Discussion

The genome of every organism is subject to damage, which is tackled either by repair or tolerance. Many DNA repair pathways have been documented inTrypanosoma brucei but less attention has been paid to damage tolerance, a reaction in which lesion bypass is needed, in particular to ensure continued genome replication. Such bypass is promoted by translesion DNA polymerases (TLS Pols), of which five putative examples can be found in T. brucei. We have used RNAi to examine the function of three proteins in bloodstream forms of T. brucei. Loss of PolN (Nu) was shown to be severely detrimental to growth, with accumulation of cells showing aberrant nuclei, suggesting a critical role in nuclear genome maintenance. RNAi of PolZ (zeta) did not impair growth, but resulted in increased sensitivity to methyl methanesulphonate (MMS) damage, suggesting a role in the response to alkylation. The sequence of PolQ (theta) suggests that the predicted protein may not be a joint polymerase-helicase like in other eukaryotes, but only a helicase. RNAi revealed that loss of the factor did not affect growth, nor did it result in increased MMS sensitivity. Taken together, these data reveal widespread and variant functions for three of five putative TLS DNA polymerases in T. brucei genome biology.