The effect of lamin A-C loss or mutation on nuclear envelope proteomics and splicing factor deregulation in Emery-Dreifuss muscular dystrophy and dilated cardiomyopathy.

Abstract

Mutations in the LMNA gene, which encodes for lamins A and C (lamin A-C) of the nuclear lamina, cause a wide array of tissue-specific pathologies, mostly affecting cardiac and skeletal muscle tissues. How mutations in a single ubiquitously expressed gene lead to a broad spectrum of phenotypes has long been a dazzling question. Lamin A-C interacts with chromatin and proteins both at the nuclear envelope (NE) and in the nuclear interior, thus serving many functions ranging from structural support of the nucleus to gene regulation. Many LMNA mutations cause Emery-Dreifuss muscular Dystrophy (EDMD), a progressive muscle weakening and wasting disorder with conduction system malfunction and dilated cardiomyopathy (DCM), as well as DCM with conduction defects in the absence of skeletal muscle involvement. To date the pathological mechanisms underlying EDMD and DCM remain incompletely understood, particularly the incurable cardiac phenotype that leads to premature sudden death. In this study, we investigated two non-mutually exclusive hypotheses that could explain the disease phenotypes of EDMD and DCM: 1) Disrupted structural integrity of the nuclear lamina, caused by lamin A-C deficiency, alters the composition of tissue-specific proteins that interact with the NE; 2) Lamin A-C loss or mutation causes deregulation in RBM20, a muscle-specific RNA binding protein that has recently been identified to be highly implicated in familial cardiomyopathies. We also sought to get a better understanding of the pathophysiological role of RBM20 in the heart by identifying novel interacting partners. Using a phage display-based approach, we identified panels of peptides and protein mimics with preferential binding to WT and lamin A-C deficient nuclei. Among the novel proteins revealed by our assay, we showed that HuR interacts with lamin A-C and has altered cleavage and subcellular localization in the absence of lamin A-C. Using the same phage display screening method in combination with co-immunoprecipitation and proximity ligat