Filamin-interacting proteins, Cfm1 and Cfm2, are essential for the formation of cartilaginous skeletal elements

Abstract Mutations of Filamingenes, which encode actin-binding proteins, cause a wide range of congenital developmental malformations in humans, mainly skeletal abnormalities. However, the molecular mechanisms underlying Filaminfunctions in skeletal system formation remain elusive. In our screen to identify skeletal development molecules, we found that Cfm (Fam101) genes, Cfm1 (Fam101b) and Cfm2 (Fam101a), are predominantly co-expressed in developing cartilage and intervertebral discs (IVDs). To investigate the functional role of Cfm genes in skeletal development, we generated single knockout mice for Cfm1 and Cfm2, as well as Cfm1/Cfm2 double-knockout (Cfm DKO) mice, by targeted gene disruption. Mice with loss of a single Cfm gene displayed no overt phenotype, whereas Cfm DKO mice showed skeletal malformations including spinal curvatures, vertebral fusionsand impairment of bone growth, showing that the phenotypes of Cfm DKO mice resemble those of FilaminB ( Flnb)-deficient mice. The number of cartilaginous cells in IVDs is remarkably reduced, and chondrocytesare moderately reduced in Cfm DKO mice. We observed increased apoptosis and decreased proliferation in Cfm DKO cartilaginous cells. In addition to direct interaction between Cfm and Filaminproteins in developing chondrocytes, we showed that Cfm is required for the interaction between Flnband Smad3, which was reported to regulate Runx2expression. Furthermore, we found that Cfm DKO primary chondrocytesshowed decreased cellular size and fewer actin bundles compared with those of wild-type chondrocytes. These results suggest that Cfms are essential partner molecules of Flnbin regulating differentiation and proliferation of chondryocytes and actin dynamics.