Investigation into the Epicardial Requirement for Crk and CrkL Adaptor Proteins in the Mammalian Heart

Date of Award

5-2025

Degree Type

Thesis

Degree Name

Master of Science in Biomedical Sciences

First Advisor

Cathy J. Hatcher, PhD

Second Advisor

Philip Reno, PhD

Third Advisor

Marina D'Angelo, PhD

Abstract

Crk and CrkL are adaptor proteins that mediate intracellular signaling critical for tissue morphogenesis, yet their roles in epicardial function and coronary vascular development remain incompletely defined. This study investigates the requirement for Crk and CrkL in the epicardium using an epicardial-specific Crk-CrkL 3-allele conditional knockout (Crk-CrkL 3acKO) mouse model. Epicardial deletion of Crk and CrkL resulted in early reductions in compact myocardium thickness and impaired migration of Wilms’ Tumor 1 (Wt1)-positive epicardium-derived cells into the myocardium at E13.5, with partial recovery by E17.5. Vascular Endothelial (VE)-Cadherin immunostaining revealed superficial displacement of developing coronary vessels at early stages. Smooth Muscle 22 alpha (SM22α) and Periostin expression patterns appeared unaffected, although limited embryo availability restricted comprehensive evaluation of vascular smooth muscle and fibroblast contributions. Transcriptomic profiling of adult Crk-CrkL 3acKO mouse hearts revealed persistent alterations in gene expression, including dysregulation of pathways related to cardiovascular development and function, extracellular matrix remodeling, cellular function and metabolism, and immune activation in comparison to Crk-CrkL control hearts. We identified key dysregulated biological functions, canonical pathways, and upstream regulators using Ingenuity Pathway Analysis. This suggests that early epicardial defects have enduring molecular consequences. These findings establish Crk and CrkL as critical regulators of epicardial behavior, coronary vessel formation, and long-term cardiac gene expression.

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