Many Ways to Loop DNA

Abstract

In the 1960s, I developed methods for directly visualizing DNA and DNA-protein complexes using an electron microscope. This made it possible to examine the shape of DNA and to visualize proteins as they fold and loop DNA. Early applications included the first visualization of true nucleosomes and linkers and the demonstration that repeating tracts of adenines can cause a curvature in DNA. The binding of DNA repair proteins, including p53 and BRCA2, has been visualized at three-and four-way junctions in DNA. The trombone model of DNA replication was directly verified, and the looping of DNA at telomeres was discovered.

RecBCD is required to complete chromosomal replication: Implications for double-strand break frequencies and repair mechanisms

“…On encountering a Chi site, the nucleolytic activity of RecBCD is attenuated, while the helicase activity remains unaffected [10, 16, 85]. The differing speeds of the dual helicases can lead to DNA loops forming as the enzyme progressively tracks along the DNA [6, 86], a phenotype similar to that seen with replisomes and mismatch repair complexes that must track and maintain polarity on DNA over large distances [87–89] (Figure 5C). These features must have arisen under strong selective pressure for specific functions, many of which are not considered in current double-strand break repair models.…”

Transcription of telomeric DNA leads to high levels of homologous recombination and t-loops

“…Agarose gel electrophoresis of linear pRST5 (BsmBI cut, no 3′ overhang) DNA in transcription buffer alone (U). If the DNA was transcribed for 30 min (Materials and Methods) and then deproteinized but not treated with RNase A, the DNA with RNA bound was present as a smear whether or not it was crosslinked with psoralen and UV (1, 2). Following transcription, deproteinization and treatment with RNase A (3) the DNA was present as a ladder of bands with the monomer band shifted upward.…”

“…The telomere specific proteins in higher eukaryotes are present in one or more multi-protein complexes termed shelterins, as the DNA at the end of the telomere must be sheltered from erroneous recognition as a double strand break (1). This function is believed to be accomplished by the combination of shelterin binding and an architectural solution in which the DNA terminus folds back to generate a duplex loop (t-loop) that hides the DNA end (2,3). …”

“…T-loops have been isolated and visualized by electron microscopy (EM) from species ranging from yeast to humans as reviewed in (2,4). The t-loop junction can occur anywhere along the length of the telomere, generating a spectrum of circle sizes; in peas, t-loops as large as 120 kb with an 80 kb circular portion were observed (5).…”

“…Our earlier suggestion that telomeres might loop back on themselves (2,3) was based on the fact that most eukaryotic telomeres end with a 3′ single stranded (ss) tail on the G-rich strand which has the same sequence as the preceding duplex (ds) telomeric DNA (6). Based on our understanding of homologous recombination (HR) reactions, the 3′ ss tail would be expected to invade the preceding duplex segment to generate a lasso structure with a D-loop at the junction.…”