ECE2016 Oral Communications Receptors & Signalling (5 abstracts)
PRKACA mutations in adrenal Cushing impair association with the PKA regulatory subunit
Kerstin Bathon 1, , Isabel Weigand 4 , Cristina L. Ronchi 3 , Guido Di Dalmazi 5 , Felix Beuschlein 5 , Silviu Sbiera 4 , Martin Fassnacht 3, & Davide Calebiro 1,
1Institute of Pharmacology and Toxicology, Würzburg, Germany; 2Rudolf Virchow Center, DFG-Research Center for Experimental Biomedicine, Würzburg, Germany; 3Comprehensive Cancer Center Mainfranken, Würzburg, Germany; 4Department of Medicine I, Endocrine and Diabetes Unit, Würzburg, Germany; 5Medizinische Klinik und Poliklinik IV, Munich, Germany.
In a previous study we found mutations in the main catalytic subunit of protein kinase A (PKA Cα) to be responsible for cortisol-secreting adrenocortical adenomas (ACAs). These mutations interfere with the formation of a stable holoenzyme, thus causing constitutive PKA activation. More recently, we identified additional mutations affecting PKA Cα in ACAs associated with overt Cushing syndrome: Ser213Arg_Leu212_Lys214insIle-Ile-Leu-Arg, Cys200_Gly201insVal, Trp197Arg, del244-248+Glu249Gln, Glu32Val.
Here, we have performed a functional characterization of these new PKA Cα mutations. Specifically, we evaluated the basal association between PKA regulatory and catalytic subunits using co-immunoprecipitation as well as PKA activity with a kemptide assay.
All analyzed mutations with the exception of Glu32Val showed a reduced interaction with the regulatory (RIIβ) subunit. The interaction with the regulatory (RIα) subunit was affected also for all mutants except Glu32Val and Trp197Arg. The control of cAMP on the dissociation of the regulatory and the catalytic subunit was not affected. Two mutations (del244-248+Glu249Gln and Trp197Arg) caused an increased basal PKA activity. Two mutations (Cys200_Gly201insVal and Ser213Arg_Leu212_Lys214insIle-Ile-Leu-Arg) were apparently inactivating, at least using kemptide as a substrate. The Glu32Val mutation did not affect the interaction with the regulatory subunit and showed a slightly increased PKA activity.
These findings suggest that interference with the formation of a stable PKA holoenzyme is a main mechanism leading to constitutive PKA activation in cortisol-secreting ACAs. Further studies are required to investigate the biological relevance and mechanisms of activation of the variants associated with decreased PKA activity.
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Endocrine Abstracts
ISSN 1470-3947 (print) | ISSN 1479-6848 (online)
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