Characterization of VSG gene expression site promoters and promoter-associated DNA rearrangement events.

Gottesdiener, Keith; Chung, Huimin; Brown, Samantha; Lee, M. G.-S.; Ploeg, Lex H.T. Van der

doi:10.1128/mcb.11.5.2467

Cited by 62 publications

(67 citation statements)

References 77 publications

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“…All of our sequence are shown the minimal promoter sequences for a PARP B gene and an rRNA gene and a consensus sequence (cons.) for the three reported bloodstream VSG promoters (30,36,76). Boxes show regions of similarity among the PARP B, rRNA, and consensus bloodstream VSG promoters (7).…”

Section: Discussionmentioning

confidence: 99%

“…Nuclear run-on assays, coupled with UV inactivation of transcription, have been used to demonstrate that for at least some VSG ESs, transcription is initiated 45 to 60 kb upstream of the VSG gene and proceeds through eight or more open translation reading frames, the last of which specifies the VSG (12,30,37,42,56). This transcription occurs in the presence of ox-amanitin, leading to suggestions that it is catalyzed by an RNA polymerase I activity or by a modified RNA polymerase II complex (26,32,59,75).…”

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confidence: 99%

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A monocistronic transcript for a trypanosome variant surface glycoprotein.

Alarcon¹,

Son²,

Hall³

et al. 1994

Mol. Cell. Biol.

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Many protein-encoding genes of African trypanosomes are transcribed as large polycistronic pre-mRNAs that are processed into individual mRNAs containing a 5' spliced leader and 3' poly(A). The 45-to 60-kb pre-mRNAs encoding some variant surface glycoproteins (VSGs) contain as many as eight unrelated coding regions. Here we identify the promoter for a metacyclic VSG gene that is expressed without duplication in a bloodstream trypanosome clone. This 70-bp promoter is located 2 kb upstream of the telomere-linked VSG gene and directs the synthesis of a monocistronic VSG pre-mRNA lacking the 5' spliced leader. Its sequence only slightly resembles those of other known trypanosome promoters, but it does cross-hybridize with several related sequences elsewhere in the genome. These results suggest that a new class of trypanosome promoters has been found, whose function is to initiate monocistronic transcription of those VSG genes normally expressed during the metacyclic stage.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

A monocistronic transcript for a trypanosome variant surface glycoprotein.

Alarcon¹,

Son²,

Hall³

et al. 1994

Mol. Cell. Biol.

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“…The ES promoter is located 40 to 60 kb upstream of the telomeric vsg (25,27). ES promoters sequenced so far show more than 90% sequence identity (18,37,53). The 5Ј end of the ES promoter is flanked by a region of imperfect direct repeats of 50 bp, which covers at least 10 kb in the vsg221 ES (54).…”

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confidence: 99%

DNA Rearrangements Associated with Multiple Consecutive Directed Antigenic Switches in Trypanosoma brucei

Navarro

Cross

1996

Molecular and Cellular Biology

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Changes in variant surface glycoprotein (Vsg) expression allow Trypanosoma brucei to elude the immune response. The expressed vsg is always located at the telomeric end of a polycistronic transcription unit known as an expression site (ES). Although there are many ESs, only one is active at any particular time. The mechanisms regulating ES transcription and switching are unknown. Chromosome rearrangements within or upstream of the ES have been described to occur in occasional switch events, but no changes have been consistently associated with switching. We inserted the drug resistance genes neo and ble, conferring resistance to G418 and phleomycin, respectively, 1 kb downstream of "silent" ES promoters. This demonstrated that short-range transcription could be achieved from a silent ES promoter. From one initial transformant clone, panels of independent consecutive on-off-on switch clones were generated and analyzed. The first activation of the neo-targeted ES was always associated with deletion of the upstream tandem promoter in this ES, but no further rearrangements were detected in consecutive off-on switches of this ES. On the other hand, direct analysis of ES promoters showed that deletions and duplications occurred elsewhere. Activation of a ble-tagged 300-kb chromosome could not be achieved, but phleomycin-resistant clones could be obtained. One such clone arose from recombination between three ESs. Taken together, our experiments suggest that ES switching may occur after a period of chromosomal interactivity that may or may not leave tangible evidence in the form of detectable sequence changes.Trypanosoma brucei is an extracellular protozoan parasite that multiplies in the tissue fluids and vascular system of its vertebrate host, causing major veterinary and medical problems in Africa. The entire surface of the bloodstream form trypanosome is covered by a dense coat, which consists largely of a single molecular species known as the variant surface glycoprotein (Vsg) (8). Sequential expression of vsg genes allows the parasite to elude the immune response of the host (reviewed in references 9, 10, and 51). The expressed vsg is always located adjacent to a telomere, at the end of a polycistronic expression site (ES) (15). Hybridization analysis suggests the presence of approximately 20 ESs, whose sequences are highly conserved (13,54). Only one ES is active, yielding a single Vsg on the surface of the infective trypanosome. The ES contains a number of ES-associated genes (esag) (13, 51). ES transcription is driven by an ␣-amanitin-resistant polymerase (26, 27). The ES promoter is located 40 to 60 kb upstream of the telomeric vsg (25, 27). ES promoters sequenced so far show more than 90% sequence identity (18, 37, 53). The 5Ј end of the ES promoter is flanked by a region of imperfect direct repeats of 50 bp, which covers at least 10 kb in the vsg221 ES (54).Antigenic variation in T. brucei can occur by duplicative transposition of a new vsg into an already active ES, displacing the resident vsg. A switch in...

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“…2, boxed regions of VSG, PRO, and RIB). However, the level of activity of the Ϫ69 to [32]; 118 [12]). In addition, the sequence of a metacyclic (Met) VSG promoter (AnTat 11.17 [30]) is aligned for maximal homology, with dashes for spacing.…”

Section: Anatomy Of the Vsg Promotermentioning

confidence: 99%

“…Because of the general organization of genes into polycistronic transcription units, there is only limited information on transcription promoters and their controls. The only promoters for protein-coding genes that are known so far are those of VSG and procyclin (11,12,23,26,31,32). There are two diploid loci for the procyclin transcription units, and it is believed that the promoters of all these units are simultaneously active in the procyclic form (11,20).…”

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confidence: 99%

Specific Binding of Proteins to the Noncoding Strand of a Crucial Element of the Variant Surface Glycoprotein, Procyclin, and Ribosomal Promoters of Trypanosoma brucei

Vanhamme¹,

Pays²,

Tebabi³

et al. 1995

Molecular and Cellular Biology

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The variant surface glycoprotein (VSG) and procyclin promoters of Trypanosoma brucei recruit an RNA polymerase sharing characteristics with polymerase I, but there is no sequence homology between them nor between these promoters and ribosomal promoters. We report the detailed characterization of the VSG promoter. The 70-bp region upstream of the transcription start site was sufficient for full promoter activity. Mutational analysis revealed three short critical stretches at positions ؊61 to ؊59 (box 1), ؊38 to ؊35 (box 2), and ؊1 to ؉1 (start site), the spacing of which was essential. These elements were conserved in the promoter for a metacyclic VSG gene. Hybrid sequences containing box 1 of the VSG promoter and box 2 of the ribosomal promoter were active. A specific binding of proteins to the noncoding strand of box 2, but not to double-stranded DNA, occurred. Competition experiments indicated that these proteins also bind to the corresponding region of the metacyclic VSG, procyclin, and ribosomal promoters. Binding of such a protein, of 40 kDa, appeared to be shared by these promoters.The protozoan parasite Trypanosoma brucei shares its life cycle between mammals and an insect vector, the tsetse fly. The bloodstream (mammalian) form and the procyclic (insect) form are uniformly covered by a surface coat, of variant surface glycoprotein (VSG) and procyclin, respectively. The VSG replaces procyclin when procyclic forms differentiate into metacyclic forms in the salivary glands of the fly. This surface antigen persists throughout the development of the parasite in the blood and rapidly disappears when procyclin is reexpressed upon differentiation from the bloodstream to the procyclic form. Therefore, the VSG and procyclin are typical markers for the major developmental stages of the parasite (for recent reviews, see references 4, 10, 24, and 25).African trypanosomes are responsible for important human and animal plagues. They escape the immune defenses of their hosts by a periodical change of their antigenic VSG coat. The study of the control of VSG expression is thus of major fundamental and economic importance. However, the mechanisms controlling gene expression in trypanosomatids are poorly understood. This is particularly true for transcriptional controls. Because of the general organization of genes into polycistronic transcription units, there is only limited information on transcription promoters and their controls. The only promoters for protein-coding genes that are known so far are those of VSG and procyclin (11,12, 23,26,31,32). There are two diploid loci for the procyclin transcription units, and it is believed that the promoters of all these units are simultaneously active in the procyclic form (11,20). The number of VSG transcription units is estimated to be between 6 and 20 (4, 24), but only a single one is expressed at a given time. So far it is not clear if this control operates at the level of promoter activity or transcription attenuation. Interestingly, although the VSG and procyclin units ar...

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Characterization of VSG gene expression site promoters and promoter-associated DNA rearrangement events.

Cited by 62 publications

References 77 publications

A monocistronic transcript for a trypanosome variant surface glycoprotein.

A monocistronic transcript for a trypanosome variant surface glycoprotein.

DNA Rearrangements Associated with Multiple Consecutive Directed Antigenic Switches in Trypanosoma brucei

Specific Binding of Proteins to the Noncoding Strand of a Crucial Element of the Variant Surface Glycoprotein, Procyclin, and Ribosomal Promoters of Trypanosoma brucei

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