👤 Amanda Vinson

Single-stranded DNA (ssDNA) containing four guanine repeats can form G-quadruplex (G4) structures. While cellular proteins and small molecules can bind G4s, it has been difficult to broadly assess their DNA-binding specificity. Here, we use custom DNA microarrays to examine the binding specificities of proteins, small molecules, and antibodies across ∼15,000 potential G4 structures. Molecules used include fluorescently labeled pyridostatin (Cy5-PDS, a small molecule), BG4 (Cy5-BG4, a G4-specific antibody), and eight proteins (GST-tagged nucleolin, IGF2, CNBP, FANCJ, PIF1, BLM, DHX36, and WRN). Cy5-PDS and Cy5-BG4 selectively bind sequences known to form G4s, confirming their formation on the microarrays. Cy5-PDS binding decreased when G4 formation was inhibited using lithium or when ssDNA features on the microarray were made double-stranded. Similar conditions inhibited the binding of all other molecules except for CNBP and PIF1. We report that proteins have different G4-binding preferences suggesting unique cellular functions. Finally, competition experiments are used to assess the binding specificity of an unlabeled small molecule, revealing the structural features in the G4 required to achieve selectivity. These data demonstrate that the microarray platform can be used to assess the binding preferences of molecules to G4s on a broad scale, helping to understand the properties that govern molecular recognition. Show less

Whereas the metabolic consequences of obesity have been studied extensively in the rhesus macaque, corollary genetic studies of obesity are nonexistent. This study assessed genetic contributions to spontaneous adiposity in this species. Phenotypic variation by age class and sex for BMI, waist to height ratio, waist to thigh ratio, and waist circumference was assessed in 583 macaques. Total and sex-specific heritability for all traits was estimated, including waist to thigh ratio adjusted for BMI, as well as genotypic and phenotypic correlations. In addition, functional genetic variation at BDNF, FTO, LEP, LEPR, MC4R, PCSK1, POMC, and SIM1 was assessed in four animals with extreme spontaneous adiposity. Trait heritability in the combined sample was low to moderate (0.14-0.32), whereas sex-specific heritability was more substantial (0.20-0.67). Heritability was greater in females for all traits except BMI. All traits were robustly correlated, with genetic correlations of 0.63 to 0.93 indicating substantial pleiotropy. Likely functional variants were discovered in the four macaques at all eight human obesity genes, including six missense mutations in BDNF, FTO, LEP, LEPR, and PCSK1 and, notably, one nonsense mutation in LEPR. A moderate polygenic contribution to adiposity in rhesus macaques was found, as well as mutations with potentially larger effects in multiple genes that influence obesity in humans. Show less