Polymerase theta-helicase promotes end joining by stripping single-stranded DNA-binding proteins and bridging DNA ends

Abstract

Homologous recombination-deficient cancers rely on DNA polymerase Theta (Pol{theta})-Mediated End Joining (TMEJ), an alternative double-strand break repair pathway. Pol{theta} is the only vertebrate polymerase that encodes an N-terminal superfamily 2 (SF2) helicase domain, but the role of this helicase domain in TMEJ remains unclear. Using single-molecule imaging, we demonstrate that Pol{theta}-helicase (Pol{theta}-h) is a highly processive single-stranded DNA (ssDNA) motor protein that can efficiently strip Replication Protein A (RPA) from ssDNA. Pol{theta}-h also has a limited capacity for disassembling RAD51 filaments but is not processive on doublestranded DNA. Pol{theta}-h can bridge two non-complementary DNA strands in trans. PARylation of Pol{theta}-h by PARP-1 resolves these DNA bridges. We conclude that Pol{theta}-h removes RPA and RAD51 filaments and mediates bridging of DNA overhangs to aid in polymerization by the Pol{theta} polymerase domain.