The human La (SS-B) autoantigen interacts with DDX15/hPrp43, a putative DEAH-box RNA helicase

Fouraux, Michael A.; Kolkman, Marloes J M; Heijden, Annemarie van der; Jong, Arjan S. de; Venrooij, Walther J. van; Pruijn, Ger J. M.

doi:10.1017/s1355838202021076

Cited by 46 publications

(32 citation statements)

References 69 publications

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“…The putative human orthologue for the yeast Prp43p is DDX15/ hPrp43p, which has been shown to be associated with the pre-mRNA splicing machinery in HeLa extracts (17) and has been localized to nuclear speckles (a site of concentrated premRNA splicing factors) as well as to human nucleoli (11). Although this is not evidence for hPrp43p function in human FIG.…”

Section: Discussionmentioning

confidence: 93%

Prp43p Is a DEAH-Box Spliceosome Disassembly Factor Essential for Ribosome Biogenesis

Combs

Nagel

Ares

et al. 2006

Molecular and Cellular Biology

109

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The known function of the DEXH/D-box protein Prp43p is the removal of the U2, U5, and U6 snRNPs from the postsplicing lariat-intron ribonucleoprotein complex. We demonstrate that affinity-purified Prp43p-associated material includes the expected spliceosomal components; however, we also identify several preribosomal complexes that are specifically purified with Prp43p. Conditional prp43 mutant alleles confer a 35S pre-rRNA processing defect, with subsequent depletion of 27S and 20S precursors. Upon a shift to a nonpermissive temperature, both large and small-ribosomal-subunit proteins accumulate in the nucleolus of prp43 mutants. Pulse-chase analysis demonstrates delayed kinetics of 35S, 27S, and 20S pre-rRNA processing with turnover of these intermediates. Microarray analysis of pre-mRNA splicing defects in prp43 mutants shows a very mild effect, similar to that of nonessential pre-mRNA splicing factors. Prp43p is the first DEXH/D-box protein shown to function in both RNA polymerase I and polymerase II transcript metabolism. Its essential function is in its newly characterized role in ribosome biogenesis of both ribosomal subunits, positioning Prp43p to regulate both pre-mRNA splicing and ribosome biogenesis.DEXH/D-box proteins comprise a family of RNA helicase and RNA helicase-like proteins that function in almost all aspects of eukaryotic gene expression. Included in these processes is RNA transcription (30, 49), pre-mRNA splicing (reviewed in reference 35), mRNA export (21, 34, 38), translation initiation (40), RNA interference (29, 41), RNA turnover (1), viral replication (31), and ribosome biogenesis (reviewed in references 13 and 44). Among this class of RNA helicases are the DEAD-box proteins and the DEAH-box proteins, which differ by subtle sequence variations in one or more of the motifs that define these proteins (reviewed in reference 42). In addition to an RNA-unwinding activity demonstrated by some of these enzymes, others exhibit an "RNPase" function that serves to displace proteins from or otherwise rearrange ribonucleoproteins (10,20). Although the specific function and mechanism of most DEXH/D-box proteins are unknown, the processes in which many of these enzymes function have been studied in great detail.The eukaryotic pre-mRNA splicing reaction is an ATP-intensive process in which at least eight DEXH/D-box proteins function (35). The genes for all eight Saccharomyces cerevisiae spliceosomal DEXH/D-box proteins are essential, although two have been shown to be dispensable in concert with bypass suppressor mutations (6, 23). DEXH/D-box proteins are required for virtually every known step of the pre-mRNA splicing reaction and are thought to monitor and modify RNA-RNA interactions and RNA-protein interactions in the highly dynamic process.Among the trans-acting spliceosomal RNA helicase-like proteins is Prp43p, which was originally identified in a PCR-based screen for RNA helicase genes in yeast (2, 9). It has been shown to be a bona fide RNA-dependent ATPase, although no RNA-unwinding activity h...

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Section: Discussionmentioning

confidence: 93%

Prp43p Is a DEAH-Box Spliceosome Disassembly Factor Essential for Ribosome Biogenesis

Combs

Nagel

Ares

et al. 2006

Molecular and Cellular Biology

109

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“…It is also possible that RNA unwinding of stem-loop IV facilitates the movement of initiation complex. In fact, it has been shown earlier that La protein interacts with DDX15/ hPrp43, which is a putative DEAH-box RNA helicase (41). Furthermore, HCV RNA toe-printing analysis in the presence of increasing concentrations of La protein revealed a decrease in certain toe-prints corresponding to the RNA structure, which suggests that La protein might help in melting some secondary structures and facilitate the extension of the primer by the reverse transcriptase (Ref.…”

Section: Discussionmentioning

confidence: 94%

La Protein Binding at the GCAC Site Near the Initiator AUG Facilitates the Ribosomal Assembly on the Hepatitis C Virus RNA to Influence Internal Ribosome Entry Site-mediated Translation

Pudi¹,

Srinivasan

Das

2004

Journal of Biological Chemistry

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Translation of hepatitis C virus (HCV) 1 is mediated by an internal ribosome entry site (IRES) located mostly within the 5Ј-untranslated region (UTR) and extending a few nucleotides into the open reading frame (1-5). HCV 5Ј-UTR is highly conserved and folds into a complex secondary structure comprising four major structural domains (I-IV) and a pseudoknot in the vicinity of the initiator AUG codon (6, 7). The cis-elements promote assembly of initiation complex independent of the 5Ј-end and thus mediate internal initiation of translation in a cap-independent manner (7). Domain I is not a part of IRES and most likely is involved in RNA replication, whereas domains II and III are complex and consist of multiple stem-loops and bulge-loops. Even minor mutations in domains II and III substantially reduce IRES activity, but this could in most cases be regained by compensatory second site mutations that restored secondary structure. Highly conserved residues are often unpaired and may thus be able to interact with the components of the translation apparatus. Domain IV consists of a stem-loop that contains initiator AUG codon and has been shown to play a key role in regulating the initiation of translation of the HCV RNA (5, 8 -10). It appears from earlier reports that both the sequence and stability of the domain IV stem might control efficiency of HCV IRES translation (11, 12). These observations have led to a model for IRES function in which the structural elements in the IRES act as a scaffold that orients the potential binding sites in such a way that their interactions with initiation factors and ribosomes lead to assembly of functional ribosomal initiation complexes (13).HCV IRES binds to the 40 S ribosomal subunit specifically and stably even in the absence of any initiation factors. Addition of eIF2/GTP/Met-tRNAi is sufficient for 40 S subunit to lock onto initiator AUG (13). eIF3, though not essential for the formation of 48 S complex formation, it has been shown to bind to the apical half of domain III and is likely to be a constituent of the 48 S-IRES complex in vivo (14,15). 48 S complex formation on HCV IRES has no requirement for eIF4A, 4B, 4E, 4G, or for ATP hydrolysis (14 -16). Because the viral 5Ј-UTR forms a binary complex with the 40 S ribosomal subunit in the absence of any canonical or non-canonical initiation factors, it is likely that the additional factors may stimulate internal initiation of translation following the assembly of RNA-40 S complex. Recently, binding of a 25-kDa cellular protein (p25) to HCV IRES has been shown to be important for the efficient translation initiation. p25 was originally suggested to be ribosomal protein S9 but later identified as rpS5 (14,17,18). In fact, HCV IRES has been suggested to have a prokaryotic-like mode of interaction with the 40 S ribosomal subunit, where the 40 S ribosomal subunit is thought to interact with the HCV-IRES through p25 (14). However, eukaryotic mRNAs and picornaviral IRESs have not been reported to require S5 protein for the ribosome asse...

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“…Since Prp43p has been reported to be a nucleolar protein (26,45) and was detected among the proteins interacting with Npa1p (17), a specific component of early pre-60S preribosomal particles, we reasoned that it might be involved in ribosome biogenesis. To assess the consequences of lack of Prp43p on RNA metabolism, we constructed a yeast strain that contains a PRP43 gene transcribed from the GAL1-10-CYC1 hybrid promoter.…”

Section: Resultsmentioning

confidence: 99%

The Splicing ATPase Prp43p Is a Component of Multiple Preribosomal Particles

Lebaron

Froment

Fromont‐Racine

et al. 2005

Molecular and Cellular Biology

116

168

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Prp43p is a putative helicase of the DEAH family which is required for the release of the lariat intron from the spliceosome. Prp43p could also play a role in ribosome synthesis, since it accumulates in the nucleolus. Consistent with this hypothesis, we find that depletion of Prp43p leads to accumulation of 35S pre-rRNA and strongly reduces levels of all downstream pre-rRNA processing intermediates. As a result, the steady-state levels of mature rRNAs are greatly diminished following Prp43p depletion. We present data arguing that such effects are unlikely to be solely due to splicing defects. Moreover, we demonstrate by a combination of a comprehensive two-hybrid screen, tandem-affinity purification followed by mass spectrometry, and Northern analyses that Prp43p is associated with 90S, pre-60S, and pre-40S ribosomal particles. Prp43p seems preferentially associated with Pfa1p, a novel specific component of pre-40S ribosomal particles. In addition, Prp43p interacts with components of the RNA polymerase I (Pol I) transcription machinery and with mature 18S and 25S rRNAs. Hence, Prp43p might be delivered to nascent 90S ribosomal particles during pre-rRNA transcription and remain associated with preribosomal particles until their final maturation steps in the cytoplasm. Our data also suggest that the ATPase activity of Prp43p is required for early steps of pre-rRNA processing and normal accumulation of mature rRNAs.Synthesis of eukaryotic ribosomes is an intricate process that is initiated by the synthesis of a precursor to 5S rRNA by RNA polymerase III and of a common precursor to 18S, 5.8S, and 25S rRNAs by RNA polymerase I (70, 93). Some nucleotides present in the sequences retained in the mature rRNAs are modified by site-specific enzymes or more generally by small nucleolar ribonucleoprotein particles (snoRNPs) during or immediately following synthesis of the primary pre-rRNA transcripts (50). These are then processed by a complex series of endonucleolytic and exonucleolytic digestion steps that remove so-called spacer regions to yield the mature rRNAs. These pre-rRNA processing events do not occur on naked pre-rRNAs but within preribosomal particles (96). These particles result from the association of pre-rRNAs at various stages of maturation with ribosomal proteins, which remain in the cytoplasmic ribosomes, as well as scores of nonribosomal proteins that only transiently interact with pre-rRNAs (25, 28). As they mature, preribosomal particles transit from the nucleolus to the nucleoplasm and are then transported to the cytoplasm where final maturation steps take place (88). With advancing maturation, the number of nonribosomal proteins present in preribosomal particles declines (2,36,63). Well over 170 nonribosomal proteins linked to ribosome biogenesis have been described previously, and the list is probably far from complete (28).Apart from the enzymes shown to modify rRNA nucleotides or to catalyze endo-or exonucleolytic digestion steps, the precise molecular functions of most nonribosomal proteins remai...

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The human La (SS-B) autoantigen interacts with DDX15/hPrp43, a putative DEAH-box RNA helicase

Cited by 46 publications

References 69 publications

Prp43p Is a DEAH-Box Spliceosome Disassembly Factor Essential for Ribosome Biogenesis

Prp43p Is a DEAH-Box Spliceosome Disassembly Factor Essential for Ribosome Biogenesis

La Protein Binding at the GCAC Site Near the Initiator AUG Facilitates the Ribosomal Assembly on the Hepatitis C Virus RNA to Influence Internal Ribosome Entry Site-mediated Translation

The Splicing ATPase Prp43p Is a Component of Multiple Preribosomal Particles

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