Cholesterol Modifies Huntingtin Binding to, Disruption of, and Aggregation on Lipid Membranes

Abstract

Huntington’s disease (HD) is an inherited neurodegenerative disease caused by abnormally long CAG-repeats in the huntingtin gene that encode an expanded polyglutamine (polyQ) domain near the N-terminus of the huntingtin (htt) protein. Expanded polyQ domains are directly correlated to disease-related htt aggregation. Htt is found highly associated with a variety of cellular and subcellular membranes that are predominantly comprised of lipids. Since cholesterol homeostasis is altered in HD, we investigated how varying cholesterol content modifies the interactions between htt and lipid membranes. A combination of Langmuir trough monolayer techniques, vesicle permeability and binding assays, and in situ atomic force microscopy were used to directly monitor the interaction of a model, synthetic htt peptide and a full-length htt-exon1 recombinant protein with model membranes comprised of total brain lipid extract (TBLE) and varying amounts of exogenously added cholesterol. As the cholesterol content of the membrane increased, the extent of htt insertion decreased. Vesicles containing extra cholesterol were resistant to htt-induced permeabilization. Morphological and mechanical changes in the bilayer associated with exposure to htt were also drastically altered by the presence of cholesterol. Disrupted regions of pure TBLE bilayers were grainy in appearance and associated with a large number of globular aggregates. In contrast, morphological changes induced by htt in bilayers enriched in cholesterol were plateau-like with a smooth appearance. Collectively, these observations suggest that the presence and amount of cholesterol in lipid membranes play a critical role in htt binding and aggregation on lipid membranes.

Membrane interactions accelerate the self-aggregation of huntingtin exon 1 fragments in a polyglutamine length-dependent manner

“…The data presented is in agreement with previous investigations where GDT-exon1 constructs have been found associated with rat postsynaptic membranes and where brain lipid vesicles accelerate nucleation and thereby fibril formation upon trypsin cleavage of 3 M GST-exon 1 encompassing 51 glutamines (21). In the same study the presence of zwitterionic DMPC or of DOPC/SM/cholesterol slowed down this effect thereby being in-line with studies where htt17 membrane interactions were weak or absent for zwitterionic and cholesterol-rich membranes (39,76).…”

“…Such lipid interactions have been shown to be modulators of aggregation and fibril formation also for α -synuclein (57,62,64), islet amyloid polypeptide (9) and βamyloid (58,84). Within these studies the detailed membrane composition including the resulting physico-chemical properties such as negative charge density, fluidity, saturation, curvature or interactions with specific lipids all play important roles in the aggregation process (9,39,56,76,85). Thereby, the membrane interactions of polyQ flanking regions and their modulation by posttranslational modifications provides a possible therapeutic intervention site which has to our knowledge not been explored in greater detail.…”

Proteins Containing Expanded Polyglutamine Tracts and Neurodegenerative Disease

“…262, 263 Although lipid bilayers can directly influence aggregation, lipid membranes can also be directly disrupted by the expanded polyQ proteins, leading to permeabilization. 179, 264 Htt aggregates can associate directly lipid membranes and induce local mechanical changes. 68, 179, 212 …”

“…198 Htt/lipid interaction is further modulated by membrane composition and polyQ length. 248, 265 For example, exogenous cholesterol protects lipid bilayers from htt induced disruption, 264 which is particularly important provided that HD patients have altered cholesterol homeostasis. 266, 267 Membrane degradation of the nuclear envelope, ER, and mitochondria has been described to the presence of mutant htt, 76, 162 and expanded polyQ can generically distort ER membranes and the nuclear envelope in cell culture.…”

Interaction of Huntingtin Exon-1 Peptides with Lipid-Based Micellar Nanoparticles Probed by Solution NMR and Q-Band Pulsed EPR

“…3 Htt NT is known to target a variety of membrane-containing structures within cells, 4 undergoes a random coil to α -helix transition upon binding lipid membranes, 5 and mediates aggregation of polyQ on membrane surfaces. 3a,6 Here we explore the interaction of two htt NT Q n ( n = 7 and 10) constructs with lipid-based micellar nanoparticles 7 using a combination of NMR chemical exchange saturation transfer (CEST) and pulsed Q-band EPR spectroscopy.…”