Structural basis for reversible amyloids of hnRNPA1 elucidates their role in stress granule assembly

Xinrui Gui,Zhenheng Qin,Dan Li,Jianhua He,Xueming Li,Feng Luo,Heng Zhou,Bo Sun,Lin He,Jinge Gu,Cong Liu,Muyun Xie,Zhenying Liu,Yichen Li,Kun Zhao,Lin Jiang,Woo Shik Shin,Minglei Zhao,Bin Dai

doi:10.1038/s41467-019-09902-7

Abstract

Subcellular membrane-less organelles consist of proteins with low complexity domains. Many of them, such as hnRNPA1, can assemble into both a polydisperse liquid phase and an ordered solid phase of amyloid fibril. The former mirrors biological granule assembly, while the latter is usually associated with neurodegenerative disease. Here, we observe a reversible amyloid formation of hnRNPA1 that synchronizes with liquid–liquid phase separation, regulates the fluidity and mobility of the liquid-like droplets, and facilitates the recruitment of hnRNPA1 into stress granules. We identify the reversible amyloid-forming cores of hnRNPA1 (named hnRACs). The atomic structures of hnRACs reveal a distinct feature of stacking Asp residues, which contributes to fibril reversibility and explains the irreversible pathological fibril formation caused by the Asp mutations identified in familial ALS. Our work characterizes the structural diversity and heterogeneity of reversible amyloid fibrils and illuminates the biological function of reversible amyloid formation in protein phase separation.

Highlights

Subcellular membrane-less organelles consist of proteins with low complexity domains
Missense mutations of low complexity (LC) identified in neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), multisystem proteinopathy (MSP), and frontotemporal dementia (FTD), can promote amyloid aggregation, which is closely associated with the pathogenesis of these diseases[12,13,14]
The results showed a specific enhancement of Thioflavin T (ThT) intensity in the droplets, which consistent with the transmission electron microscopy (TEM) imaging, indicates amyloid fibril formation in the droplets

Summary

Introduction

Subcellular membrane-less organelles consist of proteins with low complexity domains. The atomic structures of hnRACs reveal a distinct feature of stacking Asp residues, which contributes to fibril reversibility and explains the irreversible pathological fibril formation caused by the Asp mutations identified in familial ALS. Our previous work on FUS LC identified two segments that form reversible fibrils regulated by temperature and phosphorylation[24] These observations altered the view of amyloid fibrils as thermostable end-product of protein super-molecular assembly, and raised the possibility that amyloid formation may play a functional role in the assembly of membraneless organelles. The structure reveals a distinct feature of negatively charged Asp residues stacking along the fibril, which contributes to fibril instability Based on this structure, we identify another two reversible amyloid cores from hnRNPA1 LC (named hnRAC2 and hnRAC3) and more from hnRNP families. Our work bridges the liquid phase and solid phase of hnRNPA1 and illuminates the functional role of reversible amyloids in the assembly of stress granules and the pathological risk as intermediates en route to irreversible fibrils

Methods

Results

Conclusion