The BACE protease (β-site APP cleaving enzyme) is elevated in the brains of people with Alzheimer disease and takes at least part of the blame for amyloid overload because of its role in the processing of amyloid precursor protein (APP) to release β amyloid peptides. But BACE is not an APP-dedicated protease, and the search has been intense for other substrates and physiological roles of the enzyme. In a new paper, Dora Kovacs and colleagues at Harvard Medical School reveal a role for BACE in regulating the expression of voltage-gated sodium channels in neurons. Writing in the June 18 online edition of Nature Cell Biology, the researchers show that BACE cleaves the β2 regulatory subunit of the Nav1 sodium channel. The end result, a loss of functional channels at the cell membrane, reduces sodium currents in the cells and thus affects membrane excitability.
The processing, which they show is increased in AD brain tissue due to elevated BACE levels, could explain why people with AD are more prone to seizures, a known consequence of dysregulated sodium channel function. In addition, the work suggests that BACE inhibitors, developed to target Aβ production (see ARF related news story), might also prevent seizures in AD patients.
Voltage-gated sodium channels are composed of a pore-forming α subunit, and one or two regulatory β subunits. Researchers knew that BACE could cleave the β2 subunit, a type 1 membrane protein (Wong et al., 2005), but the functional ramifications of the cleavage were not known. In the new study, lead authors Doo Yeon Kim and Bryce Carey provide a wealth of information about just how the subunit is processed and how BACE elevation in AD affects cell physiology.
The processing of β2 bears a striking resemblance to that of APP, they find. Overexpression of the β2 protein or BACE1 in neuronal cells leads to the release of a soluble ectodomain fragment, which they call β2-CTF. Then, they show, the remaining, membrane-bound portion of β2 is further processed by γ-secretase to release an intracellular domain fragment, the β2- ICD. The fragments are analogous to the APP products soluble APPα and the gene regulatory fragment AICD.
In a functional parallel to APP processing, the β2-ICD also regulates gene expression. The investigators show that β2 processing increases levels of Nav1.1 α subunit mRNA and protein. In primary rat cortical neurons, blocking endogenous BACE using inhibitors decreased β2-CTF and Nav1.1 mRNA. Treatment with a γ-secretase inhibitor increased β2-CTF accumulation, as predicted, and decreased Nav1.1 mRNA. From this, the researchers conclude that serial BACE and γ-secretase cleavage of β2 comprises a physiological pathway to regulate Nav1.1 levels. To show that β2-ICD is the relevant end product, they expressed the peptide in SH-SY5Y cells and confirmed a two- to threefold increase in Nav1.1 mRNA.
In vivo studies strengthened the BACE1- Nav1.1 connection. Transgenic mice overexpressing human BACE1 had elevation of endogenous β2-CTF and both Nav1.1 mRNA and protein in their brains. The extent of processing and Nav1.1 expression correlated with BACE levels over several lines of mice. Moreover, elevation of both β2-CTF and Nav1.1 were also seen in Western blots of brain tissue from AD patients with confirmed BACE overexpression, again with some correlation with BACE levels.
What of the functional ramifications of β2 cleavage? The researchers looked at sodium channel function by whole-cell voltage-clamp recording in B104 cells stably expressing β2 or β2 plus BACE1. Surprisingly, the presence of β-secretase decreased sodium current density. Among BACE-expressing cells, 65 percent totally lost sodium currents, while the rest had decreased average current density. The investigators obtained similar results with hippocampal slices from BACE1 mice, which displayed decreased sodium currents.
The researchers confirmed elevated Nav1.1 expression in the cells. However, a closer look revealed that the protein never reached the cell surface, but was retained intracellularly. In fact, there was a decrease in cell surface α subunits in both transfected cells and in the BACE1 mouse hippocampal slices.
“These data indicate that cleavage of β2 in BACE1-overexpressing cells results in a major reduction in the transfer of the Nav1s to the membrane, and a consequent reduction in membrane excitability,” the authors write. Their results expand the role of BACE in regulating neuron function through an expanding list of substrates. BACE was recently shown to play a role in the developmental myelination of neurons in the peripheral nervous system (see ARF related news story).
The alteration in Nav1.1 subcellular localization and activity in AD could have important repercussions for network activity in the brain. People with Alzheimer disease are at increased risk for seizures, and the “unusual” processing of β2 in brains from Alzheimer disease patients may lead to or contribute to epileptic symptoms, the researchers write. Mutations in Nav1 subunits cause epilepsy, and seizures can result from either gain or loss of function of the channels, leading to imbalances in sodium channel activity. They speculate that BACE inhibitors may have an unexpected benefit for the subset of AD patients who suffer seizures.—Pat McCaffrey
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- Lozsadi DA, Larner AJ. Prevalence and causes of seizures at the time of diagnosis of probable Alzheimer's disease. Dement Geriatr Cogn Disord. 2006;22(2):121-4. PubMed.
- Kim DY, Carey BW, Wang H, Ingano LA, Binshtok AM, Wertz MH, Pettingell WH, He P, Lee VM, Woolf CJ, Kovacs DM. BACE1 regulates voltage-gated sodium channels and neuronal activity. Nat Cell Biol. 2007 Jul;9(7):755-64. PubMed.