# Characterization of EFhd2 as a novel inhibitor of necroptosis and aging-related cardiovascular pathology

> **NIH NIH R21** · UNIVERSITY OF MISSOURI-COLUMBIA · 2020 · $240,358

## Abstract

PROJECT SUMMARY
The loss of cells through necrosis is a major cause of aging-related pathologies in many different organ
systems, including the cardiovascular system. Nevertheless, our understanding of the cellular pathways that
underlie necrosis is still very far from complete. Thus, our long-term goal is to identify and characterize the
molecular mechanisms that drive/regulate necrotic death and how they contribute to aging-associated
disease. Historically, necrosis was thought of as an “accidental”, unregulated process. However, recent work
has revealed that necrosis can be tightly regulated. One form of regulated necrosis, necroptosis, is canonically
activated by tumor necrosis factor-α (TNFα). TNFα receptor binding activates the kinase receptor-interacting
protein 1 (RIPK1). RIPK1 then binds to, phosphorylates, and activates the kinase RIPK3, which in turn binds
and phosphorylates the pseudokinase mixed lineage kinase domain-like (MLKL). MLKL then oligomerizes and
translocates to the plasma membrane, where it directly perforates the membrane. Intriguingly, we have found
that oxidative stress-induced necrosis is also partially dependent on activation of this pathway. However, while
the “core” components of the pathway (RIPK1, RIPK3, MLKL) have been identified and characterized, next to
nothing is known of the key proteins that modulate these core proteins. In particular, proteins that regulate
MLKL activation, the final execution step in necroptotic signaling, have yet to be established. We have
discovered the Ca2+-binding protein EF hand domain protein 2 (EFhd2) as a potential negative regulator of
MLKL. We have found that EFhd2 can bind to MLKL, and its overexpression attenuates necroptotic cell death,
while knockdown has the opposite effect. Thus, our central hypothesis is that EFhd2 inhibits necroptotic
signaling through a Ca2+-regulated interaction with MLKL, and that loss of EFhd2 accelerates cardiac
pathology during aging. The objective of the present application is to define the mechanisms by which
EFhd2 inhibits necroptosis and the consequences this has for the development of cardiac dysfunction in aged
mice. In Aim 1 we will determine the mechanisms by which EFhd2 inhibits MLKL activation, and the role Ca2+
plays in EFhd2's ability to inhibit necroptosis. In Aim 2 we will examine indices of necroptosis and cardiac
function in aged wildtype and Efhd2-/- mice. The rationale for the proposed research is that once the key
proteins that modulate necroptosis are identified, they can be targeted to improve the clinical outcome of
many aging-related diseases such as cardiomyopathy.

## Key facts

- **NIH application ID:** 9995268
- **Project number:** 1R21AG067702-01
- **Recipient organization:** UNIVERSITY OF MISSOURI-COLUMBIA
- **Principal Investigator:** Christopher P Baines
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $240,358
- **Award type:** 1
- **Project period:** 2020-09-30 → 2022-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9995268

## Citation

> US National Institutes of Health, RePORTER application 9995268, Characterization of EFhd2 as a novel inhibitor of necroptosis and aging-related cardiovascular pathology (1R21AG067702-01). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/9995268. Licensed CC0.

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