Cohesin regulatesVSGmonoallelic expression in trypanosomes

Abstract

Antigenic variation allows Trypanosoma brucei to evade the host immune response by switching the expression of 1 out of ∼15 telomeric variant surface glycoprotein (VSG) expression sites (ESs). VSG ES transcription is mediated by RNA polymerase I in a discrete nuclear site named the ES body (ESB). However, nothing is known about how the monoallelic VSG ES transcriptional state is maintained over generations. In this study, we show that during S and G2 phases and early mitosis, the active VSG ES locus remains associated with the single ESB and exhibits a delay in the separation of sister chromatids relative to control loci. This delay is dependent on the cohesin complex, as partial knockdown of cohesin subunits resulted in premature separation of sister chromatids of the active VSG ES. Cohesin depletion also prompted transcriptional switching from the active to previously inactive VSG ESs. Thus, in addition to maintaining sister chromatid cohesion during mitosis, the cohesin complex plays an essential role in the correct epigenetic inheritance of the active transcriptional VSG ES state.

Cell Biology of the Trypanosome Genome

“…In agreement with this model, a partial reduction in the levels of the cohesin subunit TbSCC1 leads to the production of two ESBs in post-S-phase cells, presumably due to premature separation of the chromatids carrying the active ES (97). However, the full inheritance of transcriptional status does not appear to be immediate at replication, since reduction of chromatid cohesion also encourages VSG expression site switching (97). Derepression of BESs by knockdown of the telomeric protein RAP1 also leads to more than one extranucleolar pol I focus (193), suggesting a role for telomeric chromatin in maintenance of the singularity of the ESB.…”

“…With a model of a self-organized ESB, however, it can more readily be seen how inheritance of transcriptional status at replication would lead to two daughter ESBs following sister chromatid separation at mitosis. In agreement with this model, a partial reduction in the levels of the cohesin subunit TbSCC1 leads to the production of two ESBs in post-S-phase cells, presumably due to premature separation of the chromatids carrying the active ES (97). However, the full inheritance of transcriptional status does not appear to be immediate at replication, since reduction of chromatid cohesion also encourages VSG expression site switching (97).…”

Trypanosoma brucei TIF2 suppresses VSG switching by maintaining subtelomere integrity

“…The predominant VSG switching pathway is ES GC/ES loss + in situ switching in TbTIF2-depleted cells, which is consistent with this hypothesis. Interestingly, partial depletion of TbSCC1 or other cohesin subunits TbSMC1 and TbSCC3 also led to premature dissociation of sister chromatids at the active BES and increased VSG switching frequency [58], although in situ switching appears to be the preferred pathway in cohesion-defective cells, which is not exactly the same as in TbTIF2-depleted cells, where a significant portion of the VSG switchers arose from ES loss + in situ or ES GC switching. Our first model that TbTIF2 is important for telomeric DNA replication suggests that more DSB formation is the reason for more subtelomeric DSBs in TbTIF2-depleted cells, while the second model suggests that less DSB repair is the reason.…”

Third target of rapamycin complex negatively regulates development of quiescence in Trypanosoma brucei

“…Cellular localization analysis was performed by 3D immunofluorescence as described previously (29). Images displayed in the figures are maximum intensity projections from digitally deconvolved multichannel 3D image data sets using Huygens Essential software v. 2.9 (Scientific Volume Imaging).…”