Human apolipoprotein A-I natural variants: molecular mechanisms underlying amyloidogenic propensity

RAMELLA, N.; SCHINELLA, G; TRICERRI, M. A.; GADDI, G. M.; PRIETO, E. D.; ROSU, S. A.; FINARELLI, G. S.

Uppsala

Simposio; XV International Symposium on Amyloidosis; 2016

International Society on Amyloidosis

INTRODUCTION:Naturally occurring mutations of Human apolipoprotein A-I (apoA-I) have been shown to induce amyloidosis in patients, with a broad range of clinical manifestations, depending on the protein variant which is involved (1). Although the molecular mechanisms of apoA-I amyloid associated pathology remain largely unknown, the fact that the wild-type (Wt) deposits in atherosclerotic plaques supports the hypothesis that a chronic inflammatory micro environment could elicit protein aggregation (2). In order to get insight into the mechanisms inducing apoA-I misfolding, we examined the effects of point mutations in apoA-I on the structure, stability, and aggregation propensity, as well as on the ability to bind to putative ligands. MATERIAL & METHODS:Proteins were incubated under different conditions (low pH, in the presence of ligands, proteolysis enzimes etc) and structural features associated to protein folding were analyzed by fluorescence spectroscopy and western blotting. Aggregates were characterized by Atomic Force Microscopy. Pro-inflammatory Cell activation was checked by using standard macrophage cell cultures (3)RESULTS:Our results indicate that all the mutants tested (Gly26Arg, Lys107-0, Arg173Pro and Leu60Arg) are less stable than the Wt. Arg173Pro shows a higher susceptibility to partial proteolysis, and a lower efficiency to bind to phospholipid vesicles at physiological pH, which could determine the observed higher tendency to aggregate as pro-amyloidogenic complexes. But in addition, this mutant binds more efficiently to heparin or other ligands, even at physiological pH, which could explain its retention in the pro-inflammatory landscape. Lys107-0 shows higher tendency to aggregate than Gly26Arg, but, interestingly Gly26Arg and Arg173Pro (but not Lys107-0) are able to elicit macrophage activation, thus stimulating local chronic inflammation.DISCUSSION & CONCLUSIONS Our results suggest that apoA-I mutants share some but not all the same mechanism of pathogenicity. More than stability, binding to ligands as heparin or other glycosaminoglycans could be key events tuning the fine details of the interaction of apoA-I variants with the micro-environment. Moreover, induction of macrophage activation could be an attractive hypothesis to explain why certain variants contribute more than others to apoA-I-induced pathogenesis. Further studies should focus on the complex landscape mediating the roles of apoA-I in the delicate balance between health and pathology.REFERENCES(1)Eriksson M, Schonland S, Yumlu S, et al. Hereditary apolipoprotein AI-associated amyloidosis in surgical pathology specimens: identification of three novel mutations in the APOA1 gene. J Mol Diagn 2009: 11: 257?262. (2) Mucchiano GI, Haggqvist B, et al Sletten K. Apolipoprotein A-1-derived amyloid in atherosclerotic plaques of the human aorta. J Pathol 2001; 193: 270?275. (3) Ramella NA, Rimoldi OJ, Prieto ED et al. Human Apolipoprotein A-I-Derived Amyloid: Its Association with Atherosclerosis. PLoS ONE 2011; 6(7): e22532. Authors acknowledge support from UNLP (M187) and CONICET (PIP 112 201101-00648)