13 Channels Essential to Support Proper Cardiac Function

By Alomone Team • April 2024

The heart is an impressive feat of biological engineering, coordinating electrical and mechanical actions to keep us alive. At the heart of this synchrony, ion channels play a leading role. These proteins operate within functionally interdependent complexes, modulating and influencing one another to maintain cardiac rhythm. However, the full cast of these molecular interactions remains largely unknown. Currently, groundbreaking research from a group in Denmark has created a comprehensive map of these complex relationships, identifying the 13 channels indispensable for proper heart function.

Interactome and a Map of Cardiac Ion Channels

Ion channels guide the passage of ions across cell membranes. This flow of ions generates the electrical currents that drive the heart’s contractions. However, the amplitude and timing of these currents depend not only on the central ion channel proteins but also on a network of interaction partners.The channel proteins interact with a variety of other proteins, including some that help with correct protein localization and others that regulate cellular signaling pathways. This interaction network is impressively intricate, with some channels potentially interacting with hundreds of partners.

The importance of understanding these interactions becomes clear as variants of specific associated proteins have been linked with arrhythmia and other heart rhythm disorders. Thus, a thorough understanding of the cardiac ion channel interactome could offer insights into the molecular roots of heart rhythm disorders and might even uncover potential therapeutic targets.

In this groundbreaking study, the researchers provide a first-of-its-kind map of the cardiac ion channel protein network, drawn from experimental data from mouse heart tissue. They identified 13 different cardiac ion channels and their corresponding interaction networks using affinity purification and mass spectrometry (MS) based proteomics.

Identifying and Exploring a 13 Ion Channel Interaction Network

In this research, the investigators used a large collection of antibodies from Alomone Labs to isolate involved cardiac ion channels. After an initial screening involving 40 antibodies and MS-based proteomics, they identified 13 ion channels for further examination.

Their process included immunoprecipitating the corresponding 13 channels from membrane-rich cardiac lysates of male mice. They conducted this experiment in four replicates and analyzed precipitated protein components via reverse-phase liquid chromatography and MS (Figure 1 shows the team’s workflow here).

An Ion Channel Exploration Workflow

Figure 1. Workflow of the study. MS measurements of immunoprecipitated channels and their interactors and of control IPs from quadruplicate murine cardiac tissue lysates. Deep proteome measurements of the membrane-enriched mouse heart samples utilized in the IP experiments were also performed. Bioinformatics network analyses prioritized interactors for functional evaluation. A subset of interactors were evaluated for their functional impact on cardiac electrophysiology by STORM imaging, optical mapping in zebrafish KOs, and patch clamping of cardiomyocytes from mice with interactor genes silenced. From multi-omics data integration, the impact of each interactor in human electrophysiology is evaluated. b, Dendrogram from unsupervised hierarchical cluster analysis of protein intensities of proteins identified in IP experiments show that the four replicate experiments all cluster together. The clustering follows the bait replicates. c, Pearson correlation coefficients for protein intensities of the four Cacna1c replicate pulldown experiments. Pearson correlation coefficients are indicated in each scatter plot. Image and legend from Maurya, S.et al. Nat Cardiovasc Res 2, 673–692 (2023).

To separate specific protein interactors from nonspecific ones, the team used a combination of bait and control pulldowns and visualized the findings through volcano plots. They also incorporated data from their proteome measurements of the lysate samples to estimate missing values. In multiple instances, the team noticed that specific ion channel proteins immunoprecipitated with a different channel using antibodies distinct from the antibody targeting the specific ion channel protein (Figure 2). They also found that several cardiac ion channel networks intersect with one another, indicating that they are likely clustered at shared locations in the cardiomyocyte.

By conducting 64 concurrent affinity purification mass spectrometry/mass spectrometry (AP-MS/MS) experiments on cardiac tissues, this work identified 822 novel protein interactors for the 13 cardiac ion channels and detected 59 previously reported protein interactors, further validating the experimental results. Overall, the dataset marks a major stride in how we understand ion channel interactomes and this study represents the most comprehensive mapping of cardiac ion channel interactomes to date.

Networks of Shared Proteins Between Channels Found to Interact

Figure 2. Inter-channel networks. a–d, Kcnq1 and Kcnh2 (a), between Kcnj2, Kcnj3 and Kcnj5 (b), between Cacna1c, Kcnn3 and Kcnma1 (c), and between Kcnq1, Gja1 and Cacna1c (d). The inset panels show all shared interactors for these channels. The bait proteins are shown in red squares and the interactors in light-blue circles. Measured interactions are indicated by lines. Note the interactions between channel proteins. Image and legend from Maurya, S.et al. Nat Cardiovasc Res 2, 673–692 (2023).

Realizing the Connective Role of Cardiac Ion Channels

Alongside this, the team conducted a series of functional studies using optical mapping, super-resolution imaging, and electrophysiology to explore the physiological importance of ten identified interaction partners. Two key players, epsin-2 and gelsolin, were revealed to be significant regulators of the sodium current in heart cells, highlighting the value of the channel network data in unearthing potential points of therapeutic intervention.

Acknowledging the Gaps in Knowledge

The authors acknowledge significant gaps in their delineation of cardiac ion channel protein interactions. First, this research is grounded in mouse models, so further work is needed to ascertain the transferability of these findings to human hearts. Secondly, their stringent criteria for identifying protein interactors may have left out weak yet potentially relevant dynamic interactors. Thirdly, the protein interactions may occur at any point during their expression, a dynamic process from synthesis to degradation.

Translating Physical Interactions into Functional Understanding

In this impressively complex series of experiments, the researchers utilized multiple approaches, integrating immunoassays, proteomics, electrophysiology, bioinformatics, and genetic data from human populations to affirm the relevance of their findings. The team verified that over 92% of the ion channel interactors were also present in human hearts, suggesting that these findings may provide insight into human cardiac electrophysiology.

While this research marks a significant leap in understanding the cardiac ion channel interactome, it also underscores the complexity and dynamic nature of these interactions. This is just the first step in deciphering the precise mechanisms underlying these interactions and their impacts on heart function. Unraveling the mysteries of the heart’s electrical symphony may not be simple, but the benefits it could yield when tackling heart rhythm disorders make it a scientific endeavor worth pursuing.

Tools to Tackle Cardiac Channels

The researchers in this paper made extensive use of our antibodies. You can find a wide selection of tools for your own cardiac research here on the site. You might be interested in these:

Antibodies and Blocking Peptide Controls

Isotype Controls

Rabbit IgG Isotype Control (#RIC-001)