Date of Award
Cross George Laboratory
trypanosoma brucei, African T. brucei, African sleeping sickness, telomere shortening
A variety of parasites persist in their host through sequential expression of variant surface antigens. Intriguingly, the genes encoding these antigens are frequently found adjacent to the telomeres. Telomeres are nucleoprotein complexes consisting of tandem DNA repeats and proteins that bind to them. Their function is to protect chromosome ends from the DNA repair machinery that would otherwise recognize them as double-stranded breaks. With the help of the ribonucleoprotein telomerase they compensate for the gradual sequence loss that would otherwise arise from the inability of conventional DNA polymerases to replicate chromosome ends. In Trypanosoma brucei, the causative agent of African trypanosomiasis, the surface coat consists of a dense layer of Variant Surface Glycoproteins (VSG). The actively transcribed VSG is found in one of ~20 telomeric Expression Sites (ES). Antigenic variation can occur by transcriptional switching, reciprocal translocations, or duplicative gene conversion events between ES. In recent African isolates, duplicative gene conversion occurs at a high frequency and predominates, but the switching frequency decreases dramatically upon laboratory-adaptation. Very little is known about the regulation of antigenic variation. To address whether telomeres are involved in antigenic switching we created telomerasedeficient T. brucei. Telomerase-deficient parasites exhibited progressive telomere shortening at a rate of 3â€“6 bp/PD, which correlates with the end replication problem and G-overhang length. Upon reaching a critical length, short silent ES telomeres stabilized. Telomere decline was accompanied by loss of minichromosomes and rearrangements at intermediate chromosomes. Essential megabase chromosomes remained stable. After extensive telomere attrition, the active ES telomere stabilized, but the transcribed VSG was gradually lost from the population and replaced by a new VSG through duplicative gene conversion. We present a model in which subtelomeric break-induced replication-mediated repair at a short ES telomere leads to duplicative gene conversion and expression of a new VSG. By restoring telomerase, we studied telomere elongation dynamics. At the active ES, the rate of telomere elongation is inversely proportional to the initial telomere length. At silent ES, the rate of elongation remains constant. We propose a model where transcription-dependent chromatin remodeling permits telomere elongation by telomerase. We show that telomere growth at a rate of 6â€“8 bp/PD appears to be a unique feature of T. brucei. Lastly we demonstrate that fast-switching African T. brucei isolates have dramatically shorter telomeres than laboratory strains. We present a speculative model in which telomere growth and breakage affect the rate of antigenic switching.
Dreesen, Oliver, "Telomere Structure and Shortening in Telomerase-Deficient Trypanosoma brucei: Implications for Anitgenic Variation" (2006). Student Theses and Dissertations. 50.