The same journal carries a report linking a novel gene to neurodegeneration, through a pathway that is familiar to AD researchers. The pathway is endosomal trafficking and the gene is Vac14, a regulator of the signaling lipid phosphatidylinositol 3,5-bisphosphate (PI(3,5)P2). Lois Weisman and colleagues at the University of Michigan, Ann Arbor, show that Vac14 knockout mice develop large vacuoles in neurons, a result of defects in late endosomal transport. This is the same cellular domain where the sorting receptor SORL1 regulates APP processing (see ARF related news story). Polymorphisms in the SORL1 gene raise the risk of late-onset AD (see ARF related news story).

Although Vac14 is expressed in all cells, the knockout seems to affect neurons selectively, suggesting that these cells have a special weakness when it comes to deficits in endosomal function.

As described in the first report, the AMPA receptor channel mutations were discovered during a search for genes involved in X-linked metal retardation. Using cDNA microarray screening, first author Ye Wu identified one man with moderate mental retardation who showed a 10-fold reduction in expression of GRIA3, the gene encoding the ionotropic glutamate receptor subunit GluR3. Even though the investigators could not identify a mutation responsible for the low expression in that person, the discovery stimulated them to sequence the same gene from 400 additional unrelated men with X-linked mental retardation.

Doing this, they found one deletion and four mutations in GluR3, all missense mutations in highly conserved residues in functionally important areas of the protein. By expressing each variant in cultured cells, the investigators found that one mutation caused far lower expression of the Glu receptor, while the others encoded receptors with altered channel function. In whole cell recording experiments, two of these carried minimal or no current, and one displayed altered deactivation kinetics. When incorporated into heteromers with the GluR2 subunit, the mutant R3 proteins produced altered channel kinetics compared to wild-type R2/R3 receptors, showing that the mutations cause measurable changes in AMPA receptor function.

Patients with the various GluR3 mutations all showed similar clinical profiles, including moderate mental retardation, and muscular abnormalities. Thus, the authors conclude, significantly reduced iGluR3 channel function due to mutations in critical domains of the subunit is associated with a distinct neurological phenotype in humans. Whether the problem comes from failure of synaptic potentiation in the mature brain, or results from altered development is not clear, but further studies using mouse models may shed light on this question.

In the second report, lead author Yanling Zhang produced Vac14 knockout mice in order to investigate the physiological roles of PI(3,5)P2. Vac14, together with its partner protein Fig4, regulates the activity of the Fab1 kinase, which produces PI(3,5)P2 from PI3P. In the knockouts, the level of PI(3,5)P2 and PI5P were reduced by half compared to wild-type, while PI3P increased. Mice with Vac14 knockout displayed large vacuoles in neurons in parts of the brain and peripheral nervous system, and neuronal apoptosis. Some parts of the brain, notably the hippocampus and cortex, were less affected. This phenotype resembles that of the Fig4 knockout mouse, which further supports the idea that changes in PI(3,5)P2 are causing the effects. In addition, in humans, heterozygotic Fab1 mutations cause a mild disease where vacuoles appear in the cornea, again pointing to the importance of PI(3,5)P2 in vacuole formation.

Interestingly, all other organs in the mice appeared normal, despite the fact that Vac14 is present in all tissues. However, although Vac14-lacking fibroblasts appeared normal in vivo, they formed vacuoles in culture, and so the investigators used these cells to investigate the phenotype further. The vacuoles were derived from macropinocytotic vesicles, they found, and carried markers of late endosomes and lysosomes, suggesting that the vacuoles result from swelling of these organelles. The researchers also observed defects in trafficking of proteins from the trans-Golgi network to endosomes. This is the same pathway in which SORL1 and another Alzheimer-related protein, Vps35 function (Small et al., 2005).

Could PI(3,5)P2 have a role to play in AD? The idea is not far-fetched—other phosphoinositides have been linked to AD, most notably the 4,5 phosphates, where presenilin mutations linked to AD have been shown to cause an imbalance in PI(4,5)P which affects calcium channel activity in neurons (see ARF related news story).—Pat McCaffrey.

References:
Wu Y, Arai AC, Rumbaugh G, Srivastava AK, Turner G, Hayashi T, Suzuki E, Jiang Y, Zhang L, Rodriguez J, Boyle J, Tarpey P, Raymond RL, Nevelsteen J, Froyen G, Stratton M, Futreal A, Gecz J, Schwartz CE, Valle D, Huganir RL, Wang T. Mutations in ionotropic AMPA receptor 3 alter channel properties and are associated with moderate cognitive impairment in humans. Proc Natl Acad Sci U S A. 2007 Oct 24; [Epub ahead of print] Abstract

Zhang Y, Zolov SN, Chow CY, Slutsky SG, Richardson SC, Piper RC, Yang B, Nau JJ, Westrick RJ, Morrison SJ, Meisler MH, Weisman LS. Loss of Vac14, a regulator of the signaling lipid phosphatidylinositol 3,5-bisphosphate, results in neurodegeneration in mice. Proc Natl Acad Sci U S A. 2007 Oct 24; [Epub ahead of print] Abstract