Tabuchi K, Chen G, Südhof TC, Shen J.
Conditional forebrain inactivation of nicastrin causes progressive memory impairment and age-related neurodegeneration.
J Neurosci. 2009 Jun 3;29(22):7290-301.
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The paper by Tabuchi at al. is a fascinating study that uses genetic approach to investigate the importance of γ-secretase activity for neuronal survival. The authors generated floxed nicastrin mice and crossed them with the α-CaMKII-Cre transgenic driver line that expresses Cre recombinase specifically in the excitatory neurons of postnatal forebrain. The resulting nicastrin cKO mouse line appears to be essentially normal at two months of age. By the age of six and nine months, nicastrin cKO mice developed age-dependent memory deficits, displaying apoptotic loss of cortical neurons and active gliosis. The phenotype of aging nicastrin cKO mice is very similar to the phenotype of presenilin cDKO mice that Jie Shen’s group previously described (1). The authors conclude that 1) γ-secretase activity is essential for neuronal survival; and that 2) γ-secretase independent functions of presenilins, such as ER Ca2+ leak function (2), are not important for neuronal survival in this context.
The experimental design is very elegant and leaves no doubt that γ-secretase function is essential for neuronal survival. Although the exact mechanism that connects γ-secretase activity with neuronal survival is not established, these findings have obvious and important implications for developing and clinically testing γ-secretase inhibitors. The conclusion about the non-essential role of ER Ca2+ leak function of presenilins is somewhat less strong. The authors assume that ER Ca2+ signaling is normal in adult neurons from nicastrin cKO mice, but did not directly test it in the paper. In the previous studies our laboratory demonstrated that ER Ca2+ signals are normal in Aph TKO MEF cells (2), so it is already known that inactivation of γ-secretase per se does not have a major effect on ER Ca2+ handling. However, it is possible that levels of presenilin expression in aging neurons from nicastrin cKO mice are sufficiently reduced to affect ER Ca2+ signaling. When cortical lysates from two-month-old mice were analyzed by Western blotting, levels of presenilin-1 were reduced approximately twofold in nicastrin cKO mice. Unfortunately, the expression levels of presenilins are not shown for six- and nine-month cortical lysates from nicastrin cKO mice.
Another critical issue to consider is whether memory deficits and cortical neuronal loss observed in PS cDKO and nicastrin cKO mice faithfully replicate neuronal dysfunction and eventual death in AD. There is no doubt that previous (1) and present (Tabuchi at al., 2009) data from Jie Shen’s lab indicate that γ-secretase activity is important for neuronal survival. However, it remains unclear if the pathway leading to neuronal cell death in γ-secretase KO models is the same as in AD. This is a critical question that still needs to be addressed. Accumulation of amyloid has been postulated to play a critical role in AD pathogenesis (3). Abnormal neuronal Ca2+ signaling has also been implicated as one of the pathogenic pathways involved in AD (4). PS cDKO and nicastrin cKO mice do not produce amyloid and nicastrin cKO mice presumably have normal Ca2+ signaling. Thus, an open question is if neuronal cell death observed in these γ-secretase knockout mice is a faithful model for neuronal cell death in AD. In any case, the new results obtained by the authors provide very interesting and important insights into a connection between γ-secretase activity and neuronal survival. Understanding the mechanistic basis responsible for this connection will be an extremely important future task.
Saura CA, Choi SY, Beglopoulos V, Malkani S, Zhang D, Shankaranarayana Rao BS, Chattarji S, Kelleher RJ 3rd, Kandel ER, Duff K, Kirkwood A, Shen J.
Loss of presenilin function causes impairments of memory and synaptic plasticity followed by age-dependent neurodegeneration.
Neuron. 2004 Apr 8;42(1):23-36.
Tu H, Nelson O, Bezprozvanny A, Wang Z, Lee SF, Hao YH, Serneels L, De Strooper B, Yu G, Bezprozvanny I.
Presenilins form ER Ca2+ leak channels, a function disrupted by familial Alzheimer's disease-linked mutations.
Cell. 2006 Sep 8;126(5):981-93.
Hardy J, Selkoe DJ.
The amyloid hypothesis of Alzheimer's disease: progress and problems on the road to therapeutics.
Science. 2002 Jul 19;297(5580):353-6.
Bezprozvanny I, Mattson MP.
Neuronal calcium mishandling and the pathogenesis of Alzheimer's disease.
Trends Neurosci. 2008 Sep;31(9):454-63.
In this paper, Tabuchi et al. report a novel mouse that specifically lacks the nicastrin gene in excitatory neurons. Nicastrin cKO mice showed memory impairment and age-dependent cortical neuron loss, similar phenotypes to the PS cDKO mice. Of note, the nicastrin cKO mice showed significant memory impairment at two months of age. At this stage, the gross brain morphology was normal, suggesting that “functional” defects would have already occurred by the deletion of nicastrin gene, presumably the complete loss of γ-secretase activity in neurons. These data implicate that the neuronal γ-substrate is functionally important in learning and memory without significant synaptic loss or neuron death. Thus, unveiling the molecular mechanism whereby the cKO neurons showed functional defects is an important issue to consider the physiological role of the γ-secretase activity in the brain.
I think this is a very elegant study that reveals the fundamental role of nicastrin in neuronal integrity and memory in adult mice. This work also confirms in vivo, in the adult brain, that nicastrin is essential for the stabilization and activity of the γ-secretase complex. These results are nicely in line with the notion that complete or partial loss of function of presenilins is, per se, neurotoxic.
The challenge will be now to determine what are the signaling pathways—downstream from γ-secretase—involved in neuronal death in the aging brain. Cadherins may represent attractive candidates. Indeed, the cadherin family of cell-cell adhesion proteins is abundantly expressed at mature synapses, is critical for synaptic plasticity, and is cleaved by γ-secretase in neurons upon NMDA receptor stimulation (Marambaud et al., 2003). It is, therefore, reasonable to think that a loss of synaptic cadherin cleavage by γ-secretase may lead over time to defects in synaptic plasticity and neuronal integrity and thus may contribute to the phenotype observed in these mice.
Another important question relates to the integrity in these mice of the CREB/CBP transcriptional pathway, which appeared to be significantly compromised in the PS cKO mice (Saura et al., 2004).
Marambaud P, Wen PH, Dutt A, Shioi J, Takashima A, Siman R, Robakis NK.
A CBP binding transcriptional repressor produced by the PS1/epsilon-cleavage of N-cadherin is inhibited by PS1 FAD mutations.
Cell. 2003 Sep 5;114(5):635-45.
This is a very elegant knockout study reinforcing previous work of Jie Shen published in Neuron, which showed that presenilin 1 and 2 double deficient mice display a progressive neurodegenerative disorder.
In contrast with their previous paper in Neuron, Shen and colleagues now conclude that the neurodegeneration they see in both PS1 and 2 double KO mice, and nicastrin single KO mice is due to a γ-secretase defect, i.e., the loss of proteolytic function. This is part of an ongoing debate as to what extent postulated functions of presenilin outside the γ-secretase complex contribute to the overall phenotype of presenilin deficient mice. While I tend to believe that the neurodegeneration observed in their studies is indeed reflecting a real γ-secretase defect, the current paper is not conclusive in that regard. Indeed, knockout of nicastrin also destabilizes presenilin, and could theoretically affect functions of presenilin independent of its proteolytic function. However, I agree with Shen and colleagues that their current interpretation is the most likely one, as the major defect in nicastrin deficient cells is the loss of proteolytic activity, while some presenilin level is still maintained, which could fulfill these postulated other functions. For instance the Ca2+ leakage function is maintained in APh1 deficient cells (Tu et al., 2006) indicating that presenilin can exert that function outside of the complex.
The big question, as the authors discuss in their manuscript, is the identification of the substrate that is responsible for the neurodegenerative phenotype. This is a very difficult question to answer, given the many different substrates, and the possibility that any of the γ-secretase substrates which accumulate in the Nct knockout mice could theoretically contribute to this phenotype.
It is clear that γ-secretase as a drug target is not an easy one. We published recently that it is possible to knock out specifically the APh1B-γ-secretase in the brain of mice without neurodegenerative changes, and with the potential to clear Aβ peptide from the brain (Serneels et al., 2009). This indicates that a partial inhibition of the complex in brain is feasible and with acceptable side effects.
Serneels L, Van Biervliet J, Craessaerts K, Dejaegere T, Horré K, Van Houtvin T, Esselmann H, Paul S, Schäfer MK, Berezovska O, Hyman BT, Sprangers B, Sciot R, Moons L, Jucker M, Yang Z, May PC, Karran E, Wiltfang J, D'Hooge R, De Strooper B.
gamma-Secretase heterogeneity in the Aph1 subunit: relevance for Alzheimer's disease.
Science. 2009 May 1;324(5927):639-42. Epub 2009 Mar 19