may be a biomarker of some types of cancer. In the RNAs, 7-methylguanosine have been used to study and examine the ...

cap to come off and leaving the mRNA open to degradation by the

s in the cell.

Structure of the decapping complex

Both single-celled and multicellular organisms need to decap their mRNA to get rid of it, but different organisms have slightly different proteins that carry out this process. There are many proteins that stay the same, but several key differences between the single-celled (

yeast Yeasts are eukaryotic, single-celled microorganisms classified as members of the fungus kingdom. The first yeast originated hundreds of millions of years ago, and at least 1,500 species are currently recognized. They are estimated to consti ...

) and multicellular (

metazoan Animals are multicellular, eukaryotic organisms in the biological kingdom Animalia. With few exceptions, animals consume organic material, breathe oxygen, are able to move, can reproduce sexually, and go through an ontogenetic stage in ...

) decapping complexes.

Yeast decapping complex

In yeast (''

S. cerevisiae ''Saccharomyces cerevisiae'' () (brewer's yeast or baker's yeast) is a species of yeast (single-celled fungus microorganisms). The species has been instrumental in winemaking, baking, and brewing since ancient times. It is believed to have been o ...

''), Dcp2 is joined by the decapping activator Dcp1, the

helicase Helicases are a class of enzymes thought to be vital to all organisms. Their main function is to unpack an organism's genetic material. Helicases are motor proteins that move directionally along a nucleic acid phosphodiester backbone, separat ...

Dhh1, the

Xrn1,

nonsense mediated decay Nonsense-mediated mRNA decay (NMD) is a surveillance pathway that exists in all eukaryotes. Its main function is to reduce errors in gene expression by eliminating mRNA transcripts that contain premature stop codons. Translation of these aberrant ...

factors Upf1, Upf2, and Upf3, the LSm complex, Pat1, and various other proteins. These proteins all localize to cytoplasmic structures called

. Notably in yeast there are no translation factors or ribosomal proteins inside P-bodies.

Metazoan decapping complex

Higher eukaryotes have slightly different members of the decapping complex. The enzyme Dcp2 is still the catalytic subunit which forms a

holoenzyme Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrates, and the enzyme converts the substrates into different molecules known as products. A ...

with Dcp1, and interacts with auxiliary proteins such as Xrn1,

Upf1 Regulator of nonsense transcripts 1 is a protein that in humans is encoded by the ''UPF1'' gene. Function This gene encodes a protein that is part of a post-splicing multiprotein complex, the exon junction complex, involved in both mRNA nucle ...

Upf2 Regulator of nonsense transcripts 2 is a protein that in humans is encoded by the ''UPF2'' gene. Function This gene encodes a protein that is part of a post-splicing multiprotein complex, the exon junction complex, involved in both mRNA nucle ...

, Upf3, the LSm complex, and the Dhh1 ortholog DDX6. Proteins unique to plants and mammals include the

beta propeller In structural biology, a beta-propeller (β-propeller) is a type of all-β protein architecture characterized by 4 to 8 highly symmetrical blade-shaped beta sheets arranged toroidally around a central axis. Together the beta-sheets form a funnel- ...

protein Hedls and the enhancer of decapping Edc3. Researchers know how the complex physically associates because of

immunoprecipitation Immunoprecipitation (IP) is the technique of precipitating a protein antigen out of solution using an antibody that specifically binds to that particular protein. This process can be used to isolate and concentrate a particular protein from a sam ...

, while structural details of each part of the complex have been discovered by using

x-ray crystallography X-ray crystallography is the experimental science determining the atomic and molecular structure of a crystal, in which the crystalline structure causes a beam of incident X-rays to diffract into many specific directions. By measuring the angle ...

in conjunction with protein chrystallization. Each of these proteins contribute different things to the decapping complex, as discussed below.

Dcp2

, as the main

catalyst Catalysis () is the process of increasing the rate of a chemical reaction by adding a substance known as a catalyst (). Catalysts are not consumed in the reaction and remain unchanged after it. If the reaction is rapid and the catalyst recyc ...

of the decapping process, relies on a specific pattern of amino acids called a nudix domain to align itself with the

in order to

it. A nudix

domain Domain may refer to: Mathematics *Domain of a function, the set of input values for which the (total) function is defined ** Domain of definition of a partial function **Natural domain of a partial function **Domain of holomorphy of a function *Do ...

is made by packing two

beta sheet The beta sheet, (β-sheet) (also β-pleated sheet) is a common motif of the regular protein secondary structure. Beta sheets consist of beta strands (β-strands) connected laterally by at least two or three backbone hydrogen bonds, forming a gen ...

s between multiple

alpha helices The alpha helix (α-helix) is a common motif in the secondary structure of proteins and is a right hand-helix conformation in which every backbone N−H group hydrogen bonds to the backbone C=O group of the amino acid located four residues earli ...

, can be various lengths and sizes, and is generally used by proteins to carry out

dephosphorylation In biochemistry, dephosphorylation is the removal of a phosphate (PO43−) group from an organic compound by hydrolysis. It is a reversible post-translational modification. Dephosphorylation and its counterpart, phosphorylation, activate and deact ...

, getting rid of a

phosphate In chemistry, a phosphate is an anion, salt, functional group or ester derived from a phosphoric acid. It most commonly means orthophosphate, a derivative of orthophosphoric acid . The phosphate or orthophosphate ion is derived from phosph ...

by inserting a water molecule into the bond between the phosphate and the rest of the molecule. In the case of Dcp2, it contains multiple

glutamic acid Glutamic acid (symbol Glu or E; the ionic form is known as glutamate) is an α-amino acid that is used by almost all living beings in the biosynthesis of proteins. It is a non-essential nutrient for humans, meaning that the human body can syn ...

side chains that are negatively charged in normal cellular conditions, and these are what allow the protein to manipulate water molecules to hydrolyze the tri-phosphate bridge that connects the 5' end of the mRNA to the

cap. Therefore, the nudix domain is what allows Dcp2 to remove the 5' cap, which results in the creation 7mGDP, a 7-methylguanosine with two phosphate groups attached, and a monophosphorylated mRNA strand. Before the nudix domain is an N-terminal regulatory domain (NRD), which further helps hydrolyze the 5' mRNA cap. After the nudix domain is a C-terminal area called Box B, which helps bind Dcp2 to RNA. With all three of these main motifs, Dcp2 is able to find, bind firmly to, and hydrolyzes a 5' mRNA cap. It does this either by recognizing a

hairpin loop Stem-loop intramolecular base pairing is a pattern that can occur in single-stranded RNA. The structure is also known as a hairpin or hairpin loop. It occurs when two regions of the same strand, usually complementary in nucleotide sequence whe ...

in the RNA within 10 base pairs of the cap, which is called a Dcp2 binding and decapping element, or by a separate protein recognizing a base pair pattern in the mRNA and directly recruiting the Dcp2-Dcp1 holoenzyme. Unfortunately, Dcp2 works slowly, and needs a few other proteins to coordinate with it so it can decap mRNA in a timely manner.

Dcp1

Dcp1 is a regulatory subunit, it combines with Dcp2, creating a

that can decap mRNA properly. Without Dcp1, it is actually impossible for Dcp2 to decap anything

in vivo Studies that are ''in vivo'' (Latin for "within the living"; often not italicized in English) are those in which the effects of various biological entities are tested on whole, living organisms or cells, usually animals, including humans, and ...

, and it only works incredibly slowly

in vitro ''In vitro'' (meaning in glass, or ''in the glass'') studies are performed with microorganisms, cells, or biological molecules outside their normal biological context. Colloquially called "test-tube experiments", these studies in biology and ...

, which makes forming this holoenzyme an essential process in decapping. Dcp1's

secondary structure Protein secondary structure is the three dimensional form of ''local segments'' of proteins. The two most common secondary structural elements are alpha helices and beta sheets, though beta turns and omega loops occur as well. Secondary struct ...

consists of seven

s and three

. which come together to form a V-shaped

tertiary structure Protein tertiary structure is the three dimensional shape of a protein. The tertiary structure will have a single polypeptide chain "backbone" with one or more protein secondary structures, the protein domains. Amino acid side chains may int ...

. The defining features of Dcp1 are the

EVH1 domain WH1 domain is an evolutionary conserved protein domain. Therefore, it has an important function. Function WH1 domains are found on WASP proteins, which are often involved in actin polymerization. Hence, WH1 is important for all cellular process ...

and a

that recognises proline rich sequences (PRS) on other proteins. The EVH1 domain interacts directly with the earlier mentioned NRD of Dcp2, and is currently thought to directly help with the decapping of mRNA, though how it does so is unclear. The domain that recognises PRS is made of mostly

hydrophobic In chemistry, hydrophobicity is the physical property of a molecule that is seemingly repelled from a mass of water (known as a hydrophobe). In contrast, hydrophiles are attracted to water. Hydrophobic molecules tend to be nonpolar and, ...

amino acids, and is found within the cleft of the 'V' of the Dcp1 structure. It is used to bind to other proteins in the decapping complex to Dcp1.

PNRC2

PNRC2 attaches to and enhances the effect of Dcp1 to encourage decapping, and also recruits

to the decapping complex. It possesses a proline rich sequence that is hydrophobic and sticks strongly to the equally hydrophobic cleft in Dcp1, and so Dcp1 binds PNRC2's proline-rich region, which then enhances the function of Dcp2 even more. Current research suggests PNRC2 helps associate Dcp2 and Dcp1 together, making the Dcp2-Dcp1

more stable and therefore increasing the effectiveness of Dcp2, but the exact details about how it does so are vague. The recruitment of

allows the decapping complex to participate in

nonsense-mediated mRNA decay Nonsense-mediated mRNA decay (NMD) is a surveillance pathway that exists in all eukaryotes. Its main function is to reduce errors in gene expression by eliminating mRNA transcripts that contain premature stop codons. Translation of these aberrant ...

, which makes PNRC2 a way for Dcp2 to connect with the regulatory pathway in charge of destroying incorrectly transcripted mRNA.

Upf1-3

, and Upf3 are proteins involved in the regulatory pathway of

, and not the actual decapping of mRNA. Only Upf1 attaches directly to the decapping complex, whereas Upf2 and Upf3 attach to mRNA, then attach to Upf1 to facilitate the destruction of incorrect mRNA. These are activators of the complex, in that they can direct the complex at incorrectly formed mRNA, but do not actually help decap the mRNA.

DDX6

DDX6, an ortholog of Dhh1, also enhances the effectiveness of the Dcp2-Dcp1 holoenzyme while it hydrolyzes the 5' cap. It is proposed that, since it is a

, it is involved in reconfiguring the

5' end Directionality, in molecular biology and biochemistry, is the end-to-end chemical orientation of a single strand of nucleic acid. In a single strand of DNA or RNA, the chemical convention of naming carbon atoms in the nucleotide pentose-sugar- ...

of the mRNA to give Dcp2 easier access to the 5' cap, and that it stimulates Dcp1 so that it interacts better with Dcp2 when attached to the rest of the decapping complex.

Edc3

Edc3 Enhancer of mRNA-decapping protein 3 is a protein that in humans is encoded by the ''EDC3'' gene. EDC3 is associated with an mRNA-decapping complex The mRNA decapping complex is a protein complex in eukaryotic cells responsible for removal ...

further activates the Dcp2-Dcp1 holoenzyme and allows it to quickly decap mRNA. It possesses an LSm domain at its

N-terminus The N-terminus (also known as the amino-terminus, NH2-terminus, N-terminal end or amine-terminus) is the start of a protein or polypeptide, referring to the free amine group (-NH2) located at the end of a polypeptide. Within a peptide, the ami ...

, which interacts with specific amino acid motifs called HLM fragments which are found on the

C terminus The C-terminus (also known as the carboxyl-terminus, carboxy-terminus, C-terminal tail, C-terminal end, or COOH-terminus) is the end of an amino acid chain (protein or polypeptide), terminated by a free carboxyl group (-COOH). When the protein is ...

of Dcp1 and allows for Edc3 to bind to it. Another important part of this protein is the FDF linker, which is a long and unstructured stretch of amino acids that binds with DDX6 and stops it from binding directly with the mRNA, allowing it to interact with the proteins in the decapping complex instead. The final domain of note is a Yjef-N C-terminus domain which dimerizes with mRNA and helps create

around the location of the decapping complex.

are essentially stockpiled clumps of decapped or repressed mRNA mixed together with mRNA degradation factors, such as the decapping complex and the nonsense-mediated mRNA decay machinery, so they are important for the eventual destruction of the mRNA altered by Dcp2. As Edc3 creates P-bodies around the decapping complex, it becomes easier for Dcp2 to find mRNA 5' caps to hydrolyze, increasing the effectiveness of the entire complex.

Pat1

Pat1 is another protein that increases the efficiency of the decapping complex. It has three main domains. One is necessary for decapping mRNA, and directly helps the Dcp2-Dcp1 holoenzyme do so. The other two make it easier for the protein to decap mRNA, but are not directly involved in the hydrolysis of the phosphate bond. Pat1 has many interactions with the various proteins in the decapping complex, and is known as the 'scaffolding protein' because it brings everything together when it is time to decap something. The N-terminus domain interacts with DXX6 and brings it close so it can activate Dcp1, another portion helps create P-bodies along with Edc3, and the C-terminus domains attach Dcp1–Dcp2, the Lsm1–7 complex and Xrn1 to the complex. Monophosphorylated mRNA

Xrn1

Xrn1 is a 5' to 3' exonuclease that degrades the just-decapped mRNA. It targets the 5' monophosphate end of mRNA, which is what is left over when Dcp2 has hydrolyzed the cap off and taken away the

cap, along with two of the three phosphates that attach the cap to the mRNA. The current theory is that the structure of Xrn1 does not allow a capped mRNA to interact with it because the Xrn1 is structured in such a way that there is

steric hindrance Steric effects arise from the spatial arrangement of atoms. When atoms come close together there is a rise in the energy of the molecule. Steric effects are nonbonding interactions that influence the shape ( conformation) and reactivity of ions ...

that physically blocks the protein from interacting with any mRNA that Dcp2 has not already decapped.

References

{{reflist RNA