Two Trypanosome-Specific Proteins Are Essential Factors for 5S rRNA Abundance and Ribosomal Assembly in
            <i>Trypanosoma brucei</i>

Hellman, Kristina; Ciganda, Martı́n; Brown, Silvia V.; Ruyechan, William T.; Williams, Noreen

doi:10.1128/ec.00119-07

Cited by 22 publications

(62 citation statements)

References 36 publications

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“…We have previously established that p34 and p37 associate with 5S rRNA (31), a component of the 60S ribosomal subunit. Recent work from this laboratory using the same p34/p37 RNAi cells has demonstrated a dramatic decrease in the levels of 5S rRNA, in addition to disruption of a cellular complex(es) containing 5S rRNA (19). These results suggest an important role for p34 and p37 in stabilizing 5S rRNA.…”

Section: Discussionmentioning

confidence: 80%

“…RNA interference of the p34 and p37 proteins was employed in order to further characterize this interaction. We have previously established a clonal cell line which, upon addition of tetracycline, induces production of dsRNA specific to p34 and p37 (19). This leads to degradation of p34 and p37 mRNA and a phenotypic absence of these proteins, which were shown to be essential for survival of the parasite.…”

Section: Resultsmentioning

confidence: 99%

“…We have previously identified two nearly identical RNA binding proteins, p34 and p37, which are essential for Trypanosoma brucei survival (19,40). These trypanosome-specific proteins (18) interact with NOPP44/46 (30), a family of trypanosome-specific nucleolar phosphoproteins (29), suggesting that this protein-protein interaction serves a unique function within T. brucei.…”

Section: Discussionmentioning

confidence: 99%

“…These results suggest an important role for p34 and p37 in stabilizing 5S rRNA. The loss of 5S rRNA in these cells leads to a significant decrease in protein synthesis efficiency, which appears to be the result of a defect in the assembly the 60S ribosomal subunits into the 80S complex (19). At this time, we do not know whether the disrupted complex(es) in the p34/p37 RNAi cells contains both NOPP44/46 and 5S rRNA or whether they are separate and distinct complexes.…”

Section: Discussionmentioning

confidence: 99%

“…Here, we employ a T. brucei cell line expressing RNA interference (RNAi) that specifically targets both p34 and p37 (19) to further examine the cellular role of the interaction between p34 and p37 and the NOPP44/46 proteins. fractions was precipitated with trichloroacetic acid (TCA), resolved on a 12% sodium dodecyl sulfate (SDS)-polyacrylamide gel, and transferred to a nitrocellulose membrane (Schleicher & Schuell).…”

mentioning

confidence: 99%

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Trypanosoma brucei RNA Binding Proteins p34 and p37 Mediate NOPP44/46 Cellular Localization via the Exportin 1 Nuclear Export Pathway

2007

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We have previously identified and characterized two novel nuclear RNA binding proteins, p34 and p37, which have been shown to interact with a family of nucleolar phosphoproteins, NOPP44/46, in Trypanosoma brucei. These proteins are nearly identical, the major difference being an 18-amino-acid insert in the N terminus of p37. In order to characterize the interaction between p34 and p37 and NOPP44/46, we have utilized an RNA interference (RNAi) cell line that specifically targets p34 and p37. Within these RNAi cells, we detected a disruption of a higher-molecular-weight complex containing NOPP44/46, as well as a dramatic increase in nuclear NOPP44/46 protein levels. We demonstrated that no change occurred in NOPP44/46 mRNA steadystate levels or stability, nor was there a change in cellular protein levels. These results led us to investigate whether p34 and p37 regulate NOPP44/46 cellular localization. Examination of the p34 and p37 amino acid sequences revealed a leucine-rich nuclear export signal, which interacts with the nuclear export factor exportin 1. Immune capture experiments demonstrated that p34, p37, and NOPP44/46 associate with exportin 1. When these experiments were performed with p34/p37 RNAi cells, NOPP44/46 no longer associated with exportin 1. Sequential immune capture experiments demonstrated that p34, p37, NOPP44/46, and exportin 1 exist in a common complex. Inhibiting exportin 1-mediated nuclear export led to an increase in nuclear NOPP44/46 proteins, indicating that they are exported from the nucleus via this pathway. Together, our results demonstrate that p34 and p37 regulate NOPP44/46 cellular localization by facilitating their association with exportin 1.African trypanosomes are the causative agents of African sleeping sickness in humans and nagana in cattle (22). This parasite continues to pose a serious threat to human health and to cause devastating economic losses (25). African trypanosomiasis is a reemerging infectious disease with estimates ranging between 300,000 and 500,000 new cases each year (34). This rise in infection rates is due, in part, to an increase in parasite drug resistance and vector pesticide resistance.In previous work from our laboratory, two Trypanosoma brucei RNA binding proteins, p34 and p37, were identified and characterized (39, 40). The only major difference between them is an 18-amino-acid insert located within the amino terminus of p37 that is absent in p34. These proteins contain two RNA binding domains (RBD) within the central coding region, and they have been shown to interact with 5S rRNA (31). In addition to the interaction with 5S rRNA, p34 and p37 have been found to interact with a family of nucleolar phosphoproteins, NOPP44/46 (30). These abundant proteins have been identified and characterized by M. Parsons and colleagues (9,10,27,29). An interaction(s) between p34 and p37 and NOPP44/46 was mapped through yeast two-hybrid analysis and protein affinity chromatography and was shown to be mediated via the RNA binding domain of p34 and p37 (30). Thus, th...

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Section: Discussionmentioning

confidence: 80%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Trypanosoma brucei RNA Binding Proteins p34 and p37 Mediate NOPP44/46 Cellular Localization via the Exportin 1 Nuclear Export Pathway

2007

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Eukaryotic 5S rRNA biogenesis

2011

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The ribosome is a large complex containing both protein and RNA which must be assembled in a precise manner to allow proper functioning in the critical role of protein synthesis. 5S rRNA is the smallest of the RNA components of the ribosome, and although it has been studied for decades, we still do not have a clear understanding of its function within the complex ribosome machine. It is the only RNA species that binds ribosomal proteins prior to its assembly into the ribosome. Its transport into the nucleolus requires this interaction. Here we present an overview of some of the key findings concerning the structure and function of 5S rRNA and how its association with specific proteins impacts its localization and function.

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Global proteomic analysis in trypanosomes reveals unique proteins and conserved cellular processes impacted by arginine methylation

Lott

Fisk

et al. 2013

Journal of Proteomics

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Arginine methylation is a common posttranslational modification with reported functions in transcription, RNA processing and translation, and DNA repair. Trypanosomes encode five protein arginine methyltransferases, suggesting that arginine methylation exerts widespread impacts on the biology of these organisms. Here, we performed a global proteomic analysis of T. brucei to identify arginine methylated proteins and their sites of modification. Using an approach entailing two-dimensional chromatographic separation, and alternating electron transfer dissociation and collision induced dissociation, we identified 1332 methylarginines in 676 proteins. The resulting data set represents the largest compilation of arginine methylated proteins in any organism to date. Functional classification revealed numerous arginine methylated proteins involved in flagellar function, RNA metabolism, DNA replication and repair, and intracellular protein trafficking. Thus, arginine methylation has the potential to impact aspects of T. brucei gene expression, cell biology, and pathogenesis. Interestingly, pathways with known methylated proteins in higher eukaryotes were identified in this study, but often different components of the pathway were methylated in trypanosomes. Methylarginines were often identified in glycine rich contexts, although exceptions to this rule were detected. Collectively, these data inform on a multitude of aspects of trypanosome biology and serve as a guide for the identification of homologous arginine methylated proteins in higher eukaryotes.

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Two Trypanosome-Specific Proteins Are Essential Factors for 5S rRNA Abundance and Ribosomal Assembly in Trypanosoma brucei

Cited by 22 publications

References 36 publications

Trypanosoma brucei RNA Binding Proteins p34 and p37 Mediate NOPP44/46 Cellular Localization via the Exportin 1 Nuclear Export Pathway

Trypanosoma brucei RNA Binding Proteins p34 and p37 Mediate NOPP44/46 Cellular Localization via the Exportin 1 Nuclear Export Pathway

Eukaryotic 5S rRNA biogenesis

Global proteomic analysis in trypanosomes reveals unique proteins and conserved cellular processes impacted by arginine methylation

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