Minor spliceosome

Last updated September 30, 2022

The minor spliceosome is a ribonucleoprotein complex that catalyses the removal (splicing) of an atypical class of spliceosomal introns (U12-type) from messenger RNAs in some clades of eukaryotes. This process is called noncanonical splicing, as opposed to U2-dependent canonical splicing. U12-type introns represent less than 1% of all introns in human cells. However they are found in genes performing essential cellular functions.

Early evidence

A notable feature of eukaryotic nuclear pre-mRNA introns is the relatively high level of conservation of the primary sequences of 5' and 3' splice sites over a great range of organisms.

Between 1989 and 1991, several groups reported four independent examples of introns with a splice site that differed from the common intron:

Cartilage matrix protein (CMP/MATN1) gene in humans and chickens
Proliferating cell nucleolar protein P120 (NOL1) gene in humans
Mouse Rep3 gene, presumably involved in DNA repair
Drosophila prospero gene that encodes for a homeobox protein

In 1991 by comparing the intron sequences of P120 and CMP genes, IJ Jackson reported the existence of ATATCC (5') and YYCAC (3') splice sites in these introns. The finding indicated a possible novel splicing mechanism.

In 1994, S.L. Hall and R.A. Padgett compared the primary sequence of all reports on the four genes mentioned above. The results suggested a new type of introns with ATATCCTT 5' splice sites and YCCAC 3' splice sites and an almost invariant TCCTTAAC sequence near the 3' end of the introns (so called 3' upstream element). A search for small nuclear RNA sequences that are complementary to these splice sites suggested U12 snRNA (matches the 3' sequence) and U11 snRNA (matches the 5' sequence) as being putative factors involved in splicing of this new type of introns.

In all these four genes, the pre-mRNA contains other introns whose sequences conform to those of major class introns. Neither the size nor the position of the AT–AC intron within the host gene is conserved.

In 1996, Woan-Yuh Tarn and Joan A. Steitz described an in vitro system that splices a pre-mRNA substrate containing an AT–AC intron derived from the human P120 gene. Psoralen cross-linking confirms the base-pairing interaction predicted by Hall and Padgett between the branch site of the pre-mRNA substrate and U12 RNA. Native gel electrophoresis reveals that U11, U12, and U5 snRNPs assemble onto the P120 pre-mRNA to form splicing complexes.

Structure of U12-type introns

Although originally referred to as AT-AC introns, not all these introns are delimited by AT-AC dinucleotides. Some of them have GT-AG or AT-AG ends, at least. Thus, it is more correct to speak about the splicing machinery which is used to process them, differentiating between U2-type (canonical or major) and U12-type (non-canonical or minor). The main determinants for distinguishing U2- and U12-type introns are 5' splice site and branch site sequences.^[1]

U1 and U11 can be folded similarly U1-U11-2ndStructureCompared.JPG — U1 and U11 can be folded similarly

The minor spliceosome consists of U11, U12, U4atac, and U6atac, together with U5 and an unknown number of non-snRNP proteins. The U11, U12 and U4atac/U6atac snRNPs are functional analogs of the U1, U2 and U4/U6 snRNPs in the major spliceosome.^[2]^[3]^[4]^[5]^[6] Although the minor U4atac and U6atac snRNAs are functional analogs of U4 and U6, respectively, they share only limited sequence homology (c. 40%). Furthermore, the sequence of U11 in comparison with U1, as well as U12 compared with U2, are completely unrelated. Despite this fact, the minor U11, U12, U4atac and U6atac snRNAs can be folded into structures similar to U1, U2, U4 and U6, respectively.^[7]

Location of minor spliceosomal activity

The location of spliceosomal activity for the minor class spliceosome is regarded by most experts to be in the nucleus.^{[ citation needed ]} However, a single paper has claimed that the minor spliceosome is active in the cytosol.^[8] The data presented within this paper are not fully accepted within the field and directly contradict numerous other papers.

Evolution

Like the major spliceosome, the minor spliceosome had an early origin: several of its characteristic constituents are present in representative organisms from all eukaryotic supergroups for which there is any substantial genome sequence information. In addition, functionally important sequence elements contained within U12-type introns and snRNAs are highly conserved during evolution.

Related Research Articles

An intron is any nucleotide sequence within a gene that is removed by RNA processing during production of the final RNA product. The word intron is derived from the term intragenic region, i.e. a region inside a gene. The term intron refers to both the DNA sequence within a gene and the corresponding RNA sequence in RNA transcripts. The non-intron sequences that become joined by this RNA processing to form the mature RNA are called exons.

RNA splicing is a process in molecular biology where a newly-made precursor messenger RNA (pre-mRNA) transcript is transformed into a mature messenger RNA (mRNA). It works by removing all the introns and splicing back together exons. For nuclear-encoded genes, splicing occurs in the nucleus either during or immediately after transcription. For those eukaryotic genes that contain introns, splicing is usually needed to create an mRNA molecule that can be translated into protein. For many eukaryotic introns, splicing occurs in a series of reactions which are catalyzed by the spliceosome, a complex of small nuclear ribonucleoproteins (snRNPs). There exist self-splicing introns, that is, ribozymes that can catalyze their own excision from their parent RNA molecule. The process of transcription, splicing and translation is called gene expression, the central dogma of molecular biology.

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 specific proteins to form a small nuclear ribonucleoprotein complex, which in turn combines with other snRNPs to form a large ribonucleoprotein complex called a spliceosome. The spliceosome removes introns from a transcribed pre-mRNA, a type of primary transcript. This process is generally referred to as splicing. An analogy is a film editor, who selectively cuts out irrelevant or incorrect material from the initial film and sends the cleaned-up version to the director for the final cut.

SR proteins are a conserved family of proteins involved in RNA splicing. SR proteins are named because they contain a protein domain with long repeats of serine and arginine amino acid residues, whose standard abbreviations are "S" and "R" respectively. SR proteins are ~200-600 amino acids in length and composed of two domains, the RNA recognition motif (RRM) region and the RS domain. SR proteins are more commonly found in the nucleus than the cytoplasm, but several SR proteins are known to shuttle between the nucleus and the cytoplasm.

snRNPs, 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-mRNA occurs. The action of snRNPs is essential to the removal of introns from pre-mRNA, a critical aspect of post-transcriptional modification of RNA, occurring only in the nucleus of eukaryotic cells. Additionally, U7 snRNP is not involved in splicing at all, as U7 snRNP is responsible for processing the 3′ stem-loop of histone pre-mRNA.

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 transcribed by either RNA polymerase II or RNA polymerase III. Their primary function is in the processing of pre-messenger RNA (hnRNA) in the nucleus. They have also been shown to aid in the regulation of transcription factors or RNA polymerase II, and maintaining the telomeres.

Joan Elaine Argetsinger Steitz 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, including ground-breaking insights into how ribosomes interact with messenger RNA by complementary base pairing and that introns are spliced by small nuclear ribonucleic proteins (snRNPs), which occur in eukaryotes. In September 2018, Steitz won the Lasker-Koshland Award for Special Achievement in Medical Science. The Lasker award is often referred to as the 'American Nobel' because 87 of the former recipients have gone on to win Nobel prizes.

The U11 snRNA is an important non-coding RNA in the minor spliceosome protein complex, which activates the alternative splicing mechanism. The minor spliceosome is associated with similar protein components as the major spliceosome. It uses U11 snRNA to recognize the 5' splice site while U12 snRNA binds to the branchpoint to recognize the 3' splice site.

<span class="mw-page-title-main">U12 minor spliceosomal RNA</span>

U12 minor spliceosomal RNA is formed from U12 small nuclear (snRNA), together with U4atac/U6atac, U5, and U11 snRNAs and associated proteins, forms a spliceosome that cleaves a divergent class of low-abundance pre-mRNA introns. Although the U12 sequence is very divergent from that of U2, the two are functionally analogous.

<span class="mw-page-title-main">U1 spliceosomal RNA</span>

U1 spliceosomal RNA is the small nuclear RNA (snRNA) component of U1 snRNP, an RNA-protein complex that combines with other snRNPs, unmodified pre-mRNA, and various other proteins to assemble a spliceosome, a large RNA-protein molecular complex upon which splicing of pre-mRNA occurs. Splicing, or the removal of introns, is a major aspect of post-transcriptional modification, and takes place only in the nucleus of eukaryotes.

<span class="mw-page-title-main">U2 spliceosomal RNA</span>

U2 spliceosomal snRNAs are a species of small nuclear RNA (snRNA) 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 (snRNP), 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—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 RNAs (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.

<span class="mw-page-title-main">U5 spliceosomal RNA</span>

U5 snRNA is a small nuclear RNA (snRNA) that participates in RNA splicing as a component of the spliceosome. It forms the U5 snRNP by associating with several proteins including Prp8 - the largest and most conserved protein in the spliceosome, Brr2 - a helicase required for spliceosome activation, Snu114, and the 7 Sm proteins. U5 snRNA forms a coaxially-stacked series of helices that project into the active site of the spliceosome. Loop 1, which caps this series of helices, forms 4-5 base pairs with the 5'-exon during the two chemical reactions of splicing. This interaction appears to be especially important during step two of splicing, exon ligation.

<span class="mw-page-title-main">U6 spliceosomal RNA</span>

U6 snRNA is the non-coding small nuclear RNA (snRNA) component of U6 snRNP, an RNA-protein complex that combines with other snRNPs, unmodified pre-mRNA, and various other proteins to assemble a spliceosome, a large RNA-protein molecular complex that catalyzes the excision of introns from pre-mRNA. Splicing, or the removal of introns, is a major aspect of post-transcriptional modification and takes place only in the nucleus of eukaryotes.

<span class="mw-page-title-main">U4atac minor spliceosomal RNA</span>

U4atac minor spliceosomal RNA is a ncRNA which is an essential component of the minor U12-type spliceosome complex. The U12-type spliceosome is required for removal of the rarer class of eukaryotic introns.

<span class="mw-page-title-main">U6atac minor spliceosomal RNA</span>

U6atac minor spliceosomal RNA is a non-coding RNA which is an essential component of the minor U12-type spliceosome complex. The U12-type spliceosome is required for removal of the rarer class of eukaryotic introns.

Splicing factor U2AF 65 kDa subunit is a protein that in humans is encoded by the U2AF2 gene.

Splicing factor 3B subunit 2 is a protein that in humans is encoded by the SF3B2 gene.

RNA, U4atac small nuclear is a small nuclear RNA that in humans is encoded by the RNU4ATAC gene.

Messenger RNP is mRNA with bound proteins. mRNA does not exist "naked" in vivo but is always bound by various proteins while being synthesized, spliced, exported, and translated in the cytoplasm.

Prp8 refers to both the Prp8 protein and Prp8 gene. Prp8's name originates from its involvement in pre-mRNA processing. The Prp8 protein is a large, highly conserved, and unique protein that resides in the catalytic core of the spliceosome and has been found to have a central role in molecular rearrangements that occur there. Prp8 protein is a major central component of the catalytic core in the spliceosome, and the spliceosome is responsible for splicing of precursor mRNA that contains introns and exons. Unexpressed introns are removed by the spliceosome complex in order to create a more concise mRNA transcript. Splicing is just one of many different post-transcriptional modifications that mRNA must undergo before translation. Prp8 has also been hypothesized to be a cofactor in RNA catalysis.

References

Review papers:

Turunen, J. J., Niemelä, E. H., Verma, B., & Frilander, M. J (January–February 2013). "The significant other: splicing by the minor spliceosome". Wiley Interdisciplinary Reviews: RNA. 4 (1): 61–76. doi:10.1002/wrna.1141. PMC 3584512 . PMID 23074130.{{cite journal}}: CS1 maint: multiple names: authors list (link) Review.
Will CL, Lührmann R (August 2005). "Splicing of a rare class of introns by the U12-dependent spliceosome". Biol. Chem. 386 (8): 713–24. doi:10.1515/BC.2005.084. PMID 16201866. S2CID 35468060. Review.

Classic papers:

Jackson IJ (July 25, 1991). "A reappraisal of non-consensus mRNA splice sites". Nucleic Acids Res. 19 (14): 3795–8. doi:10.1093/nar/19.14.3795. PMC 328465 . PMID 1713664.
Hall SL, Padgett RA (1994). "Conserved sequences in a class of rare eukaryotic nuclear introns with non-consensus splice sites". J. Mol. Biol. 239 (3): 357–65. doi:10.1006/jmbi.1994.1377. PMID 8201617.
Tarn WY, Steitz JA (March 8, 1996). "A novel spliceosome containing U11, U12, and U5 snRNPs excises a minor class (AT-AC) intron in vitro". Cell. 84 (5): 801–11. doi: 10.1016/S0092-8674(00)81057-0 . PMID 8625417.
Russell AG, Charette JM, Spencer DF, Gray MW (October 19, 2006). "An early evolutionary origin for the minor spliceosome". Nature. 443 (7113): 863–6. Bibcode:2006Natur.443..863R. doi:10.1038/nature05228. PMID 17051219. S2CID 4419061.

Other references:

↑ Dietrich RC, Incorvaia R, Padgett RA (1997). "Terminal intron dinucleotide sequences do not distinguish between U2- and U12-dependent introns". Molecular Cell. 1 (1): 151–160. doi: 10.1016/S1097-2765(00)80016-7 . PMID 9659912.
↑ Hall SL, Padgett RA (1996). "Requirement of U12 snRNA for in vivo splicing of a minor class of eukaryotic nuclear pre-mRNA introns". Science. 271 (5256): 1716–8. Bibcode:1996Sci...271.1716H. doi:10.1126/science.271.5256.1716. PMID 8596930. S2CID 35875143.
↑ Tarn WY, Steitz JA (1996). "A novel spliceosome containing U11, U12, and U5 snRNPs excises a minor class (AT-AC) intron in vitro". Cell. 84 (5): 801–11. doi: 10.1016/S0092-8674(00)81057-0 . PMID 8625417.
↑ Kolossova I, Padgett RA (1997). "U11 snRNA interacts in vivo with the 5' splice site of U12-dependent (AU-AC) pre-mRNA introns". RNA. 3 (3): 227–33. PMC 1369475 . PMID 9056760.
↑ Yu YT, Steitz JA (1997). "Site-specific crosslinking of mammalian U11 and U6atac to the 5′ splice site of an AT–AC intron". Proc. Natl. Acad. Sci. U.S.A. 94 (12): 6030–5. Bibcode:1997PNAS...94.6030Y. doi: 10.1073/pnas.94.12.6030 . PMC 20995 . PMID 9177163.
↑ Incorvaia R, Padgett RA (1998). "Base pairing with U6atac snRNA is required for 5' splice site activation of U12-dependent introns in vivo". RNA. 4 (6): 709–18. doi:10.1017/S1355838298980207. PMC 1369652 . PMID 9622129.
↑ Tarn WY, Steitz JA (1996). "Highly diverged U4 and U6 small nuclear RNAs required for splicing rare AT-AC introns". Science. 273 (5283): 1824–32. Bibcode:1996Sci...273.1824T. doi:10.1126/science.273.5283.1824. PMID 8791582. S2CID 2638546.
↑ König H, Matter N, Bader R, Thiele W, Müller F (16 Nov 2007). "Splicing segregation: the minor spliceosome acts outside the nucleus and controls cell proliferation". Cell. 131 (4): 1718–29. doi: 10.1016/j.cell.2007.09.043 . PMID 18022366.

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[1] Dietrich RC, Incorvaia R, Padgett RA (1997). "Terminal intron dinucleotide sequences do not distinguish between U2- and U12-dependent introns". Molecular Cell. 1 (1): 151–160. doi: 10.1016/S1097-2765(00)80016-7 . PMID 9659912.

[2] Hall SL, Padgett RA (1996). "Requirement of U12 snRNA for in vivo splicing of a minor class of eukaryotic nuclear pre-mRNA introns". Science. 271 (5256): 1716–8. Bibcode:1996Sci...271.1716H. doi:10.1126/science.271.5256.1716. PMID 8596930. S2CID 35875143.

[3] Tarn WY, Steitz JA (1996). "A novel spliceosome containing U11, U12, and U5 snRNPs excises a minor class (AT-AC) intron in vitro". Cell. 84 (5): 801–11. doi: 10.1016/S0092-8674(00)81057-0 . PMID 8625417.

[4] Kolossova I, Padgett RA (1997). "U11 snRNA interacts in vivo with the 5' splice site of U12-dependent (AU-AC) pre-mRNA introns". RNA. 3 (3): 227–33. PMC 1369475 . PMID 9056760.

[5] Yu YT, Steitz JA (1997). "Site-specific crosslinking of mammalian U11 and U6atac to the 5′ splice site of an AT–AC intron". Proc. Natl. Acad. Sci. U.S.A. 94 (12): 6030–5. Bibcode:1997PNAS...94.6030Y. doi: 10.1073/pnas.94.12.6030 . PMC 20995 . PMID 9177163.

[6] Incorvaia R, Padgett RA (1998). "Base pairing with U6atac snRNA is required for 5' splice site activation of U12-dependent introns in vivo". RNA. 4 (6): 709–18. doi:10.1017/S1355838298980207. PMC 1369652 . PMID 9622129.

[7] Tarn WY, Steitz JA (1996). "Highly diverged U4 and U6 small nuclear RNAs required for splicing rare AT-AC introns". Science. 273 (5283): 1824–32. Bibcode:1996Sci...273.1824T. doi:10.1126/science.273.5283.1824. PMID 8791582. S2CID 2638546.

[8] König H, Matter N, Bader R, Thiele W, Müller F (16 Nov 2007). "Splicing segregation: the minor spliceosome acts outside the nucleus and controls cell proliferation". Cell. 131 (4): 1718–29. doi: 10.1016/j.cell.2007.09.043 . PMID 18022366.

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