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Trypanosome MTR4 is involved in rRNA processing.

ABSTRACT: The yeast putative RNA helicase Mtr4p is implicated in exosome-mediated RNA quality control in the nucleus, interacts with the exosome, and is found in the 'TRAMP' complex with a yeast nuclear poly(A) polymerase (Trf4p/Pap2p or Trf5p) and a putative RNA-binding protein, Air1p or Air2p. Depletion of the Trypanosoma brucei MTR4-like protein TbMTR4 caused growth arrest and defects in 5.8S rRNA processing similar to those seen after depletion of the exosome. TbNPAPL, a nuclear protein which is a putative homolog of Trf4p/Pap2p, was required for normal cell growth. Depletion of MTR4 resulted in the accumulation of polyadenylated rRNA precursors, while depletion of TbNPAPL had little effect. These results suggest that polyadenylation-dependent nuclear rRNA quality control is conserved in eukaryotic evolution. In contrast, there was no evidence for a trypanosome TRAMP complex since no stable interactions between TbMTR4 and the exosome, TbNPAPL or RNA-binding proteins were detected.

SUBMITTER: Cristodero M

PROVIDER: S-EPMC2175323 | biostudies-literature | 2007

REPOSITORIES: biostudies-literature

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Trypanosome MTR4 is involved in rRNA processing.

Cristodero Marina M Clayton Christine E CE

Nucleic acids research 20071016 20

The yeast putative RNA helicase Mtr4p is implicated in exosome-mediated RNA quality control in the nucleus, interacts with the exosome, and is found in the 'TRAMP' complex with a yeast nuclear poly(A) polymerase (Trf4p/Pap2p or Trf5p) and a putative RNA-binding protein, Air1p or Air2p. Depletion of the Trypanosoma brucei MTR4-like protein TbMTR4 caused growth arrest and defects in 5.8S rRNA processing similar to those seen after depletion of the exosome. TbNPAPL, a nuclear protein which is a put ...[more]

PMID: 17940093

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Project description:Kinetoplastid parasites-trypanosomes and leishmanias-infect millions of humans and cause economically devastating diseases of livestock, and the few existing drugs have serious deficiencies. Benzoxaborole-based compounds are very promising potential novel anti-trypanosomal therapies, with candidates already in human and animal clinical trials. We investigated the mechanism of action of several benzoxaboroles, including AN7973, an early candidate for veterinary trypanosomosis. In all kinetoplastids, transcription is polycistronic. Individual mRNA 5'-ends are created by trans splicing of a short leader sequence, with coupled polyadenylation of the preceding mRNA. Treatment of Trypanosoma brucei with AN7973 inhibited trans splicing within 1h, as judged by loss of the Y-structure splicing intermediate, reduced levels of mRNA, and accumulation of peri-nuclear granules. Methylation of the spliced leader precursor RNA was not affected, but more prolonged AN7973 treatment caused an increase in S-adenosyl methionine and methylated lysine. Together, the results indicate that mRNA processing is a primary target of AN7973. Polyadenylation is required for kinetoplastid trans splicing, and the EC50 for AN7973 in T. brucei was increased three-fold by over-expression of the T. brucei cleavage and polyadenylation factor CPSF3, identifying CPSF3 as a potential molecular target. Molecular modeling results suggested that inhibition of CPSF3 by AN7973 is feasible. Our results thus chemically validate mRNA processing as a viable drug target in trypanosomes. Several other benzoxaboroles showed metabolomic and splicing effects that were similar to those of AN7973, identifying splicing inhibition as a common mode of action and suggesting that it might be linked to subsequent changes in methylated metabolites. Granule formation, splicing inhibition and resistance after CPSF3 expression did not, however, always correlate and prolonged selection of trypanosomes in AN7973 resulted in only 1.5-fold resistance. It is therefore possible that the modes of action of oxaboroles that target trypanosome mRNA processing might extend beyond CPSF3 inhibition.

Project description:Although the nucleolus is known to be involved in the plant stress response, the underlying mechanisms remain unclear. In the present study, we genetically isolated and characterized the Arabidopsis thaliana salt hypersensitive mutant 1 (sahy1), which exhibits slow growth, short roots, and sterility. SAHY1 encodes an unknown protein that is predominantly expressed in root tips and mature pollens. Subcellular localization further demonstrated that SAHY1 is a nucleolar protein. Ploidy analysis and detection of S-phase cell cycle progression indicated that mitosis is reduced in the root tips of sahy1 plants. Furthermore, levels of reactive oxygen species (ROS) are reduced in the mutant. In addition to exhibiting changes in venation pattern, palisade mesophyll cells, the auxin response, and fertility, the sahy1 mutant is resistant to the protein translation inhibitors, streptomycin and cycloheximide, and has altered expression profiles of ribosome-related genes, reflecting the involvement of SAHY1 in ribosome assembly. Using a combination of transcriptome and gene coexpression clustering analyses, we identified a striking functional module in which SPIRAL 1-LIKE 4 (SPL4), which encodes a microtubule-localized protein, mediates the salt response that acts by modulating dynamic microtubule instability. Supporting evidence indicates that sahy1 is resistant to a microtubule-destabilizing drug (oryzalin), that the cortical microtubule array in sahy1 exhibits changes in dynamic instability, and that the spr1/spl2/spl3/spl4 quadruple mutant is oversensitive to salt. Collectively, these data demonstrate that a novel nucleolar protein, SAHY1, plays critical roles in salt sensitivity, plant growth, and development by integrating the diverse functions of its downstream genes. As the sahy1 mutant plants showed slow growth, short roots, and sterility, the aim of this experiment is to examine gene expression profile in root tissues mediated by SAHY1. The data obtained from this study was compared with chip database derived from root tissues and performed an analysis of gene coexpression clustering.

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Trypanosome MTR4 is involved in rRNA processing.

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Trypanosome MTR4 is involved in rRNA processing.

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