The mouse ortholog of human FACE-1, Zmpste24, is a multispanning membrane protein widely distributed in mammalian tissues 1, 2 and structurally related to Afc1p/ste24p, a yeast metalloproteinase involved in the maturation of fungal pheromones 3. Disruption of the gene Zmpste24 caused severe growth retardation and premature death in homozygous-null mice. Histopathological analysis of the mutant mice revealed several abnormalities, including dilated cardiomyopathy, muscular dystrophy and lipodystrophy. These alterations are similar to those developed by mice deficient in A-type lamin 4, a major component of the nuclear lamina 5, and phenocopy most defects observed in humans with diverse congenital laminopathies 6, 7, 8. In agreement with this finding, Zmpste24-null mice are defective in the proteolytic processing of prelamin A. This deficiency in prelamin A maturation leads to the generation of abnormalities in nuclear architecture that probably underlie the many phenotypes observed in both mice and humans with mutations in the lamin A gene. These results indicate that prelamin A is a specific substrate for Zmpste24 and demonstrate the usefulness of genetic approaches for identifying the in vivo substrates of proteolytic enzymes.