Control of variant surface glycoprotein gene‐expression sites in Trypanosoma brucei

Abstract

Trypanosoma brucei has 20 similar telomeric-expression sites for variant surface glycoprotein genes. Expression sites appear to be controlled at the level of transcription initiation, resulting in only one site being active at any time. Switching between expression sites occurs at a low rate. To analyse the switching mechanism, we used trypanosomes with two expression sites tagged with two different drug-resistance genes and selected these on agarose plates containing both drugs. Double-resistant clones arose at a low frequency of 10 -7 per cell, but these behaved as if they rapidly switched between the two tagged expression sites and lost double resistance in the absence of selection. Using in situ hybridization we found that only 10% of the double-resistant cells had two fluorescent spots corresponding to transcribed expression sites. Our results suggest that: (i) a double expressor is not a stable intermediate in expression site switching; (ii) expression sites are not independently switched on and off; and (iii) expression sites can be in a 'pre-active' silent state from which they can be readily activated.

A Histone Methyltransferase Modulates Antigenic Variation in African Trypanosomes

“…We speculate that BES17 is expressed either in the nucleolus or in the unique ESB, close to BES1. This model is consistent with what was observed in double-resistant wild-type clones, in which the two rapidly switching active BESs often were found in close proximity in the nucleus [ 12 ], and with the localization of a partially active BES at the periphery of the nucleolus [ 32 ]. Despite the localization in a Pol I competent compartment, BES17 is only partially activated, which suggests that another limiting factor is missing.…”

A Histone Methyltransferase Modulates Antigenic Variation in African Trypanosomes

“…We speculate that BES17 is expressed either in the nucleolus or in the unique ESB, close to BES1. This model is consistent with what was observed in double-resistant wild-type clones, in which the two rapidly switching active BESs often were found in close proximity in the nucleus [ 12 ], and with the localization of a partially active BES at the periphery of the nucleolus [ 32 ]. Despite the localization in a Pol I competent compartment, BES17 is only partially activated, which suggests that another limiting factor is missing.…”

Epigenetic Regulation of Virulence Gene Expression in Parasitic Protozoa

“…In another experiment revealing communication among VSG s, artificial blockade of the active VSG -ES allowed the high-frequency activation of other VSG -ESs ( Aresta-Branco et al., 2016 ). These results are consistent with a remarkably robust exclusion mechanism operating in T. brucei and failure to select two stable, simultaneously active VSG -ESs ( Chaves et al., 1999 ).…”

Bloodstream Form-specific Up-regulation of Silent VSG Expression Sites and Procyclin in Trypanosoma brucei after Inhibition of DNA Synthesis or DNA Damage

“…Partial activation of ESs in bloodstream form T. brucei has been achieved before in experiments targeting drug resistance genes into "silent" ESs, although the levels of transcription were much lower than we show here (19,64). These low levels of transcription were "erased" after activation and subsequent inactivation of the partially active ES, implying the resetting of an epigenetic state (19).…”

A Histone Methyltransferase Modulates Antigenic Variation in African Trypanosomes

“…We speculate that BES17 is expressed either in the nucleolus or in the unique ESB, close to BES1. This model is consistent with what was observed in double-resistant wild-type clones, in which the two rapidly switching active BESs often were found in close proximity in the nucleus [ 12 ], and with the localization of a partially active BES at the periphery of the nucleolus [ 32 ]. Despite the localization in a Pol I competent compartment, BES17 is only partially activated, which suggests that another limiting factor is missing.…”

Epigenetic Regulation of Virulence Gene Expression in Parasitic Protozoa

“…In another experiment revealing communication among VSG s, artificial blockade of the active VSG -ES allowed the high-frequency activation of other VSG -ESs ( Aresta-Branco et al., 2016 ). These results are consistent with a remarkably robust exclusion mechanism operating in T. brucei and failure to select two stable, simultaneously active VSG -ESs ( Chaves et al., 1999 ).…”

Bloodstream Form-specific Up-regulation of Silent VSG Expression Sites and Procyclin in Trypanosoma brucei after Inhibition of DNA Synthesis or DNA Damage

“…Partial activation of ESs in bloodstream form T. brucei has been achieved before in experiments targeting drug resistance genes into "silent" ESs, although the levels of transcription were much lower than we show here (19,64). These low levels of transcription were "erased" after activation and subsequent inactivation of the partially active ES, implying the resetting of an epigenetic state (19).…”

A Histone Methyltransferase Modulates Antigenic Variation in African Trypanosomes

“…We speculate that BES17 is expressed either in the nucleolus or in the unique ESB, close to BES1. This model is consistent with what was observed in double-resistant wild-type clones, in which the two rapidly switching active BESs often were found in close proximity in the nucleus [ 12 ], and with the localization of a partially active BES at the periphery of the nucleolus [ 32 ]. Despite the localization in a Pol I competent compartment, BES17 is only partially activated, which suggests that another limiting factor is missing.…”

Epigenetic Regulation of Virulence Gene Expression in Parasitic Protozoa

“…In another experiment revealing communication among VSG s, artificial blockade of the active VSG -ES allowed the high-frequency activation of other VSG -ESs ( Aresta-Branco et al., 2016 ). These results are consistent with a remarkably robust exclusion mechanism operating in T. brucei and failure to select two stable, simultaneously active VSG -ESs ( Chaves et al., 1999 ).…”

Bloodstream Form-specific Up-regulation of Silent VSG Expression Sites and Procyclin in Trypanosoma brucei after Inhibition of DNA Synthesis or DNA Damage

“…Partial activation of ESs in bloodstream form T. brucei has been achieved before in experiments targeting drug resistance genes into "silent" ESs, although the levels of transcription were much lower than we show here (19,64). These low levels of transcription were "erased" after activation and subsequent inactivation of the partially active ES, implying the resetting of an epigenetic state (19).…”