The Role of G Protein Activation in the Toxicity of Amyloidogenic Aβ-(1–40), Aβ-(25–35), and Bovine Calcitonin

Abstract

More than 16 different proteins have been identified as amyloid in clinical diseases; among these, ␤-amyloid (A␤) of Alzheimer's disease is the best characterized. In the present study, we performed experiments with A␤ and calcitonin, another amyloid-forming peptide, to examine the role of G protein activation in amyloid toxicity. We demonstrated that the peptides, when prepared under conditions that promoted ␤-sheet and amyloid fibril (or protofibril) formation, increased high affinity GTPase activity, but the nonamyloidogenic peptides had no discernible effects on GTP hydrolysis. These increases in GTPase activity were correlated to toxicity. In addition, G protein inhibitors significantly reduced the toxic effects of the amyloidogenic A␤ and calcitonin peptides. Our results further indicated that the amyloidogenic peptides significantly increased GTPase activity of purified G␣ o and G␣ i subunits and that the effect was not receptor-mediated. Collectively, these results imply that the amyloidogenic structure, regardless of the actual peptide or protein sequence, may be sufficient to cause toxicity and that toxicity is mediated, at least partially, through G protein activation. Our abilities to manipulate G protein activity may lead to novel treatments for Alzheimer's disease and the other amyloidoses.

Reduction in Cholesterol and Sialic Acid Content Protects Cells from the Toxic Effects of β-Amyloid Peptides

“…The final diluted concentration of calcitonin in all studies was 80 M. Prepared under these conditions, bovine calcitonin in divalent salt solution has a 55% ␤-sheet structure and binds Congo Red, typical of amyloids, whereas bovine calcitonin in deionized water has a 95% ␣-helix structure and does not bind Congo Red at detectable levels (32).…”

“…Materials-A␤- , A␤- (25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35), and A␤-(1-16) were purchased from BIOSOURCE International (Camarillo, CA). ATP and GTP were purchased from Aldrich and [␥-32 P]GTP from ICN Biochemicals (Irvine, CA).…”

“…Peptide Preparation-A␤- and A␤-(25-35) peptides were prepared analogously to methods which in our hands consistently lead to peptides that are toxic to cultured cells (32). 2 Stock solutions of 10 mg/ml were prepared by dissolving the A␤ peptides in 0.1% (v/v) trifluoroacetic acid in water.…”

“…In the present study, we examined the effects of depleting cholesterol and cell surface sialic acid residues associated with gangliosides and membrane-associated glycoproteins on the membrane-associated GTP hydrolysis induced by two A␤ fragments, A␤-(1-40) and A␤- (25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35). We had shown previously that G protein activation is an early step in A␤-induced neurotoxicity (32), and we now show that depletion of either membrane component significantly attenuated the A␤-induced increases in GTP hydrolysis observed in PC12 cells.…”

Differential prevention of morphine amnesia by antisense oligodeoxynucleotides directed against various Gi‐protein α subunits

“…Furthermore, lower Gsa levels and unmodi®ed Gia levels were observed in ®broblasts from familial Alzheimer's disease patients (Shanahan et al, 1997). Recently, it has been reported that amyloid b-peptides cause toxicity through activation of Gi proteins (Soomets et al, 1999;Rymer & Good, 2001). We can hypothesize that an intact Gi-protein functionality is essential for the induction of amnesia.…”