This Week in The Journal

Proteins Congregate in Lipid Rafts to Direct Axon Growth

Atsuko Honda, Yasuyuki Ito, Kazuko Takahashi-Niki, Natsuki Matsushita, Motohiro Nozumi, et al.

Rather than being distributed evenly throughout the plasma membrane, cellular signaling proteins are concentrated in lipid rafts, which are enriched in sphingolipids and cholesterol. Lipid rafts are particularly dense in neuronal membranes, as are their associated signaling proteins, which direct neuronal differentiation. Palmitoylated proteins aggregate in the detergent-resistant membrane (DRM) fraction of cell isolates, which contains lipid rafts.

In COS7 cells, GPM6a (red) colocalizes with cholesterol (green). Preventing GPM6a palmitoylation prevents this association. See Honda et al. for details.

To understand more about how lipid rafts coordinate signaling during growth, Honda et al. focused on the transmembrane palmitoylated glycoprotein M6a (GPM6a), a protein that congregates in lipid rafts. GPM6a is highly expressed in mature neurons and is a major component of the axon growth cone during development and synaptogenesis. The DRM from mouse brain homogenate contained GPM6a as well as a marker of lipid rafts. In cultured Neuro2a cells containing a mutant GPM6a that could not be palmitoylated, GPM6a was excluded from the DRM, suggesting that palmitoylation is required for lipid raft localization.

To probe the role of GPM6a in neuronal differentiation, the authors imaged lipid rafts in living growth cones of mouse cortical neurons and tracked the protein during the earliest phase of neural process formation. When plated on a natural substrate, laminin—but not the artificial surface polylysine—GPM6a accumulated asymmetrically in neurons and colocalized with a cholesterol indicator before neurite formation began, suggesting that GPM6a may play an early role in guiding development. Moreover, GPM6a was co-localized with downstream signaling proteins Rufy3, Rap2, and TIAM2/STEF in lipid rafts of growth cones.

Finally, to determine whether GPM6a was important for development of neural polarity and axon growth, the researchers knocked down expression of GPM6a using RNA interference in utero. A short hairpin RNA directed against GPM6a caused neurons to develop multiple, abnormally short neurites, and it delayed axon development. The defects were rescued by overexpression of wild-type GPM6a. The authors conclude that GPM6a palmitoylation is critical for localization in lipid rafts and coordination with its associated signaling proteins, which are responsible for directing neuronal polarization and development.

Astrocytes Mediate Clearance of Aβ in Alzheimer's Mouse Model

Chia-Chen Liu, Jin Hu, Na Zhao, Jian Wang, Na Wang, et al.

(see pages 4023–4031)

Early in Alzheimer's disease (AD), amyloid beta (Aβ) peptides accumulate and form plaques. The precise role Aβ deposits play in disease pathology remains unclear, but it is apparent that Aβ accumulation eventually overwhelms protein-clearing mechanisms, possibly contributing to neurodegeneration. Restoring protein clearance might therefore slow disease progression. Liu et al. asked how astrocytes might contribute to this process.

Low-density lipoprotein receptor-related protein 1 (LRP1) participates in Aβ metabolism in other cells, and blocking LRP1 impairs Aβ degradation by astrocytes. Liu et al. decreased expression of LRP1 in primary cultured astrocytes using short hairpin RNAs. Uptake of fluorescently labeled Aβ was significantly reduced in the LRP1 knockdown astrocytes compared to controls. LRP1 was not required for Aβ binding at the cell surface, but clearance of Aβ from the surface was less effective in cells with decreased LRP1 levels. Astrocytes control extracellular enzymes including insulin-degrading enzyme (IDE) and the matrix metalloproteases (MMPs), which degrade Aβ. After knock down of LRP1 by 70 percent, expression of IDE, MMP2, and MMP6 levels fell significantly.

To examine the role of astrocyte LRP1 in vivo, the researchers generated a conditional knockout mouse line lacking LRP1 only in astrocytes and crossed them with the APP/PS1 amyloid mouse model of AD (APP/PS1; aLrp1^−/−). LRP1 levels were significantly reduced in astrocytes of the cortex and hippocampus of APP/PS1; aLrp1^−/− mice compared to APP/PS1 mice. Importantly, Aβ plaques and soluble Aβ were elevated in the APP/PS1; aLrp1^−/− mice at 12 months. While loss of astrocyte LRP1 compromised degradation of soluble Aβ, it did not impair production of Aβ. In AD and in other neurodegenerative diseases, astrocytes express pro-inflammatory mediators that contribute to neuro-inflammation. Immunostaining revealed greater astrocyte activation in brains of APP/PS1; aLrp1^−/− mice compared to APP/PS1 mice. The results point to a critical role for LRP1 in astrocytes in clearing toxic Aβ from the diseased brain—one that might be exploited therapeutically for slowing AD progression.