Analysis of diverse double-strand break synapsis with Polλ reveals basis for unique substrate specificity in nonhomologous end-joining

Using X-ray crystallography and nonhomologous end-joining assays, this study reveals structural features within Pol lambda that provide it with the ability to bridge and stabilize tenuous DNA double-strand break ends, allowing for religation. DNA double-strand breaks (DSBs) threaten genomic stability, since their persistence can lead to loss of critical genetic information, chromosomal translocations or rearrangements, and cell death. DSBs can be repaired through the nonhomologous end-joining pathway (NHEJ), which processes and ligates DNA ends efficiently to prevent or minimize sequence loss. Polymerase lambda (Pol lambda), one of the Family X polymerases, fills sequence gaps of DSB substrates with a strict specificity for a base-paired primer terminus. There is little information regarding Pol lambda's approach to engaging such substrates. We used in vitro polymerization and cell-based NHEJ assays to explore the contributions of conserved loop regions toward DSB substrate specificity and utilization. In addition, we present multiple crystal structures of Pol lambda in synapsis with varying biologically relevant DSB end configurations, revealing how key structural features and hydrogen bonding networks work in concert to stabilize these tenuous, potentially cytotoxic DNA lesions during NHEJ.

Automated summary

This study reveals the structural features of Pol lambda that allow it to bridge and stabilize DNA double-strand break ends, enabling the repair of genomic instability. The findings provide valuable insights into the mechanisms of DNA repair.