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U2 spliceosomal snRNAs are a species of
small nuclear RNA Small nuclear RNA (snRNA) is a class of small RNA molecules that are found within the splicing speckles and Cajal bodies of the cell nucleus in eukaryotic cells. The length of an average snRNA is approximately 150 nucleotides. They are transcribe ...
(
snRNA Small nuclear RNA (snRNA) is a class of small RNA molecules that are found within the splicing speckles and Cajal bodies of the cell nucleus in eukaryotic cells. The length of an average snRNA is approximately 150 nucleotides. They are transcri ...
) molecules found in the major spliceosomal (Sm) machinery of virtually all eukaryotic organisms. ''In vivo'', U2 snRNA along with its associated polypeptides assemble to produce the U2
small nuclear ribonucleoprotein snRNPs (pronounced "snurps"), or small nuclear ribonucleoproteins, are RNA-protein complexes that combine with unmodified pre-mRNA and various other proteins to form a spliceosome, a large RNA-protein molecular complex upon which splicing of pre-m ...
(
snRNP snRNPs (pronounced "snurps"), or small nuclear ribonucleoproteins, are RNA-protein complexes that combine with unmodified pre-mRNA and various other proteins to form a spliceosome, a large RNA-protein molecular complex upon which splicing of pre- ...
), an essential component of the major spliceosomal complex. The major spliceosomal-splicing pathway is occasionally referred to as U2 dependent, based on a class of Sm
intron An intron is any nucleotide sequence within a gene that is not expressed or operative in the final RNA product. The word ''intron'' is derived from the term ''intragenic region'', i.e. a region inside a gene."The notion of the cistron .e., gene. ...
—found in mRNA primary transcripts—that are recognized exclusively by the U2 snRNP during early stages of spliceosomal assembly. In addition to U2 dependent intron recognition, U2 snRNA has been theorized to serve a catalytic role in the chemistry of pre-RNA splicing as well. Similar to
ribosomal RNA Ribosomal ribonucleic acid (rRNA) is a type of non-coding RNA which is the primary component of ribosomes, essential to all cells. rRNA is a ribozyme which carries out protein synthesis in ribosomes. Ribosomal RNA is transcribed from ribosomal ...
s ( rRNAs), Sm snRNAs must mediate both RNA:RNA and RNA:protein contacts and hence have evolved specialized, highly conserved, primary and secondary structural elements to facilitate these types of interactions. Shortly after the discovery that
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 Protein biosynthesis, synthesizing a protein. mRNA is ...
primary transcripts contain long, non-coding intervening sequences (
intron An intron is any nucleotide sequence within a gene that is not expressed or operative in the final RNA product. The word ''intron'' is derived from the term ''intragenic region'', i.e. a region inside a gene."The notion of the cistron .e., gene. ...
s) by
Sharp Sharp or SHARP may refer to: Acronyms * SHARP (helmet ratings) (Safety Helmet Assessment and Rating Programme), a British motorcycle helmet safety rating scheme * Self Help Addiction Recovery Program, a charitable organisation founded in 19 ...
and Roberts,
Joan Steitz Joan Elaine Argetsinger Steitz (born January 26, 1941) is Sterling Professor of Molecular Biophysics and Biochemistry at Yale University and Investigator at the Howard Hughes Medical Institute. She is known for her discoveries involving RNA, incl ...
began work to characterize the biochemical mechanism of intron excision. The curious observation that a sequence found in the 5´ region of the U1 snRNA exhibited extensive base pairing complementarity with conserved sequences across 5´ splice junctions in
hnRNA A primary transcript is the single-stranded ribonucleic acid ( RNA) product synthesized by transcription of DNA, and processed to yield various mature RNA products such as mRNAs, tRNAs, and rRNAs. The primary transcripts designated to be mRNAs ...
transcripts prompted speculation that certain snRNAs may be involved in recognizing splice site boundaries through RNA:RNA contacts. Only recently have atomic crystal structures revealed demonstrably that the original conjecture was indeed correct, even if the complexity of these interactions were not fully realized at the time.


U2 snRNA Recognition Elements

In ''
Saccharomyces 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 ...
'' the U2 snRNA is associated with 18
polypeptides Peptides (, ) are short chains of amino acids linked by peptide bonds. Long chains of amino acids are called proteins. Chains of fewer than twenty amino acids are called oligopeptides, and include dipeptides, tripeptides, and tetrapeptides. A p ...
, seven of which are structural proteins common to all Sm class snRNPs. These non-specific structural proteins associate with Sm snRNAs through a highly conserved recognition sequence (AU''n''G,''n'' = 4-6) located within the RNA called Sm-binding sites. Two other proteins, A´ and B´´, are U2-specific and require structural elements unique to U2 snRNA—specifically two 3´ stem loops—for snRNP assembly. The three-subunit SF3a and six-subunit SF3b protein complexes also associate with the U2 snRNA. U2 snRNA is implicated in intron recognition through a 7-12 nucleotide sequence between 18-40 nucleotides upstream of the 3´ splice site known as the branch point sequence (BPS). In
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 constitut ...
, the consensus BPS is 7 nucleotide residues in length and the complementary recognition sequence within the U2 snRNA is 6 nucleotides. Duplex formation between these two sequences results in bulging of a conserved
adenosine Adenosine ( symbol A) is an organic compound that occurs widely in nature in the form of diverse derivatives. The molecule consists of an adenine attached to a ribose via a β-N9-glycosidic bond. Adenosine is one of the four nucleoside building ...
residue at position 5 of the BPS. The bulged adenosine residue adopts a C3´-endo conformation that with the help of splicing factors Cwc25, Yju2 and Isy1 aligns a 2´ OH for an inline attack of a
phosphorus Phosphorus is a chemical element with the symbol P and atomic number 15. Elemental phosphorus exists in two major forms, white phosphorus and red phosphorus, but because it is highly reactive, phosphorus is never found as a free element on Ear ...
atom at the 5´ splice site. Nucleophilic attack initiates the first of two successive
transesterification In organic chemistry, transesterification is the process of exchanging the organic group R″ of an ester with the organic group R' of an alcohol. These reactions are often catalyzed by the addition of an acid or base catalyst. The reaction can ...
reactions that splices out the intron—through an unusual 2´-5´-3´ linked lariat intermediate—where the second transesterification involves ligation of the two flanking exons.


Primary and Secondary Structure

Although the sequence length of U2 snRNAs can vary by up to an order of magnitude across all
eukaryotic 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. Bacte ...
organisms, all U2 snRNAs contain many phylogenetically constant regions particularly within the first 80 nucleotides downstream of the 5´ end where 85% of the positions are conserved. Moreover, several secondary structural elements are also conserved including stem loops I, II, III, IV, and some of the single stranded regions linking these domains. Stem loop II in yeast U2 snRNA, contains an unusual sheared GA base pair leading into a characteristic U-turn loop motif that shares a geometric conformation similar to that of
tRNA Transfer RNA (abbreviated tRNA and formerly referred to as sRNA, for soluble RNA) is an adaptor molecule composed of RNA, typically 76 to 90 nucleotides in length (in eukaryotes), that serves as the physical link between the mRNA and the amino ac ...
anti-codon loops. All U2 snRNAs possess a terminal
stem 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 ...
(IV) with a 10-16 base pair helix and a conserved 11 nucleotide loop with the consensus sequence 5´-UYGCANUURYN-3´. U2 snRNAs are the most extensively modified of all the small nuclear RNAs. While the exact locations of these
post-transcriptional modification Transcriptional modification or co-transcriptional modification is a set of biological processes common to most eukaryotic cells by which an RNA primary transcript is chemically altered following transcription from a gene to produce a mature, func ...
s can vary from organism to organism, emerging evidence suggests there is a strong correlation between U2 snRNA modification and biological function. Modifications include the conversion of some
uridine Uridine (symbol U or Urd) is a glycosylated pyrimidine analog containing uracil attached to a ribose ring (or more specifically, a ribofuranose) via a β-N1-glycosidic bond. The analog is one of the five standard nucleosides which make up nuclei ...
residues to
pseudouridine Pseudouridine (abbreviated by the Greek letter psi- Ψ) is an isomer of the nucleoside uridine in which the uracil is attached via a carbon-carbon instead of a nitrogen-carbon glycosidic bond. (In this configuration, uracil is sometimes referred ...
, 2´-O-methylation, nucleobase methylation, and conversion of 5´-monomethylated guanosine cap to a 2,2,7-trimethylated guanosine cap. Many of these modifications reside in a 27-nucleotide region on the 5´ end of the molecule.


Conformational Dynamics

The
spliceosome A spliceosome is a large ribonucleoprotein (RNP) complex found primarily within the nucleus of eukaryotic cells. The spliceosome is assembled from small nuclear RNAs (snRNA) and numerous proteins. Small nuclear RNA (snRNA) molecules bind to specifi ...
is a dynamic molecular machine that undergoes several conformational rearrangements throughout assembly and splicing. Although many of the biochemical details surrounding spliceosomal rearrangements remains unclear, recent studies have visualized the formation of a critical folding complex between U2 and U6 snRNAs immediately proceeding the first step of the splicing reaction. This folding event facilitates the formation of a four-helix junction, which is believed to provide scaffolding for the critical components of the active site including aligning the 5´ splice site with the branch point adenosine for inline attack by the 2´ OH and coordinating two Mg2+ ions to stabilize negative charge formation in the proceeding steps.


Evolutionary Origins

A notable characteristic of the U2-U6 fold is its structural similarity to that of domain V in self-splicing
group II intron Group II introns are a large class of self-catalytic ribozymes and mobile genetic elements found within the genes of all three domains of life. Ribozyme activity (e.g., self- splicing) can occur under high-salt conditions ''in vitro''. However, ...
s. The AGC triad found in U6 snRNA is conserved in group II introns and has been found to favor the same tertiary stacking interactions as well. The formation of a GU wobble pair early in the U2-U6 folding event is also observed in the formation of the catalytic core of group II introns. Finally, it is likely the spliceosome utilizes the same two-metal ion mechanism as group II introns given the structural conservation of metal binding sites found within the U2-U6 fold. The extent of both secondary and tertiary structure conservation between group II introns and the U2-U6 fold in the active site of the spliceosome strongly suggests both group II introns and the spliceosome share a common evolutionary origin.


See also

*
Spliceosome A spliceosome is a large ribonucleoprotein (RNP) complex found primarily within the nucleus of eukaryotic cells. The spliceosome is assembled from small nuclear RNAs (snRNA) and numerous proteins. Small nuclear RNA (snRNA) molecules bind to specifi ...
*
Small nuclear RNA Small nuclear RNA (snRNA) is a class of small RNA molecules that are found within the splicing speckles and Cajal bodies of the cell nucleus in eukaryotic cells. The length of an average snRNA is approximately 150 nucleotides. They are transcribe ...
*
U1 spliceosomal RNA U1 spliceosomal RNA is the small nuclear RNA (snRNA) component of U1 snRNP (''small nuclear ribonucleoprotein''), an RNA-protein complex that combines with other snRNPs, unmodified pre-mRNA, and various other proteins to assemble a spliceosome, a ...


References


Further reading

* *


External links

* {{Rfam, id=RF00004, name=U2 spliceosomal RNA
U2
Small nuclear RNA Spliceosome RNA splicing