Formation and biological functions of unusual DNA/RNA structures formed by repeat sequences.

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More than 50% of most genomes are tandem repetitive DNA sequences, including telomeres, centromeres, macro-, mini- and microsatellite repeats. Repeat DNAs can exist in unusual structures beyond the canonical right-handed B-form double helix. The specific locations of repeats in genes and chromosomes highlight their biological importance, particularly with respect to the unusual structures they form. Evidence indicates that repeat structures play roles in gene regulation, splicing, recombination, epigenetic regulation, repeat length mutation, etc. Functions are likely to be reflected by the biophysical structures formed by the repeats. Importantly, the length of the repeat tracts are hyper-prone to change and these length changes can dramatically affect their biological function. Length changes likely involve unusual structures. Our goal is to understand unusual repeat structures and how they can change lengths. The formation of unusual repeat structures likely occurs during DNA replication, repair, recombination and transcription, when complementary strands become unpaired and single-stranded. Slipped-DNAs form by out-of-register misalignment of repeats. Newly transcribed RNAs can form unusual structures such as intra-strand hairpins and G-quadruplexes. Some DNA sequences can form transcriptionally induced RNA:DNA hybrids called R-loops. The specific locations of these sequences in genes and chromosomes highlight their biological importance, particularly with respect to the alternative structures they form, which are highly likely to have functional roles. These functions are poorly understood. Most structural analyses are performed on isolated DNA strands in the absence of their partner strand. Furthermore, most studies use short synthetic oligo repeats that are dramatically shorter than their biological counter-parts, which occur in the presence of both complementary strands. Such studies yield results that may only partially reflect what may occur biologically. A thorough understanding of the structures formed by these repeats, the proteins that interact and process them, must include studies using biologically relevant lengths of the complementary strands. With a focus upon a broad range of repeats (telomeres, centromeres, macro-, mini-, and micro-satellite repeats), that are and are not associated with disease, we will study the formation of unusual structures by the repeats; assess protein-nucleic acid interactions, as well as the roles of certain proteins in processing these unusual structures to repeat length changes. This proposal focuses upon the basic fundamental molecular and biological aspects of repeat sequences. The results of this proposal will shed light upon both the natural and disease related aspects of some of these repeats. Disease-related studies on a subset of the repeats studied herein are the focus of other grants.