High-Quantum-Yield Ultrasmall Ln2O3 (Ln = Eu, Tb, or Dy) Nanoparticle Colloids in Aqueous Media Obtained via Photosensitization.

Fluorescent nanoparticles used in biomedical applications should be stable in their colloidal form in aqueous media and possess a high quantum yield (QY). We report ultrasmall Ln2O3 (Ln = Eu, Tb, or Dy) nanoparticle colloids with high QYs in aqueous media. The nanoparticles are grafted with hydrophilic and biocompatible poly(acrylic acid) (PAA) to ensure colloidal stability and biocompatibility and with organic photosensitizer 2,6-pyridinedicarboxylic acid (PDA) for achieving a high QY. The PAA/PDA-Ln2O3 nanoparticle colloids were nearly monodispersed and ultrasmall (particle diameter: ∼2 nm). They exhibited excellent colloidal stability with no precipitation after synthesis (>1.5 years) in aqueous media, very low cellular toxicity, and very high absolute QYs of 87.6, 73.6, and 2.8% for Ln = Eu, Tb, and Dy, respectively. These QYs are the highest reported so far for lanthanides in aqueous media. Therefore, the results suggest their high potential as sensitive optical or imaging probes in biomedical applications.

Biophysical investigation of the dual binding surfaces of human transcription factors FOXO4 and p53.

Cellular senescence is protective against external oncogenic stress, but its accumulation causes aging-related diseases. Forkhead box O4 (FOXO4) and p53 are human transcription factors known to promote senescence by interacting with each other and activating p21 transcription. Inhibition of the interaction is a strategy for inducing apoptosis of senescent cells, but the binding surfaces that mediate the FOXO4-p53 interaction remain elusive. Here, we investigated two binding sites involved in the interaction between FOXO4 and p53 by NMR spectroscopy. NMR chemical shift perturbation analysis showed that the binding between FOXO4's forkhead domain (FHD) and p53's transactivation domain (TAD), and between FOXO4's C-terminal transactivation domain (CR3) and p53's DNA-binding domain (DBD), mediate the FOXO4-p53 interaction. Isothermal titration calorimetry data showed that both interactions have micromolar Kd values, and FOXO4 FHD-p53 TAD interaction has a higher binding affinity. We also showed that the intramolecular CR3-binding surface of FOXO4 FHD interacts with p53 TAD2, and FOXO4 CR3 interacts with the DNA/p53 TAD-binding surface of p53 DBD, suggesting a network of potentially competitive and/or coordinated interactions. Based on these results, we propose that a network of intramolecular and intermolecular interactions contributes to the two transcription factors' proper localisation on the p21 promoter and consequently promotes p21 transcription and cell senescence. This work provides structural information at the molecular level that is key to understanding the interplay of two proteins responsible for cellular senescence.

FOXO4 Transactivation Domain Interaction with Forkhead DNA Binding Domain and Effect on Selective DNA Recognition for Transcription Initiation.

Forkhead box O4 (FOXO4) is a human transcription factor (TF) that participates in cell homeostasis. While the structure and DNA binding properties of the conserved forkhead domain (FHD) have been thoroughly investigated, how the transactivation domain (TAD) regulates the DNA binding properties of the protein remains elusive. Here, we investigated the role of TAD in modulating the DNA binding properties of FOXO4 using solution NMR. We found that TAD and FHD form an intramolecular complex mainly governed by hydrophobic interaction. Remarkably, TAD and DNA share the same surface of FHD for binding. While FHD did not differentiate binding to target and non-target DNA, the FHD-TAD complex showed different behaviors depending on the DNA sequence. In the presence of TAD, free and DNA-bound FHD exhibited a slow exchange with target DNA and a fast exchange with non-target DNA. The interaction of the two domains affected the kinetic function of FHD depending on the type of DNA. Based on these findings, we suggest a transcription initiation model by which TAD modulates FOXO4 recognition of its target promoter DNA sequences. This study describes the function of TAD in FOXO4 and provides a new kinetic perspective on target sequence selection by TFs.