Energy Landscapes of Dynamic Ensembles of Rolling Triplet Repeat Bulge Loops: Implications for DNA Expansion Associated with Disease States

Abstract

DNA repeat domains can form ensembles of canonical and noncanonical states, including stable and metastable DNA secondary structures. Such sequence-induced structural diversity creates complex conformational landscapes for DNA processing pathways, including those triplet expansion events that accompany replication, recombination, and/or repair. Here we demonstrate further levels of conformational complexity within repeat domains. Specifically, we show that bulge loop structures within an extended repeat domain can form dynamic ensembles containing a distribution of loop positions, thereby yielding families of positional loop isomers, which we designate as “rollamers”. Our fluorescence, absorbance, and calorimetric data are consistent with loop migration/translocation between sites within the repeat domain (“rollamerization”). We demonstrate that such “rollameric” migration of bulge loops within repeat sequences can invade and disrupt previously formed base-paired domains via an isoenthalpic, entropy-driven process. We further demonstrate that destabilizing abasic lesions alter the loop distributions so as to favor “rollamers” with the lesion positioned at the duplex/loop junction, sites where the flexibility of the abasic “universal hinge” relaxes unfavorable interactions and/or facilitates topological accommodation. Another strategic siting of an abasic site induces directed loop migration toward denaturing domains, a phenomenon that merges destabilizing domains. In the aggregate, our data reveal that dynamic ensembles within repeat domains profoundly impact the overall energetics of such DNA constructs as well as the distribution of states by which they denature/renature. These static and dynamic influences within triplet repeat domains expand the conformational space available for selection and targeting by the DNA processing machinery. We propose that such dynamic ensembles and their associated impact on DNA properties influence pathways that lead to DNA expansion.

APE1 Incision Activity at Abasic Sites in Tandem Repeat Sequences

“…3A and 6A) on APE1 incision activity. We showed previously for a CAG8 oligomer annealed to a “CTGN” strand (N = 2, 4, 6), that the repeat bulge loop can exist in multiple isoenergentic structural arrangements, with relatively facile inter-conversion between them, prompting the designation “rollamers” ( 31 ). The relative population of loop isomers is influenced by the relative thermodynamic impact of the abasic site and its position within the repeat, with the lesion favoring isomer states that are least energetically perturbed.…”

“…The relative population of loop isomers is influenced by the relative thermodynamic impact of the abasic site and its position within the repeat, with the lesion favoring isomer states that are least energetically perturbed. CAG6F1, CAG6F3, and CAG6F5 annealed to CTG2 result in the formation of loop isomers that differ structurally in that the abasic site can be located in different structural elements of the overall bulge loop construct: i.e., in the upstream duplex (CAG6F1), the 5′ duplex loop junction (CAG6F1), the loop (CAG6F1, CAG6F3, CAG6F5), the 3′ duplex loop junction (CAG6F5), or the downstream duplex domain (CAG6F5) ( 31 ).…”

“…Oligonucleotides were purchased from Integrated DNA Technologies (Coralville, IA) or synthesized as described ( 29, 31 ). Oligonucleotide names and sequences are listed in Table 2.…”

Parity-dependent hairpin configurations of repetitive DNA sequence promote slippage associated with DNA expansion

“…DNA slippage is believed to be a primary mechanism driving the change in repeat number of various unit sizes. Repetitive DNA sequences often form alternative structures such as bulges and hairpin loops in addition to canonical DNA conformations (7,8). A repeat unit may slip between being part of a hairpin loop, a bulge, or a duplex in a dynamic fashion, which may alter the course of normal cellular DNA chemistry and ultimately lead to repeat expansion associated with neurological diseases (9).…”

Somatic microsatellite variability as a predictive marker for colorectal cancer and liver cancer progression

“…MSTs are mutational “hot-spots”, meaning they experience a significantly greater rate of somatic variability and population polymorphism than adjacent non-repetitive DNA [11-14]. The unique repetitive genomic configuration of MSTs can lead to the development of complex DNA structures susceptible to polymerase slippage and DNA breaks [11, 15-17]. This results in a distinct mutational profile for MSTs with a bias for indels, as opposed to single nucleotide polymorphisms (SNPs) which are frequently observed in non-repetitive DNA [18].…”