Mutations in presenilin associated with familial Alzheimer disease perturb calcium signaling in cells. In particular, some mutations suppress capacitative calcium entry (CCE), the mechanism for replenishing internal calcium stores from extracellular pools (Yoo et al., 2000 and Liessring et al., 2000). In a paper published this week in PNAS, Tae-Wan Kim and colleagues from Columbia University in New York reveal just how mutant presenilin proteins block CCE, with the identification of a calcium channel that is modulated by the mutated proteins. At the same time, they found that the effects of presenilins (PSs) on calcium flow are mediated by phosphatidylinositol 4,5-bisphosphate (PIP2), a natural activator of the channel. Mutant presenilins lower PIP2 levels, through an unknown, γ-secretase independent pathway, thus inhibiting channel activity.
The researchers, led by first author Natalie Landman, found that the transient potential melastatin 7 (TRPM7)-associated Mg2+-inhibited cation channel (MIC) is activated in PS1/2 knockout mouse embryonic fibroblasts. In contrast, the current through the same channel was suppressed in HEK293 cells stably transfected with either FAD-PS mutant L286V or ΔE9, consistent with the proteins’ known effects on CCE. Inhibition of the channel by FAD-PS mutants did not require γ-secretase activity or the production of Aβ40 or 42, since treatment of the cells with the γ-secretase inhibitors compound E or L-685,458 did not affect the currents. Lowering Aβ42 selectively with NSAIDS likewise had no effect. These results establish that presenilins modulate TRPM7-associated MIC channels independent of γ-secretase activity, and that their ability to affect the channel is altered by FAD-PS mutations.
TRP channels are a large family of non-voltage-dependent cation channels, which maintain calcium and magnesium homeostasis. Deregulation of the TRPM7, which forms the pore of the MIC channel, has been implicated in neurodegeneration due to ischemia and in the mysterious Guamanian neurodegenerative disorder, so it is a good candidate to play a role in Alzheimer disease.
The TRPM7-associated MIC channel is known to be activated by PIP2, and the researchers showed that even in cells expressing mutant PS1, applying PIP2 directly to the cytoplasm of cells in the patch clamp solution could activate the channels to wild-type levels. Pharmacological enhancement of PIP2 levels with edelfosine, an inhibitor of the PIP2-degrading enzyme phospholipase C, also caused recovery of MIC currents in PS1 ΔE9 cells.
These findings suggested that a PIP2 deficit might be the cause of diminished MIC channel activity in PS mutant-expressing cells. To confirm this, the researchers directly measured PIP2 levels. Cells expressing either mutant showed a small (5-10 percent) but consistent reduction in steady-state PIP2 levels compared to control cells. But PIP2 turnover was reduced by 40 percent in cells with the FAD PS1 mutations, and 25 percent in cells with the PS2 N141I FAD mutation. Treatment of cells with edelfosine elevated PIP2 levels in the mutant-expressing cells, while a γ-secretase inhibitor did not.
Though the researchers found that γ-secretase does not seem to be involved in altering PIP2 levels, they did find evidence that PIP2 affects γ-secretase activity. When they measured levels of Aβ in APP-transfected cells treated with activators or inhibitors of phospholipase C, they found that increasing steady-state levels of PIP2 with the PLC inhibitor edelfosine decreased secreted Aβ42, while decreasing PIP2 with an activator of phospholipase C increased Aβ42. Thus, raising PIP2 reverses the FAD phenotype of Aβ42 overexpression, while lowering PIP2 mimics it. The researchers saw the same effect when the readout was cleavage of transfected C99, suggesting that the differences were truly due to alterations in γ-secretase activity.
Together, the results implicate changes in PIP2 levels in two entirely separate outcomes of presenilin mutation, namely the modulation of an ion channel and altered Aβ production. Without knowing how PS might be involved in setting PIP2 levels, it’s anyone’s guess as to whether this phenomenon represents a gain of abnormal function for PS, or a loss of a normal function. (For much more on that philosophical debate, see the recent ARF Forum Discussion). Nonetheless, the finding that increasing PIP2 suppresses multiple FAD mutant phenotypes promises to get this lipid its share of attention as another approach to controlling Aβ production.—Pat McCaffrey
- Yoo AS, Cheng I, Chung S, Grenfell TZ, Lee H, Pack-Chung E, Handler M, Shen J, Xia W, Tesco G, Saunders AJ, Ding K, Frosch MP, Tanzi RE, Kim TW. Presenilin-mediated modulation of capacitative calcium entry. Neuron. 2000 Sep;27(3):561-72. PubMed.
- Leissring MA, Akbari Y, Fanger CM, Cahalan MD, Mattson MP, Laferla FM. Capacitative calcium entry deficits and elevated luminal calcium content in mutant presenilin-1 knockin mice. J Cell Biol. 2000 May 15;149(4):793-8. PubMed.
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- Landman N, Jeong SY, Shin SY, Voronov SV, Serban G, Kang MS, Park MK, Di Paolo G, Chung S, Kim TW. Presenilin mutations linked to familial Alzheimer's disease cause an imbalance in phosphatidylinositol 4,5-bisphosphate metabolism. Proc Natl Acad Sci U S A. 2006 Dec 19;103(51):19524-9. PubMed.