Identification and expression analysis of spastin gene mutations in hereditary spastic paraplegia.

Pure hereditary spastic paraplegia (SPG) type 4 is the most common form of autosomal dominant hereditary SPG, a neurodegenerative disease characterized primarily by hyperreflexia and progressive spasticity of the lower limbs. It is caused by mutations in the gene encoding spastin, a member of the AAA family of ATPases. We have screened the spastin gene for mutations in 15 families consistent with linkage to the spastin gene locus, SPG4, and have identified 11 mutations, 10 of which are novel. Five of the mutations identified are in noninvariant splice-junction sequences. Reverse transcription-PCR analysis of mRNA from patients shows that each of these five mutations results in aberrant splicing. One mutation was found to be "leaky," or partially penetrant; that is, the mutant allele produced both mutant (skipped exon) and wild-type (full-length) transcripts. This phenomenon was reproduced in in vitro splicing experiments, with a minigene splicing-vector construct only in the context of the endogenous splice junctions flanking the splice junctions of the skipped exon. In the absence of endogenous splice junctions, only mutant transcript was detected. The existence of at least one leaky mutation suggests that relatively small differences in the level of wild-type spastin expression can have significant functional consequences. This may account, at least in part, for the wide ranges in age at onset, symptom severity, and rate of symptom progression that have been reported to occur both among and within families with SPG linked to SPG4. In addition, these results suggest caution in the interpretation of data solely obtained with minigene constructs to study the effects of sequence variation on splicing. The lack of full genomic sequence context in these constructs can mask important functional consequences of the mutation.

Additional glomangioma families link to chromosome 1p: no evidence for genetic heterogeneity.

Venous malformations are a common abnormality of the vasculature that may occur sporadically or, more rarely, as an autosomal dominant trait. One familial form of venous malformations has previously been linked to chromosome 9p. Mutations in the gene encoding Tie2, an endothelial specific receptor tyrosine kinase, have been identified in four different families. Glomangiomas are a subtype of venous malformations with glomus cell involvement. These cutaneous lesions can be inherited as an autosomal dominant disease with reduced penetrance and variable expressivity. We present evidence of linkage to chromosome 1p21-1p22 using four new glomangioma families, with a combined maximum two-point lod score of 7.32 at marker D1S2804. Markers D1S2129 and D1S2881 define the 24-cM linkage interval determined by recombination within affected individuals. A recent report also showed linkage of the glomangioma locus to chromosome 1p. A total of 9 families now map to this region, suggesting a decreased likelihood of locus heterogenity in familial glomangiomas. Investigation of candidate genes within the interval should provide new insights into lesion formation in inherited venous malformations.

Allelic and locus heterogeneity in inherited venous malformations.

Venous malformations are low-flow vascular lesions consisting of disorganized thin-walled vascular channels. These can occur sporadically but also as an autosomal dominant condition termed venous malformations, cutaneous and mucosal (VMCM; OMIM 600195). In two large unrelated kindreds mapping to chromosome 9, the identical R849W missense mutation was identified in the first kinase domain of Tie2, an endothelial cell-specific receptor tyrosine kinase. We report here the identification of four new kindreds with inherited venous malformations. Unlike the initial two families described, these four families demonstrate allelic and locus heterogeneity. In one of these families, the R849W mutation co-segregates with the disease phenotype. Three other families with venous malformations lack this mutation. One of these families is linked to markers near TIE2 on chromosome 9. In this family, we identified a novel mutation within the first kinase domain of Tie2 resulting in a Y897S change. Results from COS-1 cell transfections using expression constructs containing either the R849W or the Y897S mutation suggest that the receptors containing either mutation show ligand-independent hyperphosphorylation. These results suggest a gain-of-function mechanism for development of venous malformations in these families. Of the two remaining families, one excludes linkage to the TIE2 locus, establishing the existence of at least one additional locus for dominantly inherited venous malformations.