Functional characterization of AC5 gain-of-function variants: Impact on the molecular basis of ADCY5-related dyskinesia

[Display omitted] Adenylyl cyclases are key points for the integration of stimulatory and inhibitory G protein-coupled receptor (GPCR) signals. Adenylyl cyclase type 5 (AC5) is highly expressed in striatal medium spiny neurons (MSNs), and is known to play an important role in mediating striatal dopa...

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Bibliographic Details
Published in:Biochemical pharmacology 2019-05, Vol.163, p.169-177
Main Authors: Doyle, T.B., Hayes, M.P., Chen, D.H., Raskind, W.H., Watts, V.J.
Format: Article
Language:English
Subjects:
AC5
Adenylyl cyclase
Adenylyl Cyclase Inhibitors - pharmacology
Adenylyl Cyclases - genetics
Adenylyl Cyclases - metabolism
cAMP
Dopamine
Dose-Response Relationship, Drug
Dyskinesia
Dyskinesias - genetics
Dyskinesias - metabolism
Gain of Function Mutation - physiology
Gene Knockdown Techniques
Genetic Variation - physiology
HEK293 Cells
Humans
Receptors, Dopamine D2 - agonists
Receptors, Dopamine D2 - genetics
Receptors, Dopamine D2 - metabolism
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Summary:[Display omitted] Adenylyl cyclases are key points for the integration of stimulatory and inhibitory G protein-coupled receptor (GPCR) signals. Adenylyl cyclase type 5 (AC5) is highly expressed in striatal medium spiny neurons (MSNs), and is known to play an important role in mediating striatal dopaminergic signaling. Dopaminergic signaling from the D1 expressing MSNs of the direct pathway, as well as the D2 expressing MSNs of the indirect pathway both function through the regulation of AC5 activity, controlling the production of the 2nd messenger cAMP, and subsequently the downstream effectors. Here, we used a newly developed cell line that used Crispr-Cas9 to eliminate the predominant adenylyl cyclase isoforms to more accurately characterize a series of AC5 gain-of-function mutations which have been identified in ADCY5-related dyskinesias. Our results demonstrate that these AC5 mutants exhibit enhanced activity to Gαs-mediated stimulation in both cell and membrane-based assays. We further show that the increased cAMP response at the membrane effectively translates into increased downstream gene transcription in a neuronal model. Subsequent analysis of inhibitory pathways show that the AC5 mutants exhibit significantly reduced inhibition following D2 dopamine receptor activation. Finally, we demonstrate that an adenylyl cyclase “P-site” inhibitor, SQ22536 may represent an effective future therapeutic mechanism by preferentially inhibiting the overactive AC5 gain-of-function mutants.
ISSN:0006-2952
1873-2968
DOI:10.1016/j.bcp.2019.02.005