Nanopore sequencing reveals that DNA replication compartmentalisation dictates genome stability and instability inTrypanosoma brucei

Abstract

AbstractThe genome ofTrypanosoma bruceiis structurally complex. Eleven megabase-sized chromosomes each comprise a transcribed core flanked by silent subtelomeres, housing thousands ofVariant Surface Glycoprotein(VSG) genes. Additionally, silentVSGsare also found on hundreds of sub-megabase chromosomes that harbour 177 bp repeats of unknown function, and multipleVSGtranscription sites localise to the telomeres of both chromosome types. DNA replication dynamics have been described in the megabase chromosome cores but not in the subtelomeres or sub-megabase chromosomes, and targeted early replication of the single activeVSGtranscription site is unexplained. Here, using Nanopore assembly, we mapped DNA replication across this compartmentalised genome. We show that subtelomeres display a paucity of replication initiation events relative to the core, correlating with increased instability in the silentVSGarchive. In addition, early replication of the activeVSGtranscription site is shown to originate from the telomere, likely causing targetedVSGrecombination. Lastly, we demonstrate that the 177 bp repeats act as widespread, conserved DNA replication origins, explaining mitotic stability of the abundant small chromosomes and early DNA replication of megabase chromosome centromeres. Compartmentalized DNA replication dynamics therefore explains howT. bruceibalances stable genome transmission with localised instability driving immune evasion.