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Conformational targeting of intracellular A? oligomers demonstrates their pathological oligomerization inside the endoplasmic reticulum.

ABSTRACT: A? oligomers (A?Os) are crucially involved in Alzheimer's Disease (AD). However, the lack of selective approaches for targeting these polymorphic A? assemblies represents a major hurdle in understanding their biosynthesis, traffic and actions in living cells. Here, we established a subcellularly localized conformational-selective interference (CSI) approach, based on the expression of a recombinant antibody fragment against A?Os in the endoplasmic reticulum (ER). By CSI, we can control extra- and intracellular pools of A?Os produced in an AD-relevant cell model, without interfering with the maturation and processing of the A? precursor protein. The anti-A?Os intrabody selectively intercepts critical A?O conformers in the ER, modulating their assembly and controlling their actions in pathways of cellular homeostasis and synaptic signalling. Our results demonstrate that intracellular A? undergoes pathological oligomerization through critical conformations formed inside the ER. This establishes intracellular A?Os as key targets for AD treatment and presents CSI as a potential targeting strategy.

SUBMITTER: Meli G

PROVIDER: S-EPMC4050278 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature

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Conformational targeting of intracellular Aβ oligomers demonstrates their pathological oligomerization inside the endoplasmic reticulum.

Meli Giovanni G Lecci Agnese A Manca Annalisa A Krako Nina N Albertini Valentina V Benussi Luisa L Ghidoni Roberta R Cattaneo Antonino A

Nature communications 20140527

Aβ oligomers (AβOs) are crucially involved in Alzheimer's Disease (AD). However, the lack of selective approaches for targeting these polymorphic Aβ assemblies represents a major hurdle in understanding their biosynthesis, traffic and actions in living cells. Here, we established a subcellularly localized conformational-selective interference (CSI) approach, based on the expression of a recombinant antibody fragment against AβOs in the endoplasmic reticulum (ER). By CSI, we can control extra- an ...[more]

PMID: 24861166

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Project description:Photodynamic therapy (PDT) is often used in preclinical and clinical treatment regimens. Reactive oxygen species (ROS) generated by photosensitizers (PSs) upon exposure to light induce cell death via diverse mechanisms. PSs can exert therapeutic effects in different cellular organelles, although the efficacy of organelle-specific PDT has yet to be determined as most previous studies use different PSs in different organelles. Here, we explored how a single PS, chlorin e6 (Ce6), targeted to different organelles altered the effectiveness of PDT. Ce6 intrinsically localizes to the ER after 4 h of incubation. Modification of Ce6 via conjugation with an octapeptide (LS765), a monosubstituted triphenylphosphonium (TPP) derivative (LS897), or a disubstituted TPP derivative (LS909) altered the intrinsic localization. We determined that LS765 and LS9897 predominantly accumulated in the lysosomes, but LS909 trafficked equally to both the mitochondria and the lysosomes. Moreover, the conjugation altered the type of ROS produced by Ce6, increasing the ratio of hydrogen peroxide to hydroxyl radicals. Irradiation of identical concentrations of the PSs in solution with 650 nm, 0.84 mW/cm2 light for 10 min showed that the TPP conjugates nearly doubled the hydrogen peroxide production from ∼0.2 μM for Ce6 and LS765 to ∼0.37 μM for LS897 and LS909. In contrast, Ce6 produced ∼1.5-fold higher hydroxyl radicals than its conjugates. To compare the effect of each PS on cell death, we normalized the intracellular concentration of each PS. Hydrogen peroxide-producing PSs are effective PDT agents in the lysosomes while the hydroxyl-generating PSs are very effective in the ER. Compared to the PSs that accumulated in the lysosomes, only the ER-targeted Ce6 exerted >50% cell death at either low light power or low intracellular concentration. By delineating the contributions of cellular organelles and types of ROS produced, our work suggests that targeting hydroxyl radical-producing PSs to the ER is an exciting strategy to improve the therapeutic outcome of PDT.

Dataset Information

Conformational targeting of intracellular A? oligomers demonstrates their pathological oligomerization inside the endoplasmic reticulum.

Publications

Conformational targeting of intracellular Aβ oligomers demonstrates their pathological oligomerization inside the endoplasmic reticulum.

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