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

Abstract

Functional amyloids are present in a wide variety of organisms ranging from bacteria to humans. Experience-dependent aggregation of the cytoplasmic polyadenylation element-binding (CPEB) prion-like protein to a translationally active state has emerged as a plausible biochemical substrate of long-lasting memories. CPEB aggregation is driven by prion-like domains (PLD) that are highly divergent in sequence across species. Here, we describe the amyloid-like features of the neuronal Aplysia CPEB (ApCPEB) PLD in vitro using single-molecule and bulk biophysical methods and compare them with those previously reported for neuronal Drosophila CPEB, Orb2 PLD. The existence of transient oligomers and mature filaments suggests similarities in the late stages of the assembly pathway for both PLDs. However, while prior to aggregation the Orb2 PLD monomer remains as a random coil in solution, ApCPEB PLD adopts a diversity of conformations comprising α -helical structures that evolve to coiled-coil species, suggesting structural differences at the beginning of their amyloid assembly pathways. Our results show how divergent PLDs of CPEB proteins from different species retain the ability to form a generic amyloid-like fold through different assembly mechanisms.

Structural transitions in Orb2 prion-like domain relevant for functional aggregation in memory consolidation

“…Whereas Aplysia and mammalian CPEB homologs share a similar domain organization with Drosophila Orb2, they lack the multiple His residues interspersed in their Q-rich segments (40,43,44) which are found in Orb2. This suggests that the pHregulated nature of Orb2 amyloid formation and dissociation may be substituted by other mechanisms.…”

Structural transitions in Orb2 prion-like domain relevant for functional aggregation in memory consolidation

“…The resulting helical conformations may drive association processes such as the formation of coiled-coils, as seen for other proteins which undergo liquid-liquid phase separation via intermolecular helical associations (Conicella et al, 2016). Q-rich coiled-coils and higher oligomeric species have been proposed as intermediates in amyloidogenesis of Aplysia and mammalian CPEBs (Fiumara et al, 2010;Pelassa et al, 2014;Hervás et al, 2020b;Ramírez de Mingo, et al, 2020a), which are probably heterogeneous as strongly suggested by kinetic modeling (Vitalis and Pappu, 2011). A hypothetical working model which incorporates these ideas is shown in Figure 5.…”

“…A hypothetical working model which incorporates these ideas is shown in Figure 5. Whereas Aplysia and mammalian CPEB homologs share a similar domain organization with Drosophila Orb2, they lack the multiple His residues interspersed in their Q-rich segments (Hervás et al, 2020b;Ramírez de Mingo, et al, 2020a) which are found in Orb2. This suggests that the pH-regulated nature of Orb2 amyloid formation and dissociation may be substituted by other mechanisms in higher organisms.…”

Molecular Determinants of Liquid Demixing and Amyloidogenesis in Human CPEB3

“…The polyQ binding peptide 1 (QBP1) has been previously shown to block the amyloid formation of several pathological amyloid-forming proteins, such as expanded polyQ tracts 40,41 and a TDP-43 Q/N-rich segment 42 , as well as Q/N-rich functional amyloids, such as Sup35 41 and hCPEB3 homologs from Aplysia (ApCPEB) 16 and Drosophila melanogaster (Orb2) 13 .…”

“…Moreover, the conformational trends within the PLD that precedes amyloid formation appears to be different among species. Thus, while the ApCPEB PLD forms α-helical coiled-coils in vitro and assembles into multimers to enhance amyloid aggregation [14][15][16] , the recombinant Orb2 PLD remains as a random coil conformation, which is able to oligomerize and form amyloid structures whose β-sheet core seems not to include the Q-rich domain 17 . By contrast, recent cryo-EM data revealed that Drosophila Orb2 from adult heads forms a threefold-symmetric hydrophilic amyloid core based on the Q-rich stretch 18 .…”