Post-translational regulation of retinal IMPDH1 in vivo to adjust GTP synthesis to illumination conditions
- University of Barcelona, Spain
- University of Salamanca, Spain
- CIBERDEM, Spain
- ICFO-Institut de Ciencies Fotoniques, Spain
- Universitat de Barcelona, Spain
- University of Alcalá de Henares, School of Medicine, Spain
- Universidad de Salamanca, Spain
Abstract
We report the in vivo regulation of Inosine-5´-monophosphate dehydrogenase 1 (IMPDH1) in the retina. IMPDH1 catalyzes the rate-limiting step in the de novo synthesis of guanine nucleotides, impacting the cellular pools of GMP, GDP and GTP. Guanine nucleotide homeostasis is central to photoreceptor cells, where cGMP is the signal transducing molecule in the light response. Mutations in IMPDH1 lead to inherited blindness. We unveil a light-dependent phosphorylation of retinal IMPDH1 at Thr159/Ser160 in the Bateman domain that desensitizes the enzyme to allosteric inhibition by GDP/GTP. When exposed to bright light, living mice increase the rate of GTP and ATP synthesis in their retinas; concomitant with IMPDH1 aggregate formation at the outer segment layer. Inhibiting IMPDH activity in living mice delays rod mass recovery. We unveil a novel mechanism of regulation of IMPDH1 in vivo, important for understanding GTP homeostasis in the retina and the pathogenesis of adRP10 IMPDH1 mutations.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files.
Article and author information
Author details
Santiago López-Begines
David Fernández-Justel
Funding
Ministerio de Economía y Competitividad (BFU2016-80583-R)
- Ana Méndez
Laser Lab Europe (654148)
- Pablo Loza-Alvarez
Fundación Ramón Areces (XVII Edition Rare Diseases)
- Ana Méndez
Fundacio La Marató (20141730)
- Jordi Andilla
- Pablo Loza-Alvarez
- Ana Méndez
Ministerio de Economía y Competitividad (BFU2016-79237-P)
- Ruben M Buey
Instituto de Salud Carlos III (PI18/00754)
- Pedro de la Villa
Junta de Castilla y León (Graduate student fellowship)
- David Fernández-Justel
Ministerio de Economía y Competitividad (SEV-2015-0522)
- Pablo Loza-Alvarez
Centres de Recerca de Catalunya (CERCA Institutional Support)
- Pablo Loza-Alvarez
- Ana Méndez
Fundació Privada Cellex (ICFO Institutional Support)
- Pablo Loza-Alvarez
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: Pertaining to animal research, this study was conducted in accordance with the ARVO statement for the use of animals in ophthalmic and vision research and in compliance with acts 5/1995 and 214/1997 for the welfare of experimental animals of the autonomous community (Generalitat) of Catalonia; and approved by the ethics committee on animal experiments of the University of Barcelona (Generalitat Reference #9906, protocols Bell 216/17; 217/17 and 218/17).
Reviewing Editor
- Constance L Cepko, Harvard Medical School, United States
Publication history
- Received: February 26, 2020
- Accepted: March 30, 2020
- Accepted Manuscript published: April 7, 2020 (version 1)
- Version of Record published: April 22, 2020 (version 2)
Copyright
© 2020, Plana-Bonamaisó et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Post-translational regulation of retinal IMPDH1 in vivo to adjust GTP synthesis to illumination conditions
eLife 9:e56418.
https://doi.org/10.7554/eLife.56418
Further reading
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Cyclic GMP-dependent protein kinases (PKGs) are key mediators of the nitric oxide/cGMP signaling pathway that regulates biological functions as diverse as smooth muscle contraction, cardiac function, and axon guidance. Understanding how cGMP differentially triggers mammalian PKG isoforms could lead to new therapeutics that inhibit or activate PKGs, complementing drugs that target nitric oxide synthases and cyclic nucleotide phosphodiesterases in this signaling axis. Alternate splicing of PRKG1 transcripts confers distinct leucine zippers, linkers, and auto-inhibitory pseudo-substrate sequences to PKG Iα and Iβ that result in isoform-specific activation properties, but the mechanism of enzyme auto-inhibition and its alleviation by cGMP is not well understood. Here we present a crystal structure of PKG Iβ in which the auto-inhibitory sequence and the cyclic nucleotide binding domains are bound to the catalytic domain, providing a snapshot of the auto-inhibited state. Specific contacts between the PKG Iβ auto-inhibitory sequence and the enzyme active site help explain isoform-specific activation constants and the effects of phosphorylation in the linker. We also present a crystal structure of a PKG I cyclic nucleotide binding domain with an activating mutation linked to Thoracic Aortic Aneurysms and Dissections. Similarity of this structure to wild type cGMP-bound domains and differences with the auto-inhibited enzyme provide a mechanistic basis for constitutive activation. We show that PKG Iβ auto-inhibition is mediated by contacts within each monomer of the native full-length dimeric protein, and using the available structural and biochemical data we develop a model for the regulation and cooperative activation of PKGs.
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- Biochemistry and Chemical Biology
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