Operon-driven antagonism and novel modulation of DNA damage sensor serine/threonine protein kinases-specific protein phosphatase 2C (PP2C) phosphatase in Deinococcus radiodurans.

This study identifies a novel operon-driven signaling module in Deinococcus radiodurans. The operon includes a von Willebrand A domain protein (DRA0331), a serine/threonine protein kinases; STPK (DRA0332), a canonical Forkhead-associated (FHA)-domain protein (DRA0333), and a Protein Phosphatase 2C (PP2C)-type phosphatase (DRA0334). DRA0334 shows Mn2+-dependent phosphatase activity and has a unique dual-domain structure that combines a Kinase-interacting FHA (KI-FHA) domain with a PP2C catalytic domain. Functional assays show that the FHA-domain protein, DRA0333, boosts the phosphorylation of STPKs such as DNA damage-responsive serine/threonine-protein kinase A (RqkA) and putative STPK (DR1243), while the operonic partner PP2C-type phosphatase, DRA0334, counteracts this through targeted dephosphorylation, establishing a phospho-regulatory antagonism. Notably, the KI-FHA domain of the DRA0334 phosphatase competitively interacts with the FHA domain to modulate the radiation-responsive RqkA kinase, thereby maintaining kinase-phosphatase balance. This KI-FHA domain also imparts substrate specificity and enables feedback regulation. Additionally, DRA0334 modular variants confirm separate roles of catalytic and docking modules, and STRING analyses link DRA0334 functions to DNA repair and stress recovery. Collectively, the findings suggest an operonic connection between DRA0333 and DRA0334, indicating that the KI-FHA and FHA domains may act as phospho-docking switches. These switches can regulate both kinase and phosphatase activities in a push-pull regulatory mechanism within the phosphorylation-dephosphorylation cycle of signal transduction, depending on their association with the type of catalytic domain.