Prp24

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Prp24 (precursor RNA processing, gene 24) is a protein part of the pre-messenger RNA splicing process and aids the binding of U6 snRNA to U4 snRNA during the formation of spliceosomes. Found in eukaryotes from yeast to E. coli , fungi, and humans, Prp24 was initially discovered to be an important element of RNA splicing in 1989. [1] [2] Mutations in Prp24 were later discovered in 1991 to suppress mutations in U4 that resulted in cold-sensitive strains of yeast, indicating its involvement in the reformation of the U4/U6 duplex after the catalytic steps of splicing. [3]

Contents

Prp24 RRMs 1 and 2 Prp24 domains 1 and 2 image.jpg
Prp24 RRMs 1 and 2

Biological Role

The process of spliceosome formation involves the U4 and U6 snRNPs associating and forming a di-snRNP in the cell nucleus. This di-snRNP then recruits another member (U5) to become a tri-snRNP. U6 must then dissociate from U4 to bond with U2 and become catalytically active. Once splicing has been done, U6 must dissociate from the spliceosome and bond back with U4 to restart the cycle.

Prp24 has been shown to promote the binding of U4 and U6 snRNPs. Removing Prp24 results in the accumulation of free U4 and U6, and the subsequent addition of Prp24 regenerates U4/U6 and reduces the amount of free U4 and U6. [4] Naked U6 snRNA is very compact and has little room to form base pairs with other RNA. However, when U6 snRNP associates with proteins such as Prp24, the structure is much more open, thus facilitating the binding to U4. [5] Prp24 is not present in the U6/U4 duplex itself, and it has been suggested that Prp24 must leave the complex in order for proper base pairs to be formed. [6] [7] It has also been suggested that Prp24 may play a role in destabilizing U4/U6 in order for U6 to pair bases with U2. [8]

Structure

Prp 24 RRM 3 Prp24 domain 3.jpg
Prp 24 RRM 3

Prp24 has a molecular weight of 50 kDa and has been shown to contain four RNA recognition motifs (RRMs) and a conserved 12-amino acid sequence at the C-terminus. [9] [10] RRMs 1 and 2 have been shown to be important for high-affinity binding of U6, while RRMs 3 and 4 bind at lower affinity sites on U6. [11] The first three RRMs interact extensively with each other and contain canonical folds that contain a four-stranded beta-sheet and two alpha-helices. The electropositive surface of RRMs 1 and 2 is a RNA annealing domain while the cleft between RRMs 1 and 2 including the beta-sheet face of RRM2 is a sequence-specific RNA binding site. [1] The C-terminal motif is required for association with LSm proteins and contributes to substrate (U6) binding and not the catalytic rate of splicing. [10]

U6 snRNP with the associated Prp24 and LSm proteins U6 snRNP with Prp24 and Lsm.jpg
U6 snRNP with the associated Prp24 and LSm proteins

Interactions

Prp24 interacts with the U6 snRNA via its RRMs. It has been shown through chemical modification testing that nucleotides 39–57 of U6 (40–43 in particular) [5] are involved in binding Prp24. [12]

The LSm proteins are in a consistent configuration on the U6 RNA. [9] [ clarification needed ] It has been proposed that the LSm proteins and Prp24 interact both physically and functionally [6] and the C-terminal motif of Prp24 is important for this interaction. [10] The binding of Prp24 to U6 is enhanced by the binding of Lsm proteins to U6, as is binding of U4 and U6. [13] It was revealed by electron microscopy that Prp24 may interact with the LSm protein ring at LSm2. [9]

Homologs

Prp24 has a human homolog, SART3. SART3 is a tumor rejection antigen (SART3 stands for "squamous cell carcinoma antigen recognized by T cells, gene 3). The RRMs 1 and 2 in yeast are similar to RRMs in human SART3. [1] [11] The C-terminal domain is also highly conserved from yeast to humans. [14] This protein, like Prp24, interacts with the LSm proteins [9] [15] for the recycling of U6 into the U4/U6 snRNP. It has been proposed that SART3 target U6 to a Cajal body or a nuclear inclusion as the site of assembly of the U4/U6 snRNP. [15] SART3 is located on chromosome 12, and a mutation is likely the cause of disseminated superficial actinic porokeratosis. [16]

Related Research Articles

RNA splicing Processing primary RNA to remove intron sequences and join the remaining exon sections

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 introns and so joining 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.

Spliceosome Molecular machine that removes intron RNA from the primary transcript

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 protein

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.

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.

LSm

In molecular biology, LSm proteins are a family of RNA-binding proteins found in virtually every cellular organism. LSm is a contraction of 'like Sm', because the first identified members of the LSm protein family were the Sm proteins. LSm proteins are defined by a characteristic three-dimensional structure and their assembly into rings of six or seven individual LSm protein molecules, and play a large number of various roles in mRNA processing and regulation.

U2 spliceosomal RNA

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.

U4 spliceosomal RNA Non-coding RNA component of the spliceosome

The U4 small nuclear Ribo-Nucleic Acid is a non-coding RNA component of the major or U2-dependent spliceosome – a eukaryotic molecular machine involved in the splicing of pre-messenger RNA (pre-mRNA). It forms a duplex with U6, and with each splicing round, it is displaced from the U6 snRNA in an ATP-dependent manner, allowing U6 to re-fold and create the active site for splicing catalysis. A recycling process involving protein Brr2 releases U4 from U6, while protein Prp24 re-anneals U4 and U6. The crystal structure of a 5′ stem-loop of U4 in complex with a binding protein has been solved.

U5 spliceosomal RNA

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.

U6 spliceosomal RNA

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.

U2AF2

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

PRPF8

Pre-mRNA-processing-splicing factor 8 is a protein that in humans is encoded by the PRPF8 gene.

DEAD box

DEAD box proteins are involved in an assortment of metabolic processes that typically involve RNAs, but in some cases also other nucleic acids. They are highly conserved in nine motifs and can be found in most prokaryotes and eukaryotes, but not all. Many organisms, including humans, contain DEAD-box (SF2) helicases, which are involved in RNA metabolism.

SF3A1

Splicing factor 3 subunit 1 is a protein that in humans is encoded by the SF3A1 gene.

SF3A2

Splicing factor 3A subunit 2 is a protein that in humans is encoded by the SF3A2 gene.

PRPF6

Pre-mRNA-processing factor 6 is a protein that in humans is encoded by the PRPF6 gene.

PRPF4

U4/U6 small nuclear ribonucleoprotein Prp4 is a protein that in humans is encoded by the PRPF4 gene. The removal of introns from nuclear pre-mRNAs occurs on complexes called spliceosomes, which are made up of 4 small nuclear ribonucleoprotein (snRNP) particles and an undefined number of transiently associated splicing factors. PRPF4 is 1 of several proteins that associate with U4 and U6 snRNPs.[supplied by OMIM]

RNA recognition motif

RNA recognition motif, RNP-1 is a putative RNA-binding domain of about 90 amino acids that are known to bind single-stranded RNAs. It was found in many eukaryotic proteins.

Prp8

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.

Christine Guthrie is an American yeast geneticist and American Cancer Society Research Professor of Genetics at University of California San Francisco. She showed that yeast have small nuclear RNAs (snRNAs) involved in splicing pre-messenger RNA into messenger RNA in eukaryotic cells. Guthrie cloned and sequenced the genes for yeast snRNA and established the role of base pairing between the snRNAs and their target sequences at each step in the removal of an intron. She also identified proteins that formed part of the spliceosome complex with the snRNAs. Elected to the National Academy of Sciences in 1993, Guthrie edited Guide to Yeast Genetics and Molecular Biology, an influential methods series for many years.

Kiyoshi Nagai Japanese structural biologist

Kiyoshi Nagai was a Japanese structural biologist at the MRC Laboratory of Molecular Biology Cambridge, UK. He was known for his work on the mechanism of RNA splicing and structures of the spliceosome.

References

  1. 1 2 3 Bae, E.; et al. (2007). "Structure and interactions of the first three RNA recognition motifs of splicing factor Prp24". Journal of Molecular Biology. 367 (5): 1447–1458. doi:10.1016/j.jmb.2007.01.078. PMC   1939982 . PMID   17320109.
  2. ↑ Vijayraghavan, U.; Company, M.; Abelson, J. (1989). "Isolation and characterization of pre-mRNA splicing mutants of Saccharomyces cerevisiae". Genes & Development. 3 (8): 1206–1216. doi: 10.1101/gad.3.8.1206 . PMID   2676722.
  3. ↑ Shannon, K. W.; Guthrie, C. (1991). "Suppressors of U4 snRNA mutation define a novel U6 snRNP protein with RNA-binding motifs". Genes & Development. 5 (5): 773–785. doi: 10.1101/gad.5.5.773 . PMID   1827420.
  4. ↑ Raghunathan, P. L.; Guthrie, C. A. (1998). "Spliceosomal Recycling Factor That Reanneals U4 and U6 Small Nuclear Ribonucleoprotein Particles". Science . 279 (5352): 857–860. Bibcode:1998Sci...279..857R. doi:10.1126/science.279.5352.857. PMID   9452384.
  5. 1 2 Karaduman, R.; et al. (2006). "RNA Structure and RNA–Protein Interactions in Purified Yeast U6 snRNPs". Journal of Molecular Biology. 356 (5): 1248–1262. doi:10.1016/j.jmb.2005.12.013. hdl: 11858/00-001M-0000-0012-E5F7-6 . PMID   16410014.
  6. 1 2 Vidal, V. P.; et al. (1995). "Characterization of U6 snRNA-protein interactions". RNA. 5 (11): 1470–1481. doi:10.1017/S1355838299991355. PMC   1369868 . PMID   10580475.
  7. ↑ Jandrositz, A.; Guthrie, A. (1995). "Evidence for a Prp24 binding site in U6 snRNA and in a putative intermediate in the annealing of U6 and U4 snRNAs". The EMBO Journal. 14 (4): 820–832. doi:10.1002/j.1460-2075.1995.tb07060.x. PMC   398149 . PMID   7882985.
  8. ↑ Vidaver, R. M.; Fortner, DM; Loos-Austin, LS; Brow, DA (1999). "Multiple functions of Saccharomyces cerevisiae splicing protein Prp24 in U6 RNA structural rearrangements". Genetics. 153 (3): 1205–1218. doi:10.1093/genetics/153.3.1205. PMC   1460831 . PMID   10545453.
  9. 1 2 3 4 Karaduman, R.; et al. (2008). "Structure of yeast U6 snRNPs: Arrangement of Prp24p and the LSm complex as revealed by electron microscopy". RNA. 14 (12): 1–10. doi:10.1261/rna.1369808. PMC   2590955 . PMID   18971323.
  10. 1 2 3 Rader, S. D.; Guthrie, C. (2002). "A conserved Lsm-interaction motif in Prp24 required for efficient U4/U6 di-snRNP formation". RNA. 8 (11): 1378–1392. doi:10.1017/S1355838202020010. PMC   1370345 . PMID   12458792.
  11. 1 2 Kwan, S. S.; Brow, D. A. (2005). "The N- and C-terminal RNA recognition motifs of splicing factor Prp24 have distinct functions in U6 RNA binding". RNA. 11 (5): 808–820. doi:10.1261/rna.2010905. PMC   1370765 . PMID   15811912.
  12. ↑ Ghetti, A.; Company, M.; Abelson, J. (1995). "Specificity of Prp24 binding to RNA: a role for Prp24 in the dynamic interaction of U4 and U6 snRNAs". RNA. 1 (2): 132–145. PMC   1369067 . PMID   7585243.
  13. ↑ Ryan, D. E.; Stevens, S. W.; Abelson, J. (2002). "The 5' and 3' domains of yeast U6 snRNA: Lsm proteins facilitate binding of Prp24 protein to the U6 telestem region". RNA. 8 (8): 1011–1033. doi:10.1017/S1355838202026092. PMC   1370313 . PMID   12212846.
  14. ↑ Bell, M.; et al. (2002). "p110, a novel human U6 snRNP protein and U4/U6 snRNP recycling factor". The EMBO Journal. 21 (11): 2724–2735. doi:10.1093/emboj/21.11.2724. PMC   126028 . PMID   12032085.
  15. 1 2 Stanek, D.; et al. (2003). "Targeting of U4/U6 small nuclear RNP assembly factor SART3/p110 to Cajal bodies". Journal of Cell Biology. 160 (4): 505–516. doi:10.1083/jcb.200210087. PMC   2173746 . PMID   12578909.
  16. ↑ "OMIM - POROKERATOSIS, DISSEMINATED SUPERFICIAL ACTINIC, 1; DSAP1" . Retrieved 2009-04-05.
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