Is the ApoE-induced DLK activation responsible for ApoE-stimulated ERK1/2 phosphorylation? To address this question, we inhibited DLK using shRNA-mediated knockdowns or overexpression of MBIP (for MUK/DLK binding inhibitory protein, an endogenous inhibitor of DLK; Fukuyama et al., 2000). In addition, we constitutively increased DLK activity by overexpression on the background of the DLK knockdown (Fig. 2G, S2F). We then measured the effect of these manipulations, which did not impair neuronal survival (Fig. S2G), on ApoE3-induced increases in DLK protein levels and in ERK1/2 phosphorylation, and additionally monitored phosphorylation of MKK7, a MAP-kinase kinase that is a major DLK target in neurons (Merritt et al., 1999).
