Hermann Habacher

Transcription factors play key roles in orchestrating a plethora of cellular mechanisms and controlling cellular homeostasis. Transcription factors share distinct DNA binding domains, which allows to group them into protein families. Among them, the Forkhead box O (FOXO) family contains transcription factors crucial for cellular homeostasis, longevity and response to stress. The dysregulation of FOXO signaling is linked to drug resistance in cancer therapy or cellular senescence, however, selective drugs targeting FOXOs are limited, thus knowledge about structure and dynamics of FOXO proteins is essential. Here, we provide an extensive study of structure and dynamics of all FOXO family members. We identify residues accounting for different dynamic and structural features. Furthermore, we show that the auto-inhibition of FOXO proteins by their C-terminal trans-activation domain is conserved throughout the family and that these interactions are not only possible intra-, but also inter-molecularly. This indicates a model in which FOXO transcription factors would modulate their activities by interacting mutually.

Millions of people worldwide are affected by neurodegenerative diseases (NDs), and to date, no effective treatment has been reported. The hallmark of these diseases is the formation of pathological aggregates and fibrils in neural cells. Many studies have reported that catechins, polyphenolic compounds found in a variety of plants, can directly interact with amyloidogenic proteins, prevent the formation of toxic aggregates, and in turn play neuroprotective roles. Besides harboring amyloidogenic domains, several proteins involved in NDs possess arginine-glycine/arginine-glycine-glycine (RG/RGG) regions that contribute to the formation of protein condensates. Here, we aimed to assess whether epigallocatechin gallate (EGCG) can play a role in neuroprotection via direct interaction with such RG/RGG regions. We show that EGCG directly binds to the RG/RGG region of fused in sarcoma (FUS) and that arginine methylation enhances this interaction. Unexpectedly, we found that low micromolar amounts of EGCG were sufficient to restore RNA-dependent condensate formation of methylated FUS, whereas, in the absence of EGCG, no phase separation could be observed. Our data provide new mechanistic roles of EGCG in the regulation of phase separation of RG/RGG-containing proteins, which will promote understanding of the intricate function of EGCG in cells.

Intrinsically disordered regions (IDRs) lack stable tertiary structure and instead rapidly interconvert between different conformations. This structural plasticity enables IDRs to act as key players in cellular signaling pathways. Transcription factors are enriched in IDRs, many of which are stabilized by or acquire tertiary structure in the presence of DNA or other binding partners. Using the T-cell factor/lymphoid enhancer binding factor 1 (TCF/LEF-1) transcription factor as a model system, we characterized the structure and dynamics of the high-mobility group (HMG) domain in the absence of DNA. Inclusion of the IDRs that flank the HMG domain led to enhanced solubility of the protein. Secondary 13Cα chemical shifts, 1H nuclear Overhauser effects, 15N spin relaxation, and 1HN solvent paramagnetic relaxation enhancements indicate that the three helices in the HMG domain are oriented similarly to the DNA-bound form of the protein. By contrast, the flanking IDRs do not show evidence of structure. Helix 1 and helix 3 appear to be less stable in the DNA-free conformation, indicating some form of conformational exchange or local motion in the absence of DNA. Given the high degree of sequence conservation in the TCF/LEF family of transcription factors, our results should apply to other members of the family.