To test how ApoE-induced stimulation of ERK1/2 enhances Aβ-production, we monitored the effect of ApoE on the levels of APP as the precursor to Aβ (Fig. 3A). We found that addition of ApoE2, ApoE3, and ApoE4 to human neurons co-cultured with MEFs increased both APP mRNA and protein levels 3- to 5-fold, again with the same ApoE4>ApoE3>ApoE2 potency rank order observed above (Fig. 4A–4C). This result suggests that ApoE stimulates APP gene transcription. The same ApoE-dependent increases in APP mRNA levels were observed in human neurons plated on matrigel (Fig. 4B) and were found in independently derived neurons differentiated from iPS cell lines (Fig. S3A, S3B). The effects of ApoE on APP levels were concentration- and time-dependent, and were specific to neurons (Fig. S3C–S3E). Strikingly, ApoE only enhanced APP, but not APLP1 and APLP2 mRNA levels, despite the high homology between APP and APLP1 and APLP2 (Fig. 4C, S3A, S3B). In addition to ApoE, the other glial factors that increased Aβ-levels in our initial screen also acted by stimulating APP synthesis (Fig. S4A, S4B).
