Remodeling of the cytoskeleton is central to the modulation of cell shape and migration. Filamin A, encoded by the gene FLNA, is a widely expressed protein that regulates re-organization of the actin cytoskeleton by interacting with integrins, transmembrane receptor complexes and second messengers 1,2 . We identified localized mutations in FLNA that conserve the reading frame and lead to a broad range of congenital malformations, affecting craniofacial structures, skeleton, brain, viscera and urogenital tract, in four X-linked human disorders: otopalatodigital syndrome types 1 (OPD1; OMIM 311300) and 2 (OPD2; OMIM 304120), frontometaphyseal dysplasia (FMD; OMIM 305620) and Melnick-Needles syndrome (MNS; OMIM 309350). Several mutations are recurrent, and all are clustered into four regions of the gene: the actin-binding domain and rod domain repeats 3, 10 and 14/15. Our findings contrast with previous observations that loss of function of FLNA is embryonic lethal in males but manifests in females as a localized neuronal migration disorder, called periventricular nodular heterotopia (PVNH; refs. 3-6). The patterns of mutation, X-chromosome inactivation and phenotypic manifestations in the newly described mutations indicate that they have gain-of-function effects, implicating filamin A in signaling pathways that mediate organogenesis in multiple systems during embryonic development.

X-linked hydrocephalus, spastic paraplegia type I and MASA syndrome are related disorders with loci in subchromosomal region Xq28. We have previously shown that X-linked hydrocephalus is caused by mutations in the gene for neural cell adhesion molecule L1 (L1CAM), an axonal glycoprotein involved in neuronal migration and differentiation. Here we report mutations of the L1 gene in MASA syndrome and SPG1, in addition to HSAS families. Two of the HSAS mutations would abolish cell surface expression of L1 and represent the first functional null mutations in this disorder. Our results indicate that these three syndromes from part of a clinical spectrum resulting from a heterogeneous group of mutations in the L1 gene.

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Mutations in the L1 gene induce a spectrum of human neurological disorders due to abnormal development of several brain structures and ®ber tracts. Among its binding partners, L1 immunoglobulin superfamily adhesion molecule (Ig CAM) associates with neuropilin-1 (NP-1) to form a semaphorin3A (Sema3A) receptor and soluble L1 converts Sema3A-induced axonal repulsion into attraction. Using L1 constructs containing missense pathological mutations, we show here that this reversion is initiated by a speci®c trans binding of L1 to NP-1, but not to L1 or other Ig CAMs, and leads to activation of the NO/cGMP pathway. We identi®ed the L1±NP-1-binding site in a restricted sequence of L1 Ig domain 1, as a peptide derived from this region could reverse Sema3A repulsive effects. A pathological L1 missense mutation located in this sequence speci®cally disrupts both L1±NP-1 complex formation and Sema3A reversion, suggesting that the cross-talk between L1 and Sema3A might participate in human brain development.

Familial incontinentia pigmenti (IP; MIM 308310) is a genodermatosis that segregates as an X-linked dominant disorder and is usually lethal prenatally in males. In affected females it causes highly variable abnormalities of the skin, hair, nails, teeth, eyes and central nervous system. The prominent skin signs occur in four classic cutaneous stages: perinatal inflammatory vesicles, verrucous patches, a distinctive pattern of hyperpigmentation and dermal scarring. Cells expressing the mutated X chromosome are eliminated selectively around the time of birth, so females with IP exhibit extremely skewed X-inactivation. The reasons for cell death in females and in utero lethality in males are unknown. The locus for IP has been linked genetically to the factor VIII gene in Xq28 (ref. 3). The gene for NEMO (NF-kappaB essential modulator)/IKKgamma (IkappaB kinase-gamma) has been mapped to a position 200 kilobases proximal to the factor VIII locus. NEMO is required for the activation of the transcription factor NF-kappaB and is therefore central to many immune, inflammatory and apoptotic pathways. Here we show that most cases of IP are due to mutations of this locus and that a new genomic rearrangement accounts for 80% of new mutations. As a consequence, NF-kappaB activation is defective in IP cells.

A locus for X-linked hydrocephalus (HSAS), which is characterized by mental retardation and enlarged brain ventricles, maps to the same subchromosomal region (Xq28) as the gene for neural cell adhesion molecule L1. We have found novel L1 mRNA species in cells from affected members of a HSAS family containing deletions and insertions produced by the utilization of alternative 3' splice sites. A point mutation at a potential branch point signal in an intron segregates with the disease and is likely to be responsible for the abnormal RNA processing. These results suggest that HSAS is a disorder of neuronal cell migration due to disruption of L1 protein function.

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The L1 cell adhesion molecule has six domains homologous to members of the immunoglobulin superfamily and five homologous to fibronectin type III domains. We determined the outline structure of the L1 domains by showing that they have, at the key sites that determine conformation, residues similar to those in proteins of known structure. The outline structure describes the relative positions of residues, the major secondary structures and residue solvent accessibility. We use the outline structure to investigate the likely effects of 22 mutations that cause neurological diseases. The mutations are not randomly distributed but cluster in a few regions of the structure. They can be divided into those that act mainly by changing conformation or denaturing their domain and those that alter its surface properties.