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Molecular Determinants and Signaling Effects of PKA RIα Phase Separation
Preprint   Open access

Molecular Determinants and Signaling Effects of PKA RIα Phase Separation

Julia C Hardy, Emily H Pool, Jessica G H Bruystens, Xin Zhou, Qingrong Li, Daojia R Zhou, Max Palay, Gerald Tan, Lisa Chen, Jaclyn L C Choi, …
bioRxiv : the preprint server for biology
Cold Spring Harbor laboratory
12/11/2023
DOI: 10.1101/2023.12.10.570836
PMCID: PMC10760030
PMID: 38168176
url
https://doi.org/10.1101/2023.12.10.570836View
Preprint (Author's original)This preprint has not been evaluated by subject experts through peer review. Preprints may undergo extensive changes and/or become peer-reviewed journal articles. Open Access

Abstract

Spatiotemporal regulation of intracellular signaling molecules, such as the 3',5'-cyclic adenosine monophosphate (cAMP)-dependent protein kinase (PKA), ensures the specific execution of various cellular functions. Liquid-liquid phase separation (LLPS) of the ubiquitously expressed PKA regulatory subunit RIα was recently identified as a major driver of cAMP compartmentation and signaling specificity. However, the molecular determinants of RIα LLPS remain unclear. Here, we reveal that two separate dimerization interfaces combined with the cAMP-induced release of the PKA catalytic subunit (PKA-C) from the pseudosubstrate inhibitory sequence are required to drive RIα condensate formation in cytosol, which is antagonized by docking to A-kinase anchoring proteins. Strikingly, we find that the RIα pseudosubstrate region is critically involved in the formation of a non-canonical R:C complex, which serves to maintain low basal PKA activity in the cytosol by enabling the recruitment of active PKA-C to RIα condensates. Our results suggest that RIα LLPS not only facilitates cAMP compartmentation but also spatially restrains active PKA-C, thus highlighting the functional versatility of biomolecular condensates in driving signaling specificity.

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