Research

Research Focus

Mechanisms of protein quality control in health and disease.

Research Directions

Our current research directions include:

Non-canonical protein quality control systems in protein folding and cellular homeostasis
Regulatory roles and molecular mechanisms of protein quality control in membraneless organelles and cellular homeostasis
Protein quality control in the prevention and treatment of cancer and neurodegenerative diseases

Protein Quality Control m6A / YTHDC1 ALS

YTHDC1 functions as a molecular chaperone to suppress ALS-linked hnRNPA1 mutants from aggregation

Proteostasis failure drives multiple neurodegenerative disorders (NDs), and ATP-independent chaperone pathways that support neuronal proteostasis remain poorly defined. Here, we identify the N6-methyladenosine (m6A)-binding protein YTHDC1 as an ATP-independent molecular chaperone, whose activity is mediated by a highly acidic polyaspartate/glutamate (polyD/E) segment. YTHDC1 prevents protein misfolding and aggregation, unfolds kinetically trapped substrates, and resolubilizes pre-formed aggregates. Deletion of the polyD/E segment abolishes these activities, whereas aromatic-cage mutants retain chaperone activity, demonstrating independence from m6A recognition. We identify the amyotrophic lateral sclerosis (ALS)-associated RNA-binding protein hnRNPA1 as a YTHDC1 client. YTHDC1 maintains liquid-like hnRNPA1 condensates, delays fibrillization of disease-associated mutants, and limits stress-granule sequestration, while mitigating mutant hnRNPA1-induced neurite growth defects in primary neurons. These findings define a proteostatic function of YTHDC1 and highlight its chaperone activity as a potential target for mitigating protein aggregation in ALS-related NDs.

Chaperone activity assays of YTHDC1 and its polyD/E deletion mutant YTHDC1ΔD/E — **Fig. 1 |** a Sedimentation assay of Luc-DM in HEK293T cells expressing EV, YTHDC1, or YTHDC1ΔD/E. b Luc-DM activity in HEK293T cells expressing the indicated constructs. c Representative BiFC images of α-synuclein oligomerization in HEK293T cells expressing EV, YTHDC1, or YTHDC1ΔD/E. Scale bar, 100 µm. d Quantification of BiFC intensity (EV: n = 446, YTHDC1: n = 341, YTHDC1ΔD/E: n = 480 cells). e Activity of heat-denatured luciferase incubated with MBP, YTHDC1, or YTHDC1ΔD/E. f Turbidity of heat-denatured luciferase monitored by OD600. g Reactivation of heat-denatured luciferase by the indicated proteins. h ThT fluorescence of monomeric misfolded LucD incubated with YTHDC1 or YTHDC1ΔD/E. i Time-dependent trypsin sensitivity of LucD after incubation with the indicated proteins. j Thermal shift analysis of native luciferase incubated with YTHDC1 or YTHDC1ΔD/E at the indicated molar ratios. k Thermal shift analysis of YTHDC1 and YTHDC1ΔD/E. l Schematic of freeze-thaw treatment and sedimentation to separate SN and PE fractions. m Sedimentation analysis of YTHDC1 and YTHDC1ΔD/E following freeze-thaw treatment. n Time-lapse images of RNA-induced phase separation of YTHDC1 and YTHDC1ΔD/E. Scale bar, 20 µm. **o, p** Quantification of droplet number (o) and size (p). **q, r** In vitro fluorescence recovery after photobleaching (FRAP) assays. Representative images (q) and quantification of fluorescence recovery (r). White dashed lines indicate bleached regions. Scale bar, 2 µm. **s, t** FRAP assays in HeLa cells. Representative images (s) and quantification of fluorescence recovery (t). Red dashed lines outline nuclei and white dashed lines indicate bleached regions. Scale bar, 10 µm. Data are mean ± SEM (e, g) or mean ± SD (b, d, o, p) from n = 3 independent experiments. Statistical analyses: one-way ANOVA with Tukey’s post hoc test (b, d), two-tailed unpaired Student’s t-test (o, p), extra sum-of-squares F test (e, g), and ANCOVA (f, h). Experiments in a and m were independently repeated three times with similar results. Source data are provided with this paper.

Protein Quality Control Molecular Chaperones Neurodegenerative Disease

DAXX represents a new type of protein-folding enabler

Protein quality control systems are crucial for cellular function and organismal health. At present, most known protein quality control systems are multicomponent machineries that operate via ATP-regulated interactions with non-native proteins to prevent aggregation and promote folding, and few systems that can broadly enable protein folding by a different mechanism have been identified. Moreover, proteins that contain the extensively charged poly-Asp/Glu (polyD/E) region are common in eukaryotic proteomes, but their biochemical activities remain undefined. Here we show that DAXX, a polyD/E protein that has been implicated in diverse cellular processes, possesses several protein-folding activities. DAXX prevents aggregation, solubilizes pre-existing aggregates and unfolds misfolded species of model substrates and neurodegeneration-associated proteins. Notably, DAXX effectively prevents and reverses aggregation of its in vivo-validated client proteins, the tumour suppressor p53 and its principal antagonist MDM2. DAXX can also restore native conformation and function to tumour-associated, aggregation-prone p53 mutants, reducing their oncogenic properties. These DAXX activities are ATP-independent and instead rely on the polyD/E region. Other polyD/E proteins, including ANP32A and SET, can also function as stand-alone, ATP-independent molecular chaperones, disaggregases and unfoldases. Thus, polyD/E proteins probably constitute a multifunctional protein quality control system that operates via a distinctive mechanism.

Overview of the polyD/E protein quality control system — **Fig. 2 |** PolyD/E proteins constitute a multifunctional, ATP-independent protein quality control system (Huang et al., Nature, 2021).