Figure 11.

Schematic of cell labeling with ACSA-2. This schematic attempts to summarize our understanding of ACSA-2 binding to astrocytes. Multiple lines of experimental evidence suggest that the target of ACSA-2 is ATP1B2. ATP1B2 is a single-pass transmembrane protein, consisting of a short N-terminal intracellular region (which is protected from binding by ACSA-2 by the shielding effect of the plasma membrane) and a longer extracellular C-terminal domain. Although detailed epitope mapping was not undertaken, immunoblotting experiments, performed immediately after cell isolation, indicate that ATP1B2 is sensitive to papain, because the apparent molecular mass of the protein decreases from 55 to 25 kDa (data not shown). The proteolytically sensitive region is outlined with a dashed line in this schematic. As such, ACSA-2 must be binding ATP1B2 on an intact, membrane-proximal region, as illustrated, to be useful for immunoisolation.