Gap junctions are integral membrane proteins that enable the direct cytoplasmic exchange of ions and low-molecular-mass metabolites between adjacent cells. They provide a pathway for propagating and/or amplifying the signal transduction cascades triggered by cytokines, growth factors, and other cell signaling molecules involved in growth regulation and development. Dysfunctional intercellular communication via gap junctions has been implicated in causing many human diseases. The objective of this project is to use a multi-disciplinary approach to identify the key intrinsic regulatory mechanisms that are responsible for Cx43 and Cx45 function. The central hypothesis is that unique intermolecular interactions within the divergent CT domain of Cxs affect gap junction regulation. More specifically, we hypothesize that in the failing heart, Cx43CT phosphorylation alters protein partner interactions leading to remodeling of Cx43 from the intercalated disc, and that dimerization of Cx45 CTs is, in part, responsible for the channel properties of Cx45 that distinguish it from Cx43 and for the dominant-negative effect of Cx45 in heteromeric channels with Cx43. It is well-known that the CT domains of Cxs are key regulators of channel properties, and that dimerization of cytosolic domains are key regulators of ion channels. This proposal is significant because discovery of how interactions mediated by the CT domain can be modulated would open the door to strategies to ameliorate the pathological effects of altered Cx regulation in the failing heart. The following Specific Aims are proposed to investigate this concept: 1) Define how tyrosine kinases down regulate Cx43 gap junction intercellular communication, 2) Determine how Cx43 phosphorylation alters protein partner interactions, and 3) Identify the importance and mechanism of Cx45CT dimerization.

Public Health Relevance

Mechanisms underlying the initiation and persistence of lethal cardiac rhythms are of significant clinical interest. Changes in connexin distribution, density, and/or properties are characteristic of arrhythmic heart disease; therefore, understanding what causes alterations in GJ properties in heart disease is essential for defining the pathological substrate and devising effective therapies. The rationale for the proposed research is that understanding the structural basis of Cx43 and Cx45 regulation will lead to improved strategies to therapeutically restore proper cardiac GJIC that has been altered due to ischemic injury and heart disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM072631-12
Application #
9431219
Study Section
Intercellular Interactions Study Section (ICI)
Program Officer
Nie, Zhongzhen
Project Start
2006-06-01
Project End
2020-01-31
Budget Start
2018-02-01
Budget End
2019-01-31
Support Year
12
Fiscal Year
2018
Total Cost
Indirect Cost
Name
University of Nebraska Medical Center
Department
Biochemistry
Type
Schools of Medicine
DUNS #
168559177
City
Omaha
State
NE
Country
United States
Zip Code
68198
Trease, Andrew J; Capuccino, Juan M V; Contreras, Jorge et al. (2017) Intramolecular signaling in a cardiac connexin: Role of cytoplasmic domain dimerization. J Mol Cell Cardiol 111:69-80
Jacobsen, Nicole L; Pontifex, Tasha K; Li, Hanjun et al. (2017) Regulation of Cx37 channel and growth-suppressive properties by phosphorylation. J Cell Sci 130:3308-3321
Li, Hanjun; Spagnol, Gaelle; Zheng, Li et al. (2016) Regulation of Connexin43 Function and Expression by Tyrosine Kinase 2. J Biol Chem 291:15867-80
Spagnol, Gaelle; Kieken, Fabien; Kopanic, Jennifer L et al. (2016) Structural Studies of the Nedd4 WW Domains and Their Selectivity for the Connexin43 (Cx43) Carboxyl Terminus. J Biol Chem 291:7637-50
Ambrosi, Cinzia; Ren, Cynthia; Spagnol, Gaelle et al. (2016) Connexin43 Forms Supramolecular Complexes through Non-Overlapping Binding Sites for Drebrin, Tubulin, and ZO-1. PLoS One 11:e0157073
Spagnol, Gaƫlle; Al-Mugotir, Mona; Kopanic, Jennifer L et al. (2016) Secondary structural analysis of the carboxyl-terminal domain from different connexin isoforms. Biopolymers 105:143-62
Bahl, Kriti; Xie, Shuwei; Spagnol, Gaelle et al. (2016) EHD3 Protein Is Required for Tubular Recycling Endosome Stabilization, and an Asparagine-Glutamic Acid Residue Pair within Its Eps15 Homology (EH) Domain Dictates Its Selective Binding to NPF Peptides. J Biol Chem 291:13465-78
Kopanic, Jennifer L; Schlingmann, Barbara; Koval, Michael et al. (2015) Degradation of gap junction connexins is regulated by the interaction with Cx43-interacting protein of 75 kDa (CIP75). Biochem J 466:571-85
Spagnol, Gaelle; Reiling, Calliste; Kieken, Fabien et al. (2014) Chemical shift assignments of the C-terminal Eps15 homology domain-3 EH domain. Biomol NMR Assign 8:263-267
Kopanic, Jennifer L; Al-mugotir, Mona H; Kieken, Fabien et al. (2014) Characterization of the connexin45 carboxyl-terminal domain structure and interactions with molecular partners. Biophys J 106:2184-95

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