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Regulation of the Nuclear PKA Holoenzyme
Author(s) -
Clister Terri,
Sample Vedangi,
Zhang Jin
Publication year - 2018
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2018.32.1_supplement.lb666
Subject(s) - adenylyl cyclase , microbiology and biotechnology , protein kinase a , cytosol , phosphodiesterase , chemistry , nuclear transport , kinase , nucleus , biophysics , biology , cell nucleus , signal transduction , biochemistry , enzyme
cAMP and PKA activity is regulated in time and space to ensure proper cellular responses. Recently a novel pool of activatable PKA holoenzyme was discovered to reside in the nucleus, expanding the traditional model of PKA signaling. Based on live‐cell imaging we hypothesize that the regulation of the nuclear PKA holoenzyme is dependent on phosphodiesterases (PDEs) anchored to A‐kinase Anchoring Proteins (AKAPs) which create a local microdomain with a lower concentration of cAMP, thus controlling the activation of anchored nuclear PKA. Using superresolution (pcSOFI) and FRAP techniques we've characterized AKAP95 in the nucleus. Co‐IP experiments using nuclear extracts revealed that both the RIIα subunit of the PKA holoenzyme, and multiple PDE4D isoforms interact with AKAP95, supporting our hypothesis and providing specific candidates to further explore this regulation. Our ICUE3 cAMP biosensor was localized to AKAP95 to measure the levels of cAMP directly around AKAP95 in response to differently localized sources of cAMP. We are able to generate cAMP in different cellular compartments in a dose‐dependent manner by targeting genetically encoded soluble adenylyl cyclase (sAC) and stimulating transfected cells with sodium bicarbonate. When a low concentration of cAMP is generated, only cAMP produced in the nucleus can be detected in the vicinity of AKAP95, whereas cAMP produced at the plasma membrane was not detected within this microdomain. These results support the hypothesis that AKAP95 and PDE4Ds create a microdomain that prevents the activation of scaffolded nuclear PKA holoenzyme except by local cAMP signals. We are still exploring the endogenous source of nuclear PKA activation, but hypothesize that cAMP generated by internalized GPCR signaling at the endosome could activate AKAP95‐tethered PKA. Support or Funding Information This work was funded by R01 DK073368. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .