Abstract
Expansion of CAG/CTG trinucleotide repeats causes certain familial neurological disorders. Hairpin formation in the nascent strand during DNA synthesis is considered a major path for CAG/CTG repeat expansion. However, the underlying mechanism is unclear. We show here that removal or retention of a nascent strand hairpin during DNA synthesis depends on hairpin structures and types of DNA polymerases. Polymerase (pol) δ alone removes the 3'-slipped hairpin using its 3'-5' proofreading activity when the hairpin contains no immediate 3' complementary sequences. However, in the presence of pol β, pol δ preferentially facilitates hairpin retention regardless of hairpin structures. In this reaction, pol β incorporates several nucleotides to the hairpin 3'-end, which serves as an effective primer for the continuous DNA synthesis by pol δ, thereby leading to hairpin retention and repeat expansion. These findings strongly suggest that coordinated processing of 3'-slipped (CAG)n/(CTG)n hairpins by polymerases δ and β on during DNA synthesis induces CAG/CTG repeat expansions.
Highlights
Expansion of coordinated processing of 3-slipped (CAG)/coordinated processing of 3-slipped (CAG)n/ (CTG) repeats causes familial neurological disorders, but the molecular basis is unknown
These findings strongly suggest that coordinated processing of 3-slipped (CAG)n/ (CTG)n hairpins by polymerases ␦ and  on during DNA synthesis induces CAG/CTG repeat expansions
Polymerase  Promotes CAG/CTG Repeat Expansion in Nuclear Extract-catalyzed DNA Synthesis—To explore how human cells process a CAG or CTG hairpin formed in the nascent strand at the site of DNA synthesis, a CTG or CAG hairpin substrate that mimics the nascent strand hairpin formation was constructed
Summary
Expansion of CAG/CTG repeats causes familial neurological disorders, but the molecular basis is unknown. In addition to DNA strand breaks, which provide opportunities for strand slippage, these processes share a common feature in DNA synthesis, a reaction catalyzed by DNA polymerases Taken together, these studies suggest that the disease-causing CAG/CTG repeat expansion is likely originated from hairpin formations in the nick-residing nascent strand and subsequent error-prone DNA synthesis by DNA polymerases. The left panel shows the BbvI cleavage products of a (CTG)10/(CAG) homoduplex and a (CTG)15/(CAG) heteroduplex on a 15% denaturing polyacrylamide gel, and the right panel diagrams the recognition sites and the predicted cleavage sites of the (CTG)10/(CAG) homoduplex by BbvI Their distinct roles in DNA metabolic processes, recent evidence suggests that a transient switching between TLS polymerases and replicative polymerases occurs to deal with bulky DNA lesions during DNA synthesis [19]. This synergistic stimulation suggests a concerted cooperation between pol  and pol ␦, leading a hairpin retention and TNR expansion
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