Interactions of APP with its family members APLP1 and APLP2 have complicated not only the study of its internalization and processing but also the search for its physiological function. That this is not fully understood is frequently cited as one of the gaping holes of AD research. A number of functions have been established, though their precise molecular mechanisms remain elusive. At the Eibsee, Ulrike Mueller, now at the University of Heidelberg, offered an update on her longstanding project to dissect APP function genetically with a series of knockout and, most recently, knock-in mice strains. Mueller made the first APP knockout strain (Mueller et al., 1994), which retained some residual APP expression, as well as a second, complete knockout (Li et al., 1996). The mice’s subtle phenotype initially flummoxed scientists, as they had expected a more dramatic role for this ubiquitous and conserved protein. The mice were small and weak, had small brains with mild anatomic defects, were prone to seizures, and performed poorly in behavioral paradigms of learning and exploration. Mueller subsequently generated various combinations of knockouts and crosses with APLP1 and 2 strains, and in this way found out that these family members were compensating for the physiological roles of APP (Heber et al., 2000; also von Koch et al., 1997). APLP1 and 2 are similar to APP, are both expressed in brain but lack the Aβ sequence. Their compensation made defining precisely what APP itself does a difficult task. To remove all possible interference, Mueller’s group recently generated a triple-knockout strain (Herms, 2004). These mice showed a patterning defect in their cortical architecture that resembled a human disease called type 2 lissencephaly, in which newly born neurons in the developing cortex do not build up their layered architecture in the proper order. The APP/APLP1/APLP2 knockouts also had too few reelin-producing cells.

The question then arose as to which sections of the APP protein mediate which of its functions. To address it, Mueller knocked two different fragments of APP back into the mice lacking APP, and asked which snippets of APP could rescue which aspect of the original APP single-knockout phenotype. This work is ongoing, but initial data suggest that the product of APP shedding by α secretase, i.e., the secreted APPα ectodomain, is able to restore most of the functions lost in the APP single-knockout phenotype. (Results for knock-in rescue of the double-knockout phenotypes are not available yet.) This finding appears to raise new questions about the importance of intracellular AICD signaling and subsequent gene expression to APP function.

The role of the APP cytoplasmic tail in APP function is very much in flux these days, and it came up on a related issue, as well. APP clearly plays a role in the cortical positioning of embryonic neuroblasts, but exactly which parts of it mediate this process is unclear. Tracy Young in Selkoe’s laboratory used in-utero electroporation of RNAi to delete APP from the developing forebrain of mouse embryos. This acute, localized knockdown of APP prevented neuroblasts from migrating up toward the brain’s surface after their birth. Rescue experiments with different parts of APP suggested that its C-terminus is required for cortical migration, while some of Mueller’s studies point to the ectodomain. The phenotypes are similar but not the same. While both manipulations of APP—triple-knockout or acute RNAi—disturbed the migration of cortical neuroblasts, Mueller’s triple-knockout led some to overshoot their proper place in the cortical architecture. Indeed, in certain spots they burst through the pial membrane at the top of the brain, possibly because they failed to stick to the extracellular matrix there. By contrast, Young and Selkoe’s experiment prevented the neuroblasts from ever reaching the cortical layers in the first place; they instead massed near their birthplace at the subventricular zone. The experiments are sufficiently different to preclude direct comparison at this early stage, but the scientists agreed that these apparent discrepancies are important and will be worked out experimentally.—Gabrielle Strobel.