Genome maintenance functions ofTrypanosoma bruceiDNA Polymerase N include telomere association and a role in antigenic variation

Abstract

Richard McCulloch: phone +44 (0) 1413305946; email: Richard.McCulloch@glasgow.ac.uk # These authors contributed equally to this work. AbstractMaintenance of genome integrity is critical to guarantee transfer of an intact genome from parent to offspring during cell division. DNA polymerases (Pols) provide roles in both replication of the genome and the repair of a wide range of lesions. Amongst replicative DNA Pols, translesion DNA Pols play a particular role: replication to bypass DNA damage, often at the cost of mutation. All cells express a range of translesion Pols, but little work has examined their function in parasites, including whether the enzymes might contribute to hostparasite interactions. Here, we describe a dual function of translesion PolN in African trypanosomes. Previously we demonstrated that PolN is associated with telomeric sequences and now we show that RNAi-mediated depletion of PolN results in slowed growth, altered DNA content, changes in cell morphology, and increased sensitivity to DNA damaging agents. Depletion of PolN leads to chromosome segregation defects and accumulation of DNA damage. We also show that PolN displays discrete localisation at the nuclear periphery in the absence of exogenous DNA damage. In addition, we demonstrate that PolN depletion leads to deregulation of telomeric variant surface glycoprotein genes, linking the function of this translesion DNA polymerase to host immune evasion by antigenic variation. Introduction Accurate duplication of the genome is a critical component of the cell cycle of all organisms. Two pathways contribute to accurate genome duplication: copying of the genome, and repair of DNA damage. Eukaryotic cells encode a wide range of DNA polymerases (Pols) that are required for DNA synthesis, allowing genome duplication, and for repair of DNA damage (review in [1]). Eukaryotic DNA Pols are divided into four different families (A, B, X and Y) based on sequence and structural homologies. Nuclear DNA Pols that direct the accurate copying of the genome belong to the B family, while mitochondrial genome replication is catalysed by an A family DNA Pol [2]. DNA Pols that act in DNA repair span all families, as do so-called translesion DNA Pols, which straddle DNA repair and replication activities because their activity is required whenever replicative DNA Pols encounter lesions in the template strand that must be bypassed to allow genome duplication [3];[4];[5]. In general,DNA replication is a high fidelity process with an extremely low error rate [6]. This is due to a combination of the ability of replicative DNA Pols to efficiently select the correct nucleotide to incorporate into the newly synthesized DNA strand and proofreading activity of the Pols, which permits the excision of occasionally incorrectly inserted nucleotides. Additionally, postreplicative repair mechanisms further reduce overall error rates by removing mispaired or damaged bases [7]. Although the wide range of DNA repair mechanisms available to all cells can efficiently detect and rem...