Inactivation of brain Cofilin-1 by age, Alzheimer's disease and gamma-secretase

Rapid remodeling of the actin cytoskeleton in the pre- and/or post-synaptic compartments is responsible for the regulation of neuronal plasticity, which is an important process for learning and memory. Cofilin1 plays an essential role in these processes and a dysregulation of its activity was associated with the cognitive decline observed during normal aging and Alzheimer's disease (AD). To understand the mechanism(s) regulating Cofilin1 activity we evaluated changes occurring with regard to Cofilin1 and its up-stream regulators Lim kinase-1 (LIMK1) and Slingshot phosphatase-1 (SSH1) in (i) human AD brain, (ii) 1-, 4-, and 10-months old APP/PS1 mice, (iii) wild type 3-, 8-, 12-, 18- and 26-months old mice, as well as in cellular models including (iv) mouse primary cortical neurons (PCNs, cultured for 5, 10,15 and 20 days in vitro) and (v) mouse embryonic fibroblasts (MEF). Interestingly, we found an increased Cofilin1 phosphorylation/inactivation with age and AD pathology, both in vivo and in vitro. These changes were associated with a major inactivation of SSH1. Interestingly, inhibition of gamma-secretase activity with Compound-E (10 mu M) prevented Cofilin1 phosphorylation/inactivation through an increase of SSH1 activity in PCNs. Similarly, MEF cells double knock-out for gamma-secretase catalytic subunits presenilin-1 and -2 (MEFDKO) showed a strong decrease of both Cofilin1 and SSH1 phosphorylation, which were rescued by the overexpression of human gamma-secretase. Together, these results shed new light in understanding the molecular mechanisms promoting Cofilin1 dysregulation, both during aging and AD. They further have the potential to impact the development of therapies to safely treat AD. (C) 2014 Elsevier B.V. All rights reserved.

Endogenous Regulators of Gamma-Secretase and Amyloid-Beta Production, and Engineering of Alzheimer's Disease Therapeutic Tools

Sébastien Mosser

Alzheimer s disease (AD) is a devastating neurodegenerative disease characterized by strong cognitive impairment and memory loss. These symptoms are caused by neuronal death, induced by two pathological hallmarks: extracellular senile plaques composed of aggregated amyloid-beta (Abeta) peptides, and intracellular neurofibrillary tangles generated by the hyperphosphorylation of the actin-binding protein Tau. Genetic and biochemical analysis resulted in the generation of the so-called amyloid cascade hypothesis, which stipulates that Abeta aggregation precedes neurofibrillary tangles and initiate AD pathogenesis. Abeta originates from the proteolytic cleavage of the amyloid precursor protein (APP) by gamma-secretase, an intramembrane cleaving protease. Thus, pharmacological modulation of gamma-secretase emerged as a promising strategy for the treatment of AD. However, strong side effects, including cognitive function worsening, were reported following clinical inhibition of this enzyme, caused by altered processing of the numerous gamma-secretase substrates. Pharmaceutical industries, alongside with academia, oriented their research toward the discovery of compounds selectively affecting the processing of APP, without altering the cleavage of other substrates. In the first part of this thesis work, we focused our research on endogenous modulation of gamma-secretase, and its consequences on actin cytoskeleton dynamics. We performed mass spectrometric analysis of purified gamma-secretase, and identified the adipocyte plasma membrane associated protein APMAP as a gamma-secretase interacting protein. We showed that APMAP modulates the generation of Abeta peptides without affecting the processing of other gamma-secretase substrates, and controls APP degradation by the lysosomal/autophagic pathway. We also investigated the regulation of gamma-secretase by post translational modifications, and concluded that its enzymatic activity is not affected by its phosphorylation. We then observed the effects of gamma-secretase inhibition on physiological processes, and discovered a gamma-secretase-dependent regulation of the binding of Cofilin to actin filaments. These observations showed a direct link between gamma-secretase and the actin cytoskeleton required for cell motility and neuronal plasticity. In the second part of this work, we aimed at finding a new method for monitoring brain structure alteration and provide new tools for the study of neurodegenerative disease progression in animal models. We performed ex-vivo electrical impedance measurement from intracerebral electrode in mice brain, and succeeded in the visualization of cortical layers as well as brain nuclei. Moreover, we used this method to visualize structural alteration caused by Abeta plaques in an AD mouse model. After imaging, we aimed at repairing neuronal loss causing the structural alterations observed in neurodegenerative diseases. We developed a compressible and injectable extracellular-matrix-like cell culture scaffold (cryogel) to promote neuronal growth in brain lesions. Primary cortical neuron culture on our cryogel scaffold showed good axonal sprouting, not affected by cryogel compression.

EPFL2016

Endogenous Regulators of Gamma-Secretase and Amyloid-Beta Production, and Engineering of Alzheimer's Disease Therapeutic Tools

Sébastien Mosser

EPFL2016