Divergent CPEB prion-like domains reveal different assembly mechanisms for a generic amyloid-like fold

Rubén Hervás,Margarita Menéndez,Yoshitaka Nagai,María Del Carmen Fernández-Ramírez,Mariano Carrión-Vázquez,Albert Galera-Prat,Douglas V Laurents,Mari Suzuki,Marta Bruix

doi:10.1186/s12915-021-00967-9

Abstract

BackgroundAmyloids are ordered, insoluble protein aggregates, characterized by a cross-β sheet quaternary structure in which molecules in a β-strand conformation are stacked along the filament axis via intermolecular interactions. While amyloids are typically associated with pathological conditions, functional amyloids have also been identified and are present in a wide variety of organisms ranging from bacteria to humans. The cytoplasmic polyadenylation element-binding (CPEB) prion-like protein is an mRNA-binding translation regulator, whose neuronal isoforms undergo activity-dependent aggregation, a process that has emerged as a plausible biochemical substrate for memory maintenance. CPEB aggregation is driven by prion-like domains (PLD) that are divergent in sequence across species, and it remains unknown whether such divergent PLDs follow a similar aggregating assembly pathway. Here, we describe the amyloid-like features of the neuronal Aplysia CPEB (ApCPEB) PLD and compare them to those of the Drosophila ortholog, Orb2 PLD.ResultsUsing in vitro single-molecule and bulk biophysical methods, we find transient oligomers and mature amyloid-like filaments that suggest similarities in the late stages of the assembly pathway for both ApCPEB and Orb2 PLDs. However, while prior to aggregation the Orb2 PLD monomer remains mainly as a random coil in solution, ApCPEB PLD adopts a diversity of conformations comprising α-helical structures that evolve to coiled-coil species, indicating structural differences at the beginning of their amyloid assembly pathways.ConclusionOur results indicate that divergent PLDs of CPEB proteins from different species retain the ability to form a generic amyloid-like fold through different assembly mechanisms.

Highlights

Amyloids are ordered, insoluble protein aggregates, characterized by a cross-β sheet quaternary structure in which molecules in a β-strand conformation are stacked along the filament axis via intermolecular interactions
In agreement with Coils, Aplysia cytoplasmic polyadenylation element-binding (CPEB) (ApCPEB) prion-like domains (PLD) was predicted to contain Coiled coil (CC) domains, which overlapped with the poly-Q stretches, by using three additional algorithms that detect CC motifs in protein sequence: PairCoil2 [25], Marcoil [26], or PCOILS [27] (Additional file 1: Figure S2)
ApCPEB PLD is structured in solution and its early aggregation is mediated by the formation of CCs, as previously observed for other Q/N-rich proteins [22]

Summary

Introduction

Insoluble protein aggregates, characterized by a cross-β sheet quaternary structure in which molecules in a β-strand conformation are stacked along the filament axis via intermolecular interactions. The neuronal-specific ApCPEB isoform can exist in at least two different conformational states: a soluble form and a β-sheet-rich amyloid-like form, which has an enhanced binding capacity to target mRNAs [20, 21]. In both ApCPEB and Orb proteins, in spite of their low sequence identity (Fig. 1a, b, Additional file 1: Figure S1c), the N-terminal PLD plays a key role in the transition to a β-sheet-rich fold through a neuronal activitydependent aggregation process [10, 17, 19, 20]. The existence of alternative amyloid assembly pathways, as here described, could attend to the potential, diverse regulatory mechanisms in each organism

Methods

Results

Conclusion