Prp24: Difference between revisions

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The main function of Prp24 seems to be directly related to formation of the U4/U6 complex, particularly based on the evidence that Prp24 is present in U6 and U4/U6, but not U4/U6.U5 (reference Shannon and Guthrie 1991, Ghetti et al. 1995, and Jandrositz and Guthrei 1995). Prp24 greatly increases the rate and efficiency of U4/U6 annealing (Raghunathan and Guthrie 1998) and mutations in Prp24 have been shown to prevent the formation of the U4/U6 di-snRNP (Lygerou et al. 1999). Although the exact mechanism by which Prp24 promotes annealing of U4 and U6, it has been suggested that Prp24 may stabilize the secondary structure of U6 to allow it to interact with U4 in order to allow formation of U4/U6 (reference Vidaver et al. 1999)

The interactions of Prp24 with U6 and U4/U6 are very important in proper spliceosome assembly, as evidenced by mutations in Prp24 that suppress mutations in U6 or U4. Mutations in RRM 2 and RRM 3 suppress the effects of mutations in the 3' stem of U6, in a region termed the telestem (reference Vidaver et al. 1999) (the existence of the telestem has been challenged by recent U6 secondary structure models (reference Karaduman et al. 2008 and Dunn and Rader 2010)). A later study showed that RRM 1 binds with high affinity to U6 and is important in interactions with mutations in the 3' region of U6 (reference Kwan and Brow 2005). Mutations in two RNP consensus domains of Prp24 suppress the effects of mutations in U4 in the stem II region of U4/U6 (reference Shannon and Guthrie 1991). Truncation and deletion of internal segments of U6 showed that the central region of U6 is the most important region for interaction with Prp24 and that RRM 1 and RRM 2 are the likely portions of Prp24 that interact with U6 in this region (reference Kwan and Brow 2005).

The interactions of Prp24 with U6 and U4/U6 are very important in proper spliceosome assembly, as evidenced by mutations in Prp24 that suppress mutations in U6 or U4. Mutations in RRM 2 and RRM 3 suppress the effects of mutations in the 3' stem of U6, in a region termed the telestem (reference Vidaver et al. 1999) (the existence of the telestem has been challenged by recent U6 secondary structure models (reference Karaduman et al. 2008 and Dunn and Rader 2010)). A later study showed that RRM 1 binds with high affinity to U6 and is important in interactions with mutations in the 3' region of U6 (reference Kwan and Brow 2005). Mutations in two RNP consensus domains of Prp24 suppress the effects of mutations in U4 in the stem II region of U4/U6 (reference Shannon and Guthrie 1991). Truncation and deletion of internal segments of U6 showed that the central region of U6 is the most important region for interaction with Prp24 and that RRM 1 and RRM 2 are the likely portions of Prp24 that interact with U6 in this region (reference Kwan and Brow 2005). Deletion of conserved residues in the C-terminal domain of Prp24 caused temperature sensitive growth and reduced levels of U4/U6, suggesting that this region too, containing the fourth RRM, is important for interactions in assembly of the spliceosome (Rader and Guthrie 2002).

NMR analysis of fragments of U6 sequence and regions of Prp24 have further defined and supported the interactions of the protein with U6 snRNA. Analysis of an RNA oligonucleotide containing sequences identical to nucleotides 41-46 and 83-88 of U6 (the regions proposed to bind to Prp24) with a truncated protein containing RRMs 1 and 2 of Prp24 showed that these RRMs interacted sequence specifically with these nucleotides (Bae et al. 2007). Another NMR study identified sequence specific interaction of RRM 2 with RNA, and it was determined from this that most likely region of U6 for interaction was the AGAGAU sequence of nucleotides 49-54, within the region of previously predicted interaction (Martin-Tumasz et al. 2010). From this, it was predicted that RRM 1 would interact with the GAUCAG sequence of nucleotides 55-60 (Martin-Tumasz et al. 2010).

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The Lsm proteins are a set of seven proteins found in the U6 snRNP that form a ring around the 3' end of U6 (reference _____). An interaction between Prp24 and the Lsm proteins was suggested upon identification of the Lsms and the fact that formation of U4/U6 by Prp24 was less efficient in deproteinized solutions of U4 and U6 snRNAs (reference Raghunathan and Guthrie), as well as the identification of a genetic interaction between the genes for Prp24 and Lsm4 (Mayes et al. 1999). This proposed interaction was further supported by UV cross-linking experiments in which both Prp24 and Lsm4 were immunoprecipitated with U6 after cross-linking (reference Vidal et al. 1999). It has also been shown that both Prp24 and Lsm4, as well as the other Lsm proteins, require the 3' end of U6 in order to interact with the snRNA (Vidal et al. 1999), suggesting that this may be the region where the proteins interact. Genome-wide protein interaction screens showed that Prp24 interacts additionally with Lsm 2, 5, 6, 7 and 8, and that overexpression of either Prp24 or Lsm4 can complement or intensify the effects of a defect in the other protein (Fromont-Racine et al. 2000), further supporting an interaction between the Lsm proteins and Prp24.

Evidence for a direct interaction of Prp24 and the Lsm proteins comes from a study in which the conserved residues in the C-terminal domain of Prp24 were deleted, resulting in lowered levels of U4/U6 and very literal interaction with the Lsm proteins as compared to wild-type Prp24, indicating that Prp24 interacts directly with the Lsm proteins and the C-terminal domain is necessary for this interaction (Rader and Guthrie 2002).

===Additional Interactions===

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@@ Line 54: / Line 54: @@
 The main function of Prp24 seems to be directly related to formation of the U4/U6 complex, particularly based on the evidence that Prp24 is present in U6 and U4/U6, but not U4/U6.U5 (reference Shannon and Guthrie 1991, Ghetti et al. 1995, and Jandrositz and Guthrei 1995).  Prp24 greatly increases the rate and efficiency of U4/U6 annealing (Raghunathan and Guthrie 1998) and mutations in Prp24 have been shown to prevent the formation of the U4/U6 di-snRNP (Lygerou et al. 1999).  Although the exact mechanism by which Prp24 promotes annealing of U4 and U6, it has been suggested that Prp24 may stabilize the secondary structure of U6 to allow it to interact with U4 in order to allow formation of U4/U6 (reference Vidaver et al. 1999)
-The interactions of Prp24 with U6 and U4/U6 are very important in proper spliceosome assembly, as evidenced by mutations in Prp24 that suppress mutations in U6 or U4.  Mutations in RRM 2 and RRM 3 suppress the effects of mutations in the 3' stem of U6, in a region termed the telestem (reference Vidaver et al. 1999) (the existence of the telestem has been challenged by recent U6 secondary structure models (reference Karaduman et al. 2008 and Dunn and Rader 2010)).  A later study showed that RRM 1 binds with high affinity to U6 and is important in interactions with mutations in the 3' region of U6 (reference Kwan and Brow 2005).  Mutations in two RNP consensus domains of Prp24 suppress the effects of mutations in U4 in the stem II region of U4/U6 (reference Shannon and Guthrie 1991).  Truncation and deletion of internal segments of U6 showed that the central region of U6 is the most important region for interaction with Prp24 and that RRM 1 and RRM 2 are the likely portions of Prp24 that interact with U6 in this region (reference Kwan and Brow 2005).
+The interactions of Prp24 with U6 and U4/U6 are very important in proper spliceosome assembly, as evidenced by mutations in Prp24 that suppress mutations in U6 or U4.  Mutations in RRM 2 and RRM 3 suppress the effects of mutations in the 3' stem of U6, in a region termed the telestem (reference Vidaver et al. 1999) (the existence of the telestem has been challenged by recent U6 secondary structure models (reference Karaduman et al. 2008 and Dunn and Rader 2010)).  A later study showed that RRM 1 binds with high affinity to U6 and is important in interactions with mutations in the 3' region of U6 (reference Kwan and Brow 2005).  Mutations in two RNP consensus domains of Prp24 suppress the effects of mutations in U4 in the stem II region of U4/U6 (reference Shannon and Guthrie 1991).  Truncation and deletion of internal segments of U6 showed that the central region of U6 is the most important region for interaction with Prp24 and that RRM 1 and RRM 2 are the likely portions of Prp24 that interact with U6 in this region (reference Kwan and Brow 2005).  Deletion of conserved residues in the C-terminal domain of Prp24 caused temperature sensitive growth and reduced levels of U4/U6, suggesting that this region too, containing the fourth RRM, is important for interactions in assembly of the spliceosome (Rader and Guthrie 2002).
 NMR analysis of fragments of U6 sequence and regions of Prp24 have further defined and supported the interactions of the protein with U6 snRNA.  Analysis of an RNA oligonucleotide containing sequences identical to nucleotides 41-46 and 83-88 of U6 (the regions proposed to bind to Prp24) with a truncated protein containing RRMs 1 and 2 of Prp24 showed that these RRMs interacted sequence specifically with these nucleotides (Bae et al. 2007).  Another NMR study identified sequence specific interaction of RRM 2 with RNA, and it was determined from this that most likely region of U6 for interaction was the AGAGAU sequence of nucleotides 49-54, within the region of previously predicted interaction (Martin-Tumasz et al. 2010).  From this, it was predicted that RRM 1 would interact with the GAUCAG sequence of nucleotides 55-60 (Martin-Tumasz et al. 2010).
@@ Line 61: / Line 61: @@
 The Lsm proteins are a set of seven proteins found in the U6 snRNP that form a ring around the 3' end of U6 (reference _____).  An interaction between Prp24 and the Lsm proteins was suggested upon identification of the Lsms and the fact that formation of U4/U6 by Prp24 was less efficient in deproteinized solutions of U4 and U6 snRNAs (reference Raghunathan and Guthrie), as well as the identification of a genetic interaction between the genes for Prp24 and Lsm4 (Mayes et al. 1999).  This proposed interaction was further supported by UV cross-linking experiments in which both Prp24 and Lsm4 were immunoprecipitated with U6 after cross-linking (reference Vidal et al. 1999).  It has also been shown that both Prp24 and Lsm4, as well as the other Lsm proteins, require the 3' end of U6 in order to interact with the snRNA (Vidal et al. 1999), suggesting that this may be the region where the proteins interact.  Genome-wide protein interaction screens showed that Prp24 interacts additionally with Lsm 2, 5, 6, 7 and 8, and that overexpression of either Prp24 or Lsm4 can complement or intensify the effects of a defect in the other protein (Fromont-Racine et al. 2000), further supporting an interaction between the Lsm proteins and Prp24.
+Evidence for a direct interaction of Prp24 and the Lsm proteins comes from a study in which the conserved residues in the C-terminal domain of Prp24 were deleted, resulting in lowered levels of U4/U6 and very literal interaction with the Lsm proteins as compared to wild-type Prp24, indicating that Prp24 interacts directly with the Lsm proteins and the C-terminal domain is necessary for this interaction (Rader and Guthrie 2002).
 ===Additional Interactions===