Differential regulation of protein phosphatase-1I by neurabin

Neurabin is a brain-specific actin and protein phosphatase-1 (PP-1) binding protein that inhibits the purified catalytic subunit of protein phosphatase-1 (PP-1C). However, endogenous PP-1 exists primarily as multimeric complexes of PP-1C bound to various regulatory proteins that determine its activity, substrate specificity, subcellular localization and function. The major form of endogenous PP-1 in brain is protein phosphatase-1I (PP-1I), a Mg2+/ATP-dependent form of PP-1 that consists of PP-1C, the inhibitor-2 regulatory subunit, an activating protein kinase and other unidentified proteins. We have identified four PP-1I holoenzyme fractions (PP-1IA, PP-1IB, PP-1IC, and PP-1ID) in freshly harvested pig brain separable by poly-l-lysine chromatography. Purified recombinant neurabin (amino acid residues 1–485) inhibited PP-1IB (IC50 = 1.1 μM), PP-1IC (IC50 = 0.1 μM), and PP-1ID (IC50 = 0.2 μM), but activated PP-1IA by up to threefold (EC50 = 40 nM). The PP-1IA activation domain was localized to neurabin1–210. Our results indicate a novel mechanism of PP-1 regulation by neurabin as both an inhibitor and an activator of distinct forms of PP-1I in brain.