ZBP1 subcellular localization and association with stress granules is controlled by its Z-DNA binding domains

Z-DNA binding protein 1 (ZBP1) belongs to a family of proteins that contain the Z alpha domain, which binds specifically to left-handed Z-DNA and Z-RNA. Like all vertebrate proteins in the Z alpha family, it contains two Z alpha-like domains and is highly inducible by immunostimulation. Using circular dichroism spectroscopy and electrophoretic mobility shift assays we show that both Z alpha domains can bind Z-DNA independently and that substrate binding is greatly enhanced when both domains are linked. Full length ZBP1 and a prominent splice variant lacking the first Z alpha domain (Delta Z alpha) showed strikingly different subcellular localizations. While the full length protein showed a finely punctate cytoplasmatic distribution, ZBP1 Delta Z alpha accumulated in large cytoplasmic granules. Mutation of residues important for Z-DNA binding in the first Z alpha domain resulted in a distribution comparable to that of ZBP1 Delta Z alpha. The ZBP1 Delta Z alpha granules are distinct from stress granules (SGs) and processing bodies but dynamically interacted with these. Polysome stabilization led to the disassembly of ZBP1 Delta Z alpha granules, indicating that mRNA are integral components. Heat shock and arsenite exposure had opposing effects on ZBP1 isoforms: while ZBP1 Delta Z alpha granules disassembled, full length ZBP1 accumulated in SGs. Our data link ZBP1 to mRNA sorting and metabolism and indicate distinct roles for ZBP1 isoforms.