Poster abstracts

Poster number 157 submitted by Sk Abdus Sayeed

Investigating TERRA functions in VSG switching and telomere maintenance in Trypanosoma brucei

SK ABDUS SAYEED (Cleveland State University), Bibo Li (Cleveland State University)

Human African Trypanosomiasis (HAT) is caused by the protozoan parasite Trypanosoma brucei, which regularly switches its major surface antigen (variant surface glycoprotein, VSG) to evade the human immune response. VSGs are expressed from subtelomeric loci in a monoallelic manner. Work from our lab has shown that telomere proteins play important roles in VSG monoallelic expression and VSG switching. The active VSG gene is flanked by upstream 70 bp repeats and downstream telomeric repeats. The telomeric repeats downstream of the active VSG are transcribed into the telomeric repeat-containing RNA (TERRA), a long non-coding RNA, as a result of read-through transcription by RNA polymerase I. It has been shown that TbRAP1 and TbTRF, two telomere proteins, suppress TERRA and telomeric R-loop (RNA: DNA hybrid) levels, thereby suppressing DNA recombination mediated VSG switching events. We previously showed that excessive amounts of TERRA and the telomeric R-loops cause more DNA breaks at the telomere and threatens cell viability. Recently, we successfully generated T. brucei strains with LacO repeats targeted to the subtelomeric region immediately downstream of the active VSG and immediately upstream of the telomere repeats. We observed that inducing the LacI-GFP expression significantly reduces the TERRA level, presumably because binding of LacI-GFP to the LacO repeats blocks the read-through transcription of the telomeric repeats downstream of the active VSG gene. This is the first conditional repression system established in trypanosomes, which effectively solves the problem of inefficient depletion of TERRA by RNAi. TERRA depletion does not affect T. brucei growth. However, TERRA depletion increases the TbTRF protein level and the amount of telomere bound TbTRF. Whether TERRA depletion affects the VSG switching frequency and/or telomere length maintenance are under investigation now.

Keywords: TERRA, Telomere