Antigenic variation inTrypanosoma brucei: a telomeric expression site for variant-specific surface glycoprotein genes with novel features

Abstract

African trypanosomes evade the immune response of their host by periodically changing their variant surface glycoprotein (VSG) coat. Each coat is encoded by a separate VSG gene. Expressed genes are in a telomeric expression site (ES) and there are several sites in each trypanosome. To study the transcription control of VSG genes in Trypanosoma brucei we have analyzed an ES, called the dominant ES (DES), that readily switches off and on. The promoter area of the DES is very similar to that of the 221 ES (Zomerdijk et al., 1990). It can be switched off and on in vivo without detectable DNA alterations in the vicinity of the transcription start and it can drive high transient expression of a reporter gene in transfection experiments. However, there are also two major differences between the DES and the 221 ES. First, one version of the DES contains an additional upstream transcription unit overlapping the VSG gene ES promoter. The presence of this upstram transcription is dispensable, however, for the VSG gene ES promoter is active, even if transcription through this start from the upstream promoter is blocked using UV light. Moreover, a second version of the DES present in another trypanosome variant does not produce these upstream transcripts. Secondly, we find that the inactivation of DES transcription in one trypanosome variant is accompanied by DNA alterations in the DES upstream (greater than 2 kb) of the transcription start; reactivation of DES transcription is accompanied by another alteration far upstream. Although we cannot exclude that these DNA rearrangements are incidental, our results raise the possibility that the activity of ES promoters is negatively controlled in cis by far upstream sequences not included in transfection constructs and that alterations in these sequences may lead to (in)activation of the promoter.

Transcription is initiated on silent variant surface glycoprotein expression sites despite monoallelic expression in Trypanosoma brucei

“…Most sequences were present in 1-11 copies, except for G1, which was present in 30-40 copies. Because G1 is present in both FM162566 and FM162572 (33), and FM162566 is the active ES in this isolate, the relatively high proportion of this PCR product was in keeping with the higher transcription efficiency in the dominant ES (36) and with the more efficient processing of transcripts from the active ES (29). Here again, 11-17 outliers (12-19% of the total sequences) were generated.…”

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

“…The neo probe was the RsaI-BamHI restriction fragment containing the 3Ј end of the gene. The core ES promoter probe was a 315-bp HpaI-SalI fragment (positions 1359 to 1673 in the sequence described in reference 54) isolated from plasmid pRK8(ϩ) (52).…”

“…DNA rearrangements have been described to occur in the region upstream of the promoter, in a few ES inactivation events. Some of these rearrangements involved either the deletion of the upstream promoter, in an ES where the ES promoter, embracing a region of about 13 kb, was tandemly duplicated (18), or changes within the 50-bp repeat array upstream of the promoter (52). To determine the significance of these observations, we decided to search in detail, in panels of independent clones with consecutive on-off-on switches at a specific ES, for any DNA rearrangements that might occur consistently upon ES activation and inactivation.…”

Telomere as an Important Player in Regulation of Microbial Pathogen Virulence

“…Lister 427 has 15 different B-ESs), usually carrying different VSGs, but all B-ESs have very similar genomic organization with ~90% sequence identity (Hertz-Fowler et al 2008). Earlier studies focused on B-ES promoters also showed that they are almost always identical (Pham et al 1996;Zomerdijk et al 1990Zomerdijk et al , 1991. Therefore, how T. brucei manages to fully express only one B-ES and VSG had been a great puzzle for more than a couple of decades.…”