Telomere shortening and cell cycle arrest in Trypanosoma brucei expressing human telomeric repeat factor TRF1

Muñoz‐Jordán, Jorge L.; Cross, George A.M.

doi:10.1016/s0166-6851(01)00259-6

Cited by 18 publications

(9 citation statements)

References 48 publications

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“…Consistent with this possibility, different cyclins are used in bloodstream and procyclic stages to control cell-cycle progress (28,69). Interestingly, the overexpression of hTRF1 in procyclic T. brucei also resulted in S-phase arrest, though the phenotypes are not exactly the same as with tbTRF RNAi (52). It was proposed that hTRF1 partially displaces endogenous TTAGGG-binding factor(s) in procyclic cells.…”

Section: Discussionmentioning

confidence: 99%

“…It is interesting that tbTRF has higher affinity for double-stranded TTAGGG repeats than for any permuta- (73,74), and two J-binding proteins have been identified (20,59). Thus, tbTRF may bind the J-containing repeats with lower affinity, which also appears to be the case for hTRF1 expressed in T. brucei (52). However, the details of tbTRF DNA-binding specificity remain to be determined.…”

Section: Discussionmentioning

confidence: 99%

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Trypanosome Telomeres Are Protected by a Homologue of Mammalian TRF2

Espinal

Cross

2005

Molecular and Cellular Biology

186

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Putative TTAGGG repeat-binding factor (TRF) homologues in the genomes of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major were identified. They have significant sequence similarity to higher eukaryotic TRFs in their C-terminal DNA-binding myb domains but only weak similarity in their N-terminal domains. T. brucei TRF (tbTRF) is essential and was shown to bind to duplex TTAGGG repeats. The RNA interferencemediated knockdown of tbTRF arrested bloodstream cells at G 2 /M and procyclic cells partly at S phase. Functionally, tbTRF resembles mammalian TRF2 more than TRF1, as knockdown diminished telomere single-stranded G-overhang signals. This suggests that tbTRF, like vertebrate TRF2, is essential for telomere end protection, and this also supports the hypothesis that TRF rather than Rap1 is the more ancient DNA-binding component of the telomere protein complex. Identification of the first T. brucei telomere DNAbinding protein and characterization of its function provide a new route to explore the roles of telomeres in pathogenesis of this organism. This work also establishes T. brucei as an attractive model for telomere biology.Telomeres are specialized protein-DNA complexes at the ends of eukaryotic chromosomes. Telomere DNA generally consists of simple, repetitive, TG-rich sequences that are maintained by telomerase (5) and end with a single-stranded G-rich overhang (78). A specialized telomere structure, the T loop, formed by the invasion of telomere G overhangs into the telomeric double-stranded region, in mammals, hypotrichous ciliates, and Trypanosoma brucei has been identified (26,51,53). It is hypothesized that hiding the telomere single-stranded G overhangs in a T-loop structure helps to protect the telomere ends.Proteins that bind to duplex or single-stranded telomere DNA are integral components of the telomere complex and play critical roles in both telomere length regulation and end protection (36). In mammalian cells, two paralogues, TT AGGG repeat-binding factor 1 (TRF1) and TRF2, bind duplex TTAGGG repeats (4,9,12). Both TRF1 and TRF2 have a C-terminal myb motif for DNA binding (3, 9) and an upstream TRF homology (TRFH) domain for homodimerization (24). However, the N termini are quite different: TRF1 is acidic, while TRF2 is basic. Mammalian TRF1 negatively regulates telomere length through a telomerase-dependent pathway (61, 75), whereas TRF2 is involved in both telomere length regulation (38, 61) and telomere end protection (17). Removal of TRF2 from telomeres by overexpression of a dominantnegative mutant of TRF2 resulted in an at least 30%

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Trypanosome Telomeres Are Protected by a Homologue of Mammalian TRF2

Espinal

Cross

2005

Molecular and Cellular Biology

186

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Section: Discussionmentioning

confidence: 99%

“…A previous study showed that human TRF1 could be expressed in PF T. brucei cells and was located at the telomere that does not have J modification (67). However, human TRF1, when expressed in BF T. brucei cells where the DNA is modified by J, is not located at the telomere (67). Presumably, the presence of J in BF telomeres interfered with efficient binding of human TRF1, although the possibility that other telomere factors are involved in such reduced TRF1 binding cannot be ruled out.…”

Section: Discussionmentioning

confidence: 99%

Suppression of subtelomeric VSG switching by Trypanosoma brucei TRF requires its TTAGGG repeat-binding activity

Jehi

Sandhu

et al. 2014

Nucleic Acids Research

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Trypanosoma brucei causes human African trypanosomiasis and regularly switches its major surface antigen, VSG, in the bloodstream of its mammalian host to evade the host immune response. VSGs are expressed exclusively from subtelomeric loci, and we have previously shown that telomere proteins TbTIF2 and TbRAP1 play important roles in VSG switching and VSG silencing regulation, respectively. We now discover that the telomere duplex DNA-binding factor, TbTRF, also plays a critical role in VSG switching regulation, as a transient depletion of TbTRF leads to significantly more VSG switching events. We solved the NMR structure of the DNA-binding Myb domain of TbTRF, which folds into a canonical helix-loop-helix structure that is conserved to the Myb domains of mammalian TRF proteins. The TbTRF Myb domain tolerates well the bulky J base in T. brucei telomere DNA, and the DNA-binding affinity of TbTRF is not affected by the presence of J both in vitro and in vivo. In addition, we find that point mutations in TbTRF Myb that significantly reduced its in vivo telomere DNA-binding affinity also led to significantly increased VSG switching frequencies, indicating that the telomere DNA-binding activity is critical for TbTRF's role in VSG switching regulation.

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“…The reversion experiment was done by culturing the amplicon-containing cell lines in the absence of drug pressure. The DNA measurement by flow-cytometry was done as described in Munoz-Jordan and Cross (28). …”

Section: Methodsmentioning

confidence: 99%

Formation of linear inverted repeat amplicons following targeting of an essential gene in Leishmania

Genest

Riet²,

Dumas³

et al. 2005

Nucleic Acids Research

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Attempts to inactivate an essential gene in the protozoan parasite Leishmania have often led to the generation of extra copies of the wild-type alleles of the gene. In experiments with Leishmania tarentolae set up to disrupt the gene encoding the J-binding protein 1 (JBP1), a protein binding to the unusual base β-d-glucosyl-hydroxymethyluracil (J) of Leishmania, we obtained JBP1 mutants containing linear DNA elements (amplicons) of ∼100 kb. These amplicons consist of a long inverted repeat with telomeric repeats at both ends and contain either the two different targeting cassettes used to inactivate JBP1, or one cassette and one JBP1 gene. Each long repeat within the linear amplicons corresponds to sequences covering the JBP1 locus, starting at the telomeres upstream of JBP1 and ending in a ∼220 bp sequence repeated in an inverted (palindromic) orientation downstream of the JBP1 locus. We propose that these amplicons have arisen by a template switch inside a DNA replication fork involving the inverted DNA repeats and helped by the gene targeting.

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Telomere shortening and cell cycle arrest in Trypanosoma brucei expressing human telomeric repeat factor TRF1

Cited by 18 publications

References 48 publications

Trypanosome Telomeres Are Protected by a Homologue of Mammalian TRF2

Trypanosome Telomeres Are Protected by a Homologue of Mammalian TRF2

Suppression of subtelomeric VSG switching by Trypanosoma brucei TRF requires its TTAGGG repeat-binding activity

Formation of linear inverted repeat amplicons following targeting of an essential gene in Leishmania

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