Structural basis of PP2A phosphatase diseases

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PROJECT SUMMARY Protein phosphatase 2A (PP2A) is a major Ser/Thr phosphatase with complex regulation and composition. Deregulation of PP2A holoenzymes leads to devastating human diseases, including multiple types of cancer and neurological disorders. In recent years, whole exome/genome sequencing for the molecular diagnosis identified broad disease mutations in PP2A subunits and substrates in cancer and neurological disorders. The diagnosis of de novo PP2A mutations in developmental disorders enthused unprecedented multidisciplinary phosphatase research and interactions with patient families. De novo mutations in several members of B56 family of PP2A regulatory subunits, such as B56δ and B56γ, cause neurological disorders, known as Jordan Syndrome. The same somatic mutations were found in cancer patients. Built on our preliminary cryo-EM, molecular dynamic simulations, biochemical, single molecule, reverse phase protein array, and cell biology studies, here we aim to decipher the mechanisms of B56 disease mutations in cancer and intellectual disabilities. We will determine the cryo-EM structures of WT and disease variants of the PP2A-B56δ holoenzyme (Specific Aim 1), investigate how intellectual disability (ID) mutations alter holoenzyme dynamics, activation phosphorylation, and interactions with regulatory proteins, combining biochemical dissections, advanced molecular dynamic simulation approaches, and single-molecule fluorescence resonance energy transfer (smFRET) (Specific Aim 2), and test our hypothesis on the discriminating and merging mechanisms of B56δ and B56γ ID mutations in perturbing the cAMP negative feedback signaling via MAPK and CREB signaling, respectively (Specific Aim 3). These studies will shed light on a highly dynamic regulation machinery involving the folding of long disordered regions against the holoenzyme core and a super-long dynamic interface harboring the majority of residues mutated in Jordan Syndrome, multiple activation phophorylation sites, and regulatory elements that suppress both the phosphatase active site and the substrate-binding groove.