Diversity and dynamics of the minichromosomal karyotype in Trypanosoma brucei

Abstract

The genome of African trypanosomes contains a large number of minichromosomes. Their only proposed role is in the expansion of the parasites' repertoire of telomeric variant surface glycoprotein (VSG) genes as minichromosomes carry silent VSG gene copies in telomeric locations. Despite their importance as VSG gene donors, little is known about the actual composition of the minichromosomal karyotype and the stability of its inheritance. In this study we show, by using high-resolution pulsed-field electrophoresis, that a non-clonal trypanosome population contains an extremely diverse pattern of minichromosomes, which can be resolved into less complex clone-specific karyotypes by non-selective cloning. We show that the minichromosome patterns of such clones are stable over at least 360 generations. Furthermore, using DNA markers for specific minichromosomes, we demonstrate the mitotic stability of these minichromosomes within the population over a period of more than 5 years. Length variation is observed for an individual minichromosome and is most likely caused by a continuous telomeric growth of approximately 6 bp per telomere per cell division. This steady telomeric growth, counteracted by stochastic large losses of telomeric sequences is the most likely cause of minichromosome karyotype heterogeneity within a population.

Inactivation of Mre11 Does Not Affect VSG Gene Duplication Mediated by Homologous Recombination in Trypanosoma brucei

“…The latter two repeat sequences are thought to have important roles in gene rearrangements and as insulators, respectively. The potential importance of the minichromosome repeats is seen in the fact that they are remarkably stable in size, despite the repeats being identical in sequence over most of their length (64). Even though Mre11 is thought to help in the removal of secondary structures arising in some types of repeat, thus preventing replication errors (67,68), we find no such problems arise in trypanosome minichromosomes when Mre11 is absent (data not shown).…”

“…A second genomic feature in the trypanosome that may relate to this proposed demarcation between repair and recombination is the abundance of repetitive sequence, estimated at 32% (62). Much of this is the satellite sequence that constitutes most of the structure of the ϳ100 nuclear minichromosomes (63,64), whereas other highly repetitive regions flank VSG genes (65, 66) and bloodstream VSG transcription units (61). The latter two repeat sequences are thought to have important roles in gene rearrangements and as insulators, respectively.…”

Cell Biology of the Trypanosome Genome

“…The MC and IC populations are VOL. 74,2010 CELL BIOLOGY OF THE TRYPANOSOME GENOME 553 probably aneuploid, being inherited in an apparently nonMendelian fashion (186) and showing a mixed ploidy for genetic markers (2). The genetic information of T. brucei totals ϳ35 Mb (per haploid genome) and is enclosed in a nucleus with a diameter of ϳ2.5 m (51,138).…”

Telomere as an Important Player in Regulation of Microbial Pathogen Virulence

“…There are 11 pairs of megabase chromosomes (0.9-5.7 Mb), several intermediate chromosomes (300-900 kb), and ~100 copies of minichromosomes (50-100 kb) in T. brucei genome (Alsford et al 2001;Berriman et al 2005;Melville et al 2000). The majority of VSG genes are found in long tandem arrays of repeated genes at subtelomeric locations on megabase chromosomes (Fig.…”

“…Approximately 200 copies of VSG genes are found immediately upstream of telomeres of the minichromosomes, which carry besides the VSG genes, only repetitive sequences, including 177 bp repeats in the chromosome internal region and telomere repeats at the chromosome ends (Fig. 7D) (Alsford et al 2001). The rest of VSGs are found in VSG expression sites.…”