G-quadruplexes are four-stranded nucleic acid structures involved in multiple cellular pathways including DNA replication and telomere maintenance. Such structures are formed by G-rich DNA sequences typified by telomeric DNA repeats. Whilst there is evidence for proteins that bind and regulate G-quadruplex formation, the molecular basis for this remains poorly understood. The budding yeast telomeric protein Rap1, originally identified as a transcriptional regulator functioning by recognizing double-stranded DNA binding sites, was one of the first proteins to be discovered to also bind and promote G-quadruplex formation in vitro. Here, we present the 2.4 angstrom resolution crystal structure of the Rap1 DNA-binding domain in complex with a G-quadruplex. Our structure not only provides a detailed insight into the structural basis for G-quadruplex recognition by a protein, but also gives a mechanistic understanding of how the same DNA-binding domain adapts to specifically recognize different DNA structures. The key observation is the DNA-recognition helix functions in a bimodal manner: In double-stranded DNA recognition one helix face makes electrostatic interactions with the major groove of DNA, whereas in G-quadruplex recognition a different helix face is used to make primarily hydrophobic interactions with the planar face of a G-tetrad.

Several transcription factors ( TF s) have been implicated in neuroectoderm ( NE ) development, and recently, the TF PAX 6 was shown to be critical for human NE specification. However, micro RNA networks regulating human NE development have been poorly documented. We hypothesized that micro RNA s activated by PAX 6 should promote NE development. Using a genomics approach, we identified PAX 6 binding sites and active enhancers genome‐wide in an in vitro model of human NE development that was based on neural differentiation of human embryonic stem cells ( hESC ). PAX 6 binding to active enhancers was found in the proximity of several micro RNA s, including hsa‐mi R ‐135b. Mi R ‐135b was activated during NE development, and ectopic expression of mi R ‐135b in hESC promoted differentiation toward NE . Mi R ‐135b promotes neural conversion by targeting components of the TGF ‐β and BMP signaling pathways, thereby inhibiting differentiation into alternate developmental lineages. Our results demonstrate a novel TF ‐mi RNA module that is activated during human neuroectoderm development and promotes the irreversible fate specification of human pluripotent cells toward the neural lineage.