Nebulette knockout mice have normal cardiac function but show Z-line widening and upregulation of cardiac stress markers

Aims: Nebulette is a 109 kDa modular protein localized in the sarcomeric Z-line of the heart. In vitro studies have suggested a role of nebulette in stabilizing the thin filament, and missense mutations in the nebulette gene were recently shown to be causative for dilated cardiomyopathy and endocardial fibroelastosis in human and mice. However, the role of nebulette in vivo has remained elusive. To provide insights into the function of nebulette in vivo, we generated and studied nebulette-deficient (nebl-/-) mice. Methods and results: Nebl-/- mice were generated by replacement of exon 1 by Cre under the control of the endogenous nebulette pro- moter, allowing for lineage analysis using the ROSA26 Cre reporter strain. This revealed specific expression of nebul- ette in the heart, consistent with in situ hybridization results. Nebl-/- mice exhibited normal cardiac function both under basal conditions and in response to transaortic constriction as assessed by echocardiography and haemodynamic analyses. Furthermore, histological, IF, and western blot analysis showed no cardiac abnormalities in nebl-/- mice up to 8 months of age. In contrast, transmission electron microscopy showed Z-line widening starting from 5 months of age, suggesting that nebulette is important for the integrity of the Z-line. Furthermore, up-regulation of cardiac stress responsive genes suggests the presence of chronic cardiac stress in nebl-/- mice. Conclusion Nebulette is dispensable for normal cardiac function, although Z-line widening and up-regulation of cardiac stress markers were found in nebl-/- heart. These results suggest that the nebulette disease causing mutations have dominant gain-of-function effects.