The small ubiquitin-like modifier (SUMO) is essential in cell cycle regulation in Trypanosoma brucei

Liao, Shanhui; Wang, Tao; Fan, Kai; Tu, Xiaoming

doi:10.1016/j.yexcr.2009.12.017

Cited by 41 publications

(49 citation statements)

References 40 publications

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“…This construct generated TbTim50 double-stranded RNA from two opposing tetracycline-regulated T7 promoters and expressed the phleomycin-resistant gene constitutively for selection purposes. The TbTim50 overexpression construct was generated using the modified pLEW100 -3HA vector (a gift from Dr. Xiaoming Tu) (38). The entire open reading frame (ORF) of TbTim50 was PCR-amplified from T. brucei genomic DNA using primers shown in supplemental Table 1.…”

Section: Methodsmentioning

confidence: 99%

Tim50 in Trypanosoma brucei Possesses a Dual Specificity Phosphatase Activity and Is Critical for Mitochondrial Protein Import

Duncan

Fullerton

Chaudhuri

2013

Journal of Biological Chemistry

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Background: Tim50 is a preprotein receptor for the TIM23 complex in mitochondria. Results: T. brucei Tim50 (TbTim50), a protein phosphatase, is involved in the regulation of VDAC expression, interacts with TbTim17, and imports precursor proteins into mitochondria. Conclusion: TbTim50 is evolutionarily distinct from Tim50 in other eukaryotes. Significance: Characterization of TbTim50 is critical for understanding mitochondrial protein import in T. brucei.

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Section: Methodsmentioning

confidence: 99%

Tim50 in Trypanosoma brucei Possesses a Dual Specificity Phosphatase Activity and Is Critical for Mitochondrial Protein Import

Duncan

Fullerton

Chaudhuri

2013

Journal of Biological Chemistry

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“…PCRs were performed using appropriate forward primers (see Table S1) for generation of N-terminal deletion constructs (⌬10TAO-3HA, ⌬20TAO-3HA, ⌬30TAO-3HA, and ⌬40TAO-3HA), and the same reverse primer was used for generation of the full-length TAO construct. Digested and purified PCR products were subcloned into a pLEW100-3HA vector (a generous gift from Xiaoming Tu) (27) between the HindIII and XhoI sites. For generation of the TAO-DHFR fusion constructs, FLTAO and TAO fragments (amino acid residues 1 to 30 and 31 to 329 of TAO) were PCR amplified using forward and reverse primers (see Table S1) containing HindIII and BamHI restriction sites at the 5=ends, respectively.…”

Section: Cellsmentioning

confidence: 99%

Trypanosome Alternative Oxidase Possesses both an N-Terminal and Internal Mitochondrial Targeting Signal

et al. 2014

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Recognition of mitochondrial targeting signals (MTS) by receptor translocases of outer and inner membranes of mitochondria is one of the prerequisites for import of nucleus-encoded proteins into this organelle. The MTS for a majority of trypanosomatid mitochondrial proteins have not been well defined. Here we analyzed the targeting signal for trypanosome alternative oxidase (TAO), which functions as the sole terminal oxidase in the infective form of Trypanosoma brucei. Deleting the first 10 of 24 amino acids predicted to be the classical N-terminal MTS of TAO did not affect its import into mitochondria in vitro. Furthermore, ectopically expressed TAO was targeted to mitochondria in both forms of the parasite even after deletion of first 40 amino acid residues. However, deletion of more than 20 amino acid residues from the N terminus reduced the efficiency of import. These data suggest that besides an N-terminal MTS, TAO possesses an internal mitochondrial targeting signal. In addition, both the N-terminal MTS and the mature TAO protein were able to target a cytosolic protein, dihydrofolate reductase (DHFR), to a T. brucei mitochondrion. Further analysis identified a cryptic internal MTS of TAO, located within amino acid residues 115 to 146, which was fully capable of targeting DHFR to mitochondria. The internal signal was more efficient than the N-terminal MTS for import of this heterologous protein. Together, these results show that TAO possesses a cleavable N-terminal MTS as well as an internal MTS and that these signals act together for efficient import of TAO into mitochondria.

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“…The fact SUMO resulted essential for T. brucei viability (18,19) reflects that the SUMOylation system is also important in these early divergent eukaryotes, and emphasizes the relevance of studying the proteins modified by SUMO. According to the conditions in which the purification of SUMOylated proteins is performed, a different kind of information can be extracted from a proteomic study.…”

Section: Fig 5 Validation Of Metacaspase-3 As a Sumoylation Target mentioning

confidence: 99%

“…Ub controls protein fate by triggering degradation through the 26S proteasome and the Ubl Atg8 is involved in autophagosome biogenesis that recycles cellular material. In the case of SUMO studies performed in T. brucei have shown that SUMO is essential in cell cycle regulation in procyclic and bloodstream forms (18,19). However, the SUMOylation system has not been characterized yet at the molecular level nor have the targets of SUMO been identified in any Trypanosomatid.…”

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confidence: 99%

SUMOylation Pathway in Trypanosoma cruzi: Functional Characterization and Proteomic Analysis of Target Proteins

Bayona

Nakayasu

Laverrière

et al. 2011

Molecular & Cellular Proteomics

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SUMOylation is a relevant protein post-translational modification in eukaryotes. The C terminus of proteolytically activated small ubiquitin-like modifier (SUMO) is covalently linked to a lysine residue of the target protein by an isopeptide bond, through a mechanism that includes an E1-activating enzyme, an E2-conjugating enzyme, and transfer to the target, sometimes with the assistance of a ligase. The modification is reversed by a protease, also responsible for SUMO maturation. A number of proteins have been identified as SUMO targets, participating in the regulation of cell cycle progression, transcription, translation, ubiquitination, and DNA repair. In this study, we report that orthologous genes corresponding to the SUMOylation pathway are present in the etiological agent of Chagas disease, Trypanosoma cruzi. Furthermore, the SUMOylation system is functionally active in this protozoan parasite, having the requirements for SUMO maturation and conjugation. Immunofluorescence analysis showed that T. cruzi SUMO (TcSUMO) is predominantly found in the nucleus. To identify SUMOylation targets and get an insight into their physiological roles we generated transfectant T. cruzi epimastigote lines expressing a double-tagged T. cruzi SUMO, and SUMOylated proteins were enriched by tandem affinity chromatography. By two-dimensional liquid chromatography-mass spectrometry a total of 236 proteins with diverse biological functions were identified as potential T. cruzi SUMO targets. Of these, metacaspase-3 was biochemically validated as a bona fide SUMOylation substrate. Proteomic studies in other organisms have reported that orthologs of putative T.

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The small ubiquitin-like modifier (SUMO) is essential in cell cycle regulation in Trypanosoma brucei

Cited by 41 publications

References 40 publications

Tim50 in Trypanosoma brucei Possesses a Dual Specificity Phosphatase Activity and Is Critical for Mitochondrial Protein Import

Tim50 in Trypanosoma brucei Possesses a Dual Specificity Phosphatase Activity and Is Critical for Mitochondrial Protein Import

Trypanosome Alternative Oxidase Possesses both an N-Terminal and Internal Mitochondrial Targeting Signal

SUMOylation Pathway in Trypanosoma cruzi: Functional Characterization and Proteomic Analysis of Target Proteins

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