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Abstract: Lamin A/C mutations are the most common cause of familial dilated cardiomyopathy (DCM) but the pathogenetic mechanisms are incompletely understood. Nesprins are spectrin repeat-containing proteins that interact with lamin A/C and are components of the linker-of-nucleoskeleton-and-cytoskeleton (LINC) complex that connects the nuclear envelope to the actin cytoskeleton. Our aim was to determine whether changes in nesprin-1 and actin might contribute to DCM in homozygous Lmna knockout (Lmna −/−) mice. Here we find that Lmna −/− cardiomyocytes have altered nuclear envelope morphology, disorganization of nesprin-1 and heterogeneity in the distribution of nuclear and cytoskeletal actin. Functional interactions of nesprin-1 with nuclear G-actin and with the cytoskeletal γ-actin, α-cardiac actin and α-smooth muscle actin (α-SMA) isoforms were shown by immunoprecipitation and Western blotting. At 4–6weeks of age, Lmna −/− mice had normal levels of γ-actin and α-cardiac actin, but α-SMA expression was increased by 50%. In contrast to the predominant vascular distribution of α-SMA in WT ventricular sections, α-SMA had a diffuse staining pattern in Lmna −/− sections. Osmotic swelling studies showed enhanced radial swelling in Lmna −/− cardiomyocytes indicative of cytoskeletal instability. The distensibility of Lmna −/− cardiomyocytes with osmotic stress was reduced by addition of α-SMA-specific fusion peptide. Our findings support a model in which uncoupling of the nucleus and cytoskeleton associated with disruption of the LINC complex promotes mechanical instability and defective force transmission in cardiomyocytes. Changes in the distribution and expression patterns of nuclear and cytoskeletal actin suggest that diverse transcriptional and structural defects may also contribute to DCM in Lmna −/− mice. [Copyright &y& Elsevier]