Trypanosoma brucei repeated element with unusual structural and transcriptional properties

Murphy, Noel B.; Pays, Annette; Tebabi, Patricia; Coquelet, H; Guyaux, Michel; Steinert, Maurice; Pays, Étienne

doi:10.1016/0022-2836(87)90490-6

Cited by 141 publications

(94 citation statements)

References 45 publications

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“…7A). This is consistent with previous reports by others (25) and reflects the highly dispersed genomic organization of the Ingi retroposon family (22). We observed that the decay rate of the majority of the discrete transcripts visible by Northern blotting was similar in ago1 Ϫ/Ϫ and wild-type cells, with the exception of one transcript, indicated by an asterisk in Fig.…”

Section: Ago1 Affects the Accumulation Of Retroposon Transcriptssupporting

confidence: 93%

Argonaute Protein in the Early Divergent Eukaryote Trypanosoma brucei: Control of Small Interfering RNA Accumulation and Retroposon Transcript Abundance

Shi¹,

Djikeng²,

Tschudi³

et al. 2004

Molecular and Cellular Biology

110

140

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Members of the Argonaute protein family have been linked through a combination of genetic and biochemical studies to RNA interference (RNAi) and related phenomena. Here, we describe the characterization of the first Argonaute protein (AGO1) in Trypanosoma brucei, the earliest divergent eukaryote where RNAi has been described so far. AGO1 is predominantly cytoplasmic and is found in a ribonucleoprotein particle with small interfering RNAs (siRNAs), and this particle is present in a soluble form, as well as associated with polyribosomes. A genetic knockout of AGO1 leads to a loss of RNAi, and concomitantly, endogenous retroposonderived siRNAs as well as siRNAs derived from transgenic double-stranded RNA are reduced to almost undetectable levels. Furthermore, AGO1 deficiency leads to an increase in retroposon transcript abundance via mechanisms operating at the transcriptional level and at the RNA stability level. Our results suggest that AGO1 function is required for production and/or stabilization of siRNAs and provide the first evidence for an Argonaute protein being involved in the regulation of retroposon transcript levels.

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Section: Ago1 Affects the Accumulation Of Retroposon Transcriptssupporting

confidence: 93%

Argonaute Protein in the Early Divergent Eukaryote Trypanosoma brucei: Control of Small Interfering RNA Accumulation and Retroposon Transcript Abundance

Shi¹,

Djikeng²,

Tschudi³

et al. 2004

Molecular and Cellular Biology

110

140

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“…2B) demonstrates that it contains three different, closely spaced genes. The first one is a RIME element (nucleotides 376 -876), a member of a family of abundant, highly transcribed, repetitive transposable elements (44). Within this element, nucleotides 868 -632 on the reverse strand represent the open reading frame coding for a RIME-associated protein.…”

Section: Resultsmentioning

confidence: 99%

Characterization of TbPDE2A, a Novel Cyclic Nucleotide-specific Phosphodiesterase from the Protozoan Parasite Trypanosoma brucei

Zoraghi¹,

Kunz

Gong³

et al. 2001

Journal of Biological Chemistry

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This study reports the identification and characterization of a cAMP-specific phosphodiesterase from the parasitic hemoflagellate Trypanosoma brucei. TbPDE2A is a class I phosphodiesterase. Its catalytic domain exhibits 30 -40% sequence identity with those of all 11 mammalian phosphodiesterase (PDE) families, as well as with PDE2 from Saccharomyces cerevisiae, dunce from Drosophila melanogaster, and regA from Dictyostelium discoideum. The overall structure of TbPDE2A resembles that of human PDE11A in that its N-terminal region contains a single GAF domain. This domain is very similar to those of the mammalian PDE2, -5, -6, -10, and -11, where it constitutes a potential cGMP binding site. TbPDE2A can be expressed in S. cerevisiae, and it complements an S. cerevisiae PDE deletion strain. Recombinant TbPDE2A is specific for cAMP, with a K m of ϳ2 M. It is entirely resistant to the nonselective PDE inhibitor 3-isobutyl-1-methylxanthine, but it is sensitive to trequinsin, dipyridamole, sildenafil, and ethaverine with IC 50 values of 5.4, 5.9, 9.4, and 14.2 M, respectively. All four compounds inhibit proliferation of bloodstream form trypanosomes in culture, indicating that TbPDE2A is an essential enzyme.

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“…Isolation of nuclei, labeling of nascent transcripts, and hybridization to filters were done as described previously (19,32 P]UTP in the presence or absence of 1 mg of ␣-amanitin per ml and isolated, after DNase I (Gibco BRL) and proteinase K (Sigma) digestion, by trichloroacetic acid precipitation. Filters were prepared for hybridization by linearizing 5 g each of plasmid DNA with the appropriate enzyme, denaturing with 0.1 M NaOH, neutralizing with 6ϫ SSC (1ϫ SSC is 0.15 M NaCl plus 0.015 M sodium citrate), and transferring to Nytran filters with a slot blot apparatus (Schleicher and Schuell).…”

Section: Strainsmentioning

confidence: 99%

“…Filters were prehybridized, hybridized, and washed as previously described (30). [␣- 32 P]UTP-labeled riboprobes were prepared by transcribing gel-isolated restriction-enzyme-digested fragments with T3 or T7 RNA polymerase (Stratagene). Unincorporated label was removed with a Bio-Gel P-30 (Bio-Rad) column.…”

Section: Strainsmentioning

confidence: 99%

Increased Expression of LD1 Genes Transcribed by RNA Polymerase I in Leishmania donovani as a Result of Duplication into the rRNA Gene Locus

Lodes

Merlin

deVos

et al. 1995

Molecular and Cellular Biology

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Eukaryotic protein-coding genes are generally transcribed by RNA polymerase II (Pol II), which has a lower transcription rate than that of Pol I. We report here the duplication of two LD1 genes into the rRNA locus and their resultant transcription by Pol I. The multigenic LD1 locus is present in a 2.2-Mb chromosome in all stocks of Leishmania spp. and is also present in multicopy 200-to 450-kb linear chromosomes or multicopy circular DNAs in over 15% of stocks examined. Genomic rearrangement in Leishmania donovani LSB-51.1 resulted in duplication of a 3.9-kb segment of LD1 containing two genes (orfF and orfG) and of a 1.3-kb segment from ϳ10 kb downstream into the rRNA gene repeat region of the 1.2-Mb chromosome. Short sequences (12 or 13 bp) common to the 2.2-Mb LD1 and 1.2-Mb rRNA loci suggest that this gene conversion occurred by homologous recombination. Transcription of the duplicated genes is ␣-amanitin resistant, indicating transcription by Pol I, in contrast to the ␣-amanitin-sensitive (Pol II) transcription of the genes in the 2.2-Mb LD1 locus. This results in higher transcript abundance than expected from the gene copy number in LSB-51.1 and in elevated expression of at least the orfF gene product.Several multigene loci undergo amplification in Leishmania spp. (for a review, see reference 3). These gene amplifications have often been the result of selection for drug resistance in vitro (4,5,21,29,37). However, unselected strains of Leishmania spp. also have amplified DNAs, including LD1 (27, 52), but their significance is unknown. The amplicons are circular or linear molecules of various sizes and copy numbers, and the mechanism of their amplification appears to involve homologous recombination based on the presence of direct or inverted repeats at the termini of the amplified sequence. For example, rearrangement of the H region of Leishmania tropica has been shown to involve either direct or inverted DNA repeats of approximately 200 bp or greater (38, 55), while the circular LD1 (CD1) of Leishmania mexicana M379 is thought to be generated by recombination involving short (13-bp) direct repeats (28).We have been investigating the multigene LD1 locus that is present within a 2.2-Mb chromosome in Leishmania donovani and a similar-sized chromosome in all Leishmania strains. LD1 locus sequences are amplified as 20-to 60-copy, 200-to 450-kb chromosomes in some strains or 100-to 200-copy circular DNAs in others (52, 53). The LD1 locus contains nine genes, including one (orfB) which encodes ribosomal protein L37 (35), one (orfC) with homology to genes which appear to have a role in regulation of cell growth (36), and another (orfG) with homology to Trypanosoma brucei ESAG10 (18, 34). ESAG10 and orfG predict proteins with 10 to 12 potential membranespanning domains, indicating possible transporter function (34).In the Kinetoplastida, most protein-coding genes are transcribed as polycistronic units by RNA polymerase II (Pol II), similar to other eukaryotic systems. However, in T. brucei, the variant surface gly...

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Trypanosoma brucei repeated element with unusual structural and transcriptional properties

Cited by 141 publications

References 45 publications

Argonaute Protein in the Early Divergent Eukaryote Trypanosoma brucei: Control of Small Interfering RNA Accumulation and Retroposon Transcript Abundance

Argonaute Protein in the Early Divergent Eukaryote Trypanosoma brucei: Control of Small Interfering RNA Accumulation and Retroposon Transcript Abundance

Characterization of TbPDE2A, a Novel Cyclic Nucleotide-specific Phosphodiesterase from the Protozoan Parasite Trypanosoma brucei

Increased Expression of LD1 Genes Transcribed by RNA Polymerase I in Leishmania donovani as a Result of Duplication into the rRNA Gene Locus

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