FMRP-PKA Activity Negative Feedback Regulates RNA Binding-Dependent Fibrillation in Brain Learning and Memory Circuitry

Fragile X mental retardation protein (FMRP) promotes cyclic AMP (cAMP) signaling. Using an in vivo protein kinase A activity sensor (PKA-SPARK), we find that Drosophila FMRP (dFMRP) and human FMRP (hFMRP) enhance PKA activity in a central brain learning and memory center. Increasing neuronal PKA activity suppresses FMRP in Kenyon cells, demonstrating an FMRP-PKA negative feedback loop. A patient-derived R140Q FMRP point mutation mislocalizes PKA-SPARK activity, whereas deletion of the RNA-binding argi-nine-glycine-glycine (RGG) box (hFMRP-ΔRGG) produces fibrillar PKA-SPARK assemblies colocalizing with ribonucleoprotein (RNP) and aggregation (thioflavin T) markers, demonstrating fibrillar partitioning of cytosolic protein aggregates. hFMRP-ΔRGG reduces dFMRP levels, indicating RGG-independent regulation. Short-term hFMRP-ΔRGG induction produces activated PKA-SPARK puncta, whereas long induction drives fibrillar assembly. Elevated temperature disassociates hFMRP-ΔRGG aggregates and blocks activated PKA-SPARK localization. These results suggest that FMRP regulates compartmentalized signaling via complex assembly, directing PKA activity localization, with FMRP RGG box RNA binding restricting separation via low-complexity interactions.