A synthetic peptide that prevents cAMP regulation in mammalian Hyperpolarization-activated Cyclic Nucleotide-regulated (HCN) channels
- University of Milan, Italy
- University of Florence, Italy
- Université de Montpellier, CNRS, INSERM, France
- Technische Universität Darmstadt, Germany
- Columbia University, United States
Abstract
Binding of TRIP8b to the cyclic nucleotide binding domain (CNBD) of mammalian HCN channels prevents their regulation by cAMP. Since TRIP8b is expressed exclusively in the brain, we envisage that it can be used for orthogonal control of HCN channels beyond the central nervous system. To this end, we have identified by rational design a 40-aa long peptide (TRIP8bnano) that recapitulates affinity and gating effects of TRIP8b in HCN isoforms (human hHCN1, mHCn2, rbHCN4) and in the cardiac current If in rabbit and mouse sinoatrial node cardiomyocytes. Guided by a NMR-derived structural model that identifies the key molecular interactions between TRIP8bnano and HCN CNBD, we further designed a cell-penetrating peptide (TAT-TRIP8bnano) which successfully prevented b-adrenergic activation of mouse If leaving the stimulation of the L-type calcium current (ICaL) unaffected. TRIP8bnano represents a novel approach to selectively control HCN activation, which yields the promise of a more targeted pharmacology compared to pore blockers.
Data availability
All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Figure 3.
Article and author information
Author details
Matteo Elia Mangoni
Funding
Fondazione Cariplo
- Anna Moroni
National Institutes of Health
- Anna Moroni
H2020 European Research Council
- Anna Moroni
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Animal experimentation: Experiments on rabbit SAN cells were performed using left-over cells obtained during experiments approved by the Animal Welfare Body of the University of Milan and by the Italian Ministry of Health (license n.1127/2015-PR). Animal procedures were conformed to the guidelines of the care and use of laboratory animals established by Italian and European Directives (D. Lgs no 2014/26, 2010/63/UE).Mouse primary pacemaker cells were isolated from adult C5B6/J mice as previously described (Mangoni and Nargeot, Cardiovasc Res 2001), in accordance with the Guide for the Care and Use of Laboratory Animals (eighth edition, 2011), published by the US National Institute of Health and European directives (2010/63/EU). The protocol was approved by the ethical committee of the University of Montpellier and the French Ministry of Agriculture (protocol N{degree sign}: 2017010310594939).
Copyright
© 2018, Saponaro 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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A synthetic peptide that prevents cAMP regulation in mammalian Hyperpolarization-activated Cyclic Nucleotide-regulated (HCN) channels
eLife 7:e35753.
https://doi.org/10.7554/eLife.35753
