Chromatin-Associated Pea Apyrase psNTP9 Function as a DNA-Binding Regulatory Protein in Yeast and Arabidopsis

As reported in earlier work, when a pea apyrase, psNTP9 (PS), and a modified version of it, psNTP9-DM (DM), are expressed in Saccharomyces cerevisiae, they localize to nuclei, binding to largely non-overlapping promoter regions of chromatin. PS- and DM-expressing yeast also exhibit different expression profiles for potentially regulated target genes, consistent with observed phenotypes. In the present study, we use ChIP-seq assays to show that PS and DM also associate with largely different promoter regions of Arabidopsis genes, with similar non-overlapping expression profiles for potential target genes. Functional studies, using electrophoretic mobility shift assays (EMSA), verified PS-specific binding to yeast or plant promoter binding sites. DM binding to both heterologous dsDNA and to PS-specific binding site sequences was minimal. AlphaFold3 modeling of PS protein binding to a yeast PHM6 promoter sequence identified potential DNA-binding residues and a potential binding site motif (5'-(G/T)GG(G/T)A-3') that is also present in two Arabidopsis promoter binding sites. These novel findings extend the previously known functions of PS and other plant apyrases in the Golgi or extracellular matrix, and support their potential function as DNA-binding proteins that can regulate gene expression in both yeast and Arabidopsis.