ABSTRACT
Ultraviolet (UV) radiation generates crosslinked DNA lesions—primarily cyclobutane pyrimidine dimers (CPDs) and [6–4] photoproducts ([6–4] PPs)—that block the progression of replicative DNA polymerases. In plants, these lesions are efficiently removed from nuclear DNA by dedicated repair pathways; however, comparable repair mechanisms are absent in plastids and mitochondria. Consequently, how plant organellar DNA polymerases (POPs) tolerate or bypass UV-induced damage has remained unclear. Here, we show that the two Arabidopsis thaliana organellar polymerases, AtPolIs, possess robust translesion synthesis (TLS) activity across CPDs. Although wild-type enzymes display only limited extension across [6–4] PPs, removal of their exonuclease function dramatically enhances bypass, yielding an efficiency of replication across the [6–4] PP that closely resembles that observed on an undamaged template. This establishes AtPolI as the first known replicative DNA polymerase capable of efficiently bypassing a [6–4] PP. We further demonstrate that TLS across UV photoproducts relies on three unique amino acid insertions within the AtPolI polymerase domain, as deletion of any single insertion abolishes TLS. Notably, Mn²⁺ can restore TLS activity in these variants, but only for CPD lesions. Together, these findings identify AtPolIs as the first plant organellar replicases with intrinsic [6–4] PP bypass capability and define the structural features that enable this function.
GRAPHICAL ABSTRACT
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