Non-Canonical G-quadruplexes cause the hCEB1 minisatellite instability in <i>Saccharomyces cerevisiae</i>.

Abstract

G-quadruplexes (G4) are polymorphic four-stranded structures formed by certain G-rich nucleic acids in vitro, but the sequence and structural features dictating their formation and function in vivo remains uncertain. Here we report a structure-function analysis of the complex hCEB1 G4-forming sequence. We isolated four G4 conformations in vitro, all of which bear unusual structural features: <i>Form 1</i> bears a V-shaped loop and a snapback guanine; <i>Form 2</i> contains a terminal G-triad; <i>Form 3</i> bears a zero-nucleotide loop; and <i>Form 4</i> is a zero-nucleotide loop monomer or an interlocked dimer. In vivo, <i>Form 1</i> and <i>Form 2</i> differently account for 2/3^rd of the genomic instability of hCEB1 in two G4-stabilizing conditions. <i>Form 3</i> and an unidentified form contribute to the remaining instability, while <i>Form 4</i> has no detectable effect. This work underscores the structural polymorphisms originated from a single highly G-rich sequence and demonstrates the existence of non-canonical G4s in cells, thus broadening the definition of G4-forming sequences.