Electron tomography reveals the fibril structure and lipid interactions in amyloid deposits

Abstract

Electron tomography is an increasingly powerful method to study the detailed architecture of macromolecular complexes or cellular structures. Applied to amyloid deposits formed in a cell culture model of systemic amyloid A amyloidosis, we could determine the structural morphology of the fibrils directly in the deposit. The deposited fibrils are arranged in different networks, and depending on the relative fibril orientation, we can distinguish between fibril meshworks, fibril bundles, and amyloid stars. These networks are frequently infiltrated by vesicular lipid inclusions that may originate from the death of the amyloid-forming cells. Our data support the role of nonfibril components for constructing fibril deposits and provide structural views of different types of lipid-fibril interactions.aggregation | conformational disease | electron tomography | protein assembly | prion

Serum amyloid A forms stable oligomers that disrupt vesicles at lysosomal pH and contribute to the pathogenesis of reactive amyloidosis

“…Recombinant murine SAA1 (mSAA1, 103 aa, 11.6 kDa) was expressed in Escherichia coli and purified to 95% purity as described previously (9). All other materials, which were commercially obtained as described in SI Materials and Methods, were of highest available purity.…”

“…26 and references therein). In vitro fibril formation by SAA and AA requires acidic conditions (27) that are optimal circa pH 3 (9). Moreover, ex vivo and cell-based immunohistochemical and electron microscopic (EM) studies have clearly established lysosomes as the initial sites of AA fibril formation in macrophages and other cells (26,28,29).…”

“…Furthermore, compact soluble polymorphic low-order oligomers of various amyloidogenic proteins, including SAA (21)(22)(23)(24), are thought to comprise the pathogenic species that can disrupt cell membranes and/or mediate prion-like transmission (7,8,(13)(14)(15)40). Notably, studies of soluble low-order oligomeric forms of Aβ have revealed a direct correlation between the ANS binding and cytotoxicity (9), whereas studies of a model amyloidogenic protein HypF reported that "structural flexibility and hydrophobic exposure are primary determinants of the ability of oligomeric assemblies to cause cellular dysfunction" (13). We hypothesize that small soluble oligomers of SAA, such as those formed circa pH 4.3, which bind ANS, disrupt phospholipid bilayers, and convert from an α-helix to β-sheet on interacting with these bilayers, may be involved in the pathogenesis of AA amyloidosis.…”

“…Although binding to the lipid surface mediates normal functions of SAA in lipid transport, SAA interactions with cell membranes may also modulate pathologic effects in AA amyloidosis (7,8). Molecular underpinnings for these effects are beginning to be elucidated (9) and are explored in the current study.…”

Serum amyloid A self-assembles with phospholipids to form stable protein-rich nanoparticles with a distinct structure: A hypothetical function of SAA as a “molecular mop” in immune response

“…Our ultimate goal is to understand the delicate balance between the normal functions of SAA in innate immunity and lipid homeostasis and the pathologic pathway of SAA misfolding and deposition in AA amyloidosis. Both normal and pathologic effects of SAA are critically hinged upon its binding to numerous ligands, particularly lipids (Frame and Gursky, 2016; Kollmer et al, 2016; Tanaka et al, 2017) that are in the focus of the current study.…”