A single G-to-C change causes human centromere TGGAA repeats to fold back into hairpins.

Abstract

Recently, we established that satellite III (TGGAA). tandem repeats, which occur at the centromeres of human chromosomes, pair with themselves to form an unusual "self-complementary" antiparallel duplex containing (GGA)2 motifs in which two unpaired guanines from opposite strands intercalate between sheared GoA base pairs. In separate studies, we have also established that the GCA triplet does not form bimolecular (GCA)2 motifs but instead promotes the formation of hairpins containing a GCA-turn motif in which the loop contains a single cytidine closed by a sheared GoA pair. Since TGCAA is the most frequent variant of TGGAA found in satellite III repeats, we reasoned that the potential of this variant to form GCA-turn miniloop fold-back structures might be an important factor in modulating the local structure in natural (TGGAA The classical satellite III of human DNA, which has been located in the centromere region of chromosomes (1), is composed of (TGGAA)n repeats with the consensus sequence 5'-CAACCCGAM(TGGAA), (2). From the cooperative UVmelting transition Grady et al. (1) found that, in the absence of the complementary TTCCA strand, (TGGAA)6 alone can somehow pair with itself to form a stable duplex with a Tm of 65°C. We recently solved the solution structure of the antiparallel "self-complementary" (TGGAATGGAA)2 duplex containing a tandem TGGAA repeat; this structure contains two (GGA)2 motifs in which the central guanines from opposite strands are unpaired and intercalatively stacked on each other between flanking GoA pairs (3, 4). The flanking G°A pairs are in the sheared, or side-by-side, configuration (5-7) and the guanines of the (GGA)2 motif form a four-guanine interstrand stack. Although the (TGGAA)n repeat is highly conserved in satellite III, perfect repeats rarely continue for more than 12-15 repeats (60-75 bp) with several variants that differ from the consensus pentamer by only a single base. The most frequent among these variants is TGCAA-i.e., the central GGA triplet [which forms the self-paired (GGA)2 motifl is changed to GCA. Interestingly, we recently found that the triplet GCA is a very strong hairpin promoter (8, 9). Among the four GNA triplets in a d(NAATGNAATG) sequence context, GGA predominantly forms bimolecular duplexes containing a (GGA)2 motif, as described previously (3, 4), while GCA forms a unimolecular hairpin (8, 9) and GAA and GTA form a mixture of mostly hairpin in equilibrium with a minor (GNA)2 duplex form (9). If (TGGAA), repeats self-pair to form antiparallel (GGA)2 motifs in vivo, we reasoned that occasional TGCAA pentamers interspersed among (TG-GAA), repeats might promote folding back of the chain on itself by forming GCA-turn loops, and thus modulate centromere folding. In the present paper, we have used NMR, distance geometry, restrained molecular dynamics, and backcalculation refinement to study the solution structure of the DNA heptadecamer (G)TGGAATGCAATGGAA(C), which contains three centromeric pentamer repeats, the middle one of which is changed from TG...

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

“…By combining this information with our high-resolution structure of the dTGGAA repeat duplex, the structural basis of the molecular behavior observed in the smFRET experiments becomes clear. For sequences containing an odd number of dTGGAA repeats, the end-to-end configuration is optimal because it forms the highest number of duplex interactions while maintaining the 1-nt loop structure proposed by Zhu et al (11). In contrast, sequences containing an even number of dTGGAA repeats can either maintain the 1-nt loop structure and leave a dangling overhang or maintain the end-to-end configuration and form octaloop structures (Fig.…”

“…Because dTGGAA repeat expansion is associated with the neurodegenerative disease SCA31 (13), understanding its structural and chemical bases of configuration slippage may also be important in a physiological context. It has been shown by NMR that dTGGAA tends to form a hairpin with a single G residue in the loop closed by a sheared G:A mismatch (11,20). Nucleic acids that contain 5′-GGA/AGG-5′ or 5′-GAAA/AAAG-5′ motifs can form antiparallel duplexes stabilized by unpaired purine bases that extend their stacking interactions until reaching the sheared G:A base pair.…”

“…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). (TGGAA) n repeats, for example, may form noncanonical structures such as a hairpin arm (10,11) or an antiparallel duplex (12). Expansion of this pentanucleotide sequence has been associated with spinocerebellar ataxia 31 (SCA31), an adult-onset autosomal-dominant neurodegenerative disorder (13).…”

Sheared purine·purine pairing in biology

“…While DNA sequences with three TGGAA repeats bracketed by a Watson-Crick G Á C pair exclusively form duplex structures with three (GGA) 2 intercalation motifs, changing the central G of such heptadecamers to C converts them completely to hairpins that contain a (GGA) 2 intercalation motif in the stem and a GCA-turn single-residue loop (Chou et al, 1996b;Zhu et al, 1996). This extraordinary hairpin-promoting capability of the (GCA) triplet may perhaps be used to fold back (TGGAA) tandem repeats and perhaps cause the condensation of human centromeres.…”

“…However, recent experiments have shown that sheared G ÁA base-pairing is an exception to this rule. The special geometry of sheared or side-by-side G Á A pairing has been found to be quite stable and compatible with¯anking WatsonCrick base-paired B-form DNA structure either as tandem base-pairs in (CGAG) 2 or (TGAA) 2 contexts (Li et al, 1991a,b;Chou et al, 1992aChou et al, ,b, 1994aCheng et al, 1992;Lane et al, 1992Lane et al, , 1994Katahira et al, 1993;Green et al, 1994), as base-pairs closing single-residue hairpin loops (Hirao et al, 1994;Zhu et al, 1995aZhu et al, , 1996Chou et al, 1996a,b), as basepairs closing two-residue hairpin loops (Hirao et al, 1989(Hirao et al, , 1990, as base-pairs bracketing unpaired purines (Chou et al, 1994b;Zhu et al, 1995bZhu et al, , 1996, or as base-pairs bracketing a single unpaired purine (our unpublished results). Due to their great potential in important biological functions such as DNA replication and centromere formation, we will describe structural characteristics of this type of basepairing and discuss its occurrence in the chromosomes of several single-stranded DNA viruses and in the centromere of human chromosomes.…”