Gene for Identifying Individuals with Familial Dysautonomia

Abstract

This invention relates to methods and compositions useful for detecting mutations which cause Familial Dysautonomia. Familial dysautonomia (FD; Riley-Day syndrome), an Ashkenazi Jewish disorder, is the best known and most frequent of a group of congenital sensory neuropathies and is characterized by widespread sensory and variable autonomic dysfunction. Previously, we mapped the FD gene, DYS, to a 0.5 cM region of chromosome 9q31 and showed that the ethnic bias is due to a founder effect, with >99.5% of disease alleles sharing a common ancestral haplotype. To investigate the molecular basis of FD, we sequenced the minimal candidate region and cloned and characterized its 5 genes. One of these, IKBKAP, harbors two mutations that can cause FD. The major haplotype mutation is located in the donor splice site of intron 20. This mutation can result in skipping of exon 20 in the mRNA from FD patients, although they continue to express varying levels of wild-type message in a tissue-specific manner. RNA isolated from patient lymphoblasts is primarily wild-type, whereas only the deleted message is seen in RNA isolated from brain. The mutation associated with the minor haplotype in four patients is a missense (R696P) mutation in exon 19 that is predicted to disrupt a potential phosphorylation site. Our findings indicate that almost all cases of FD are caused by an unusual splice defect that displays tissue-specific expression; and they also provide the basis for rapid carrier screening in the Ashkenazi Jewish population.

1 - 43 . (canceled) 
     
     
         44 . A kit for assaying for the presence of a mutation associated with Familial Dysautonomia in an individual comprising primers consisting of at least 16 contiguous nucleotides of the human IKBKAP gene as defined according to its cDNA sequence in SEQ ID NO: 2 or the complement thereof, said primers being complementary to sequences flanking the mutation site and capable of amplifying a region of the IKBKAP gene of sufficient size to detect the G-C mutation at position 73 of exon 19 of the human IKBKAP gene, wherein said amplified region comprises a G-C mutation at position 73 of exon 19 of the human IKBKAP gene. 
     
     
         45 . An isolated oligonucleotide probe suitable for the detection of a mutation associated with Familial Dysautonomia in an individual, the oligonucleotide probe consisting of at 16 contiguous nucleotides of the human IKBKAP gene as defined according to its cDNA sequence in SEQ ID NO: 2 or the complement thereof, said oligonucleotide probe being complementary to either the coding or non-coding strand and being suitable for detection of the G-C mutation at position 73 of exon 19 of the human IKBKAP gene. 
     
     
         46 . A method for detecting a mutation associated with Familial Dysautonomia in a sample, comprising DNA isolated from an individual, by amplifying the DNA sequence in the region flanking the portion of the human IKBKAP gene as defined according to its cDNA sequence in SEQ ID NO: 2 using primers consisting of at least 16 contiguous nucleotides of the human IKBKAP or the complement thereof, said primers being complementary to sequences flanking the mutation site and being capable of amplifying a region of the IKBKAP gene of sufficient size to detect the G-C mutation at position 73 of exon 19 of the human IKBKAP gene, wherein said amplified region comprises a G-C mutation at position 73 of exon 19 of the human IKBKAP gene, wherein said amplified region comprises a G-C mutation at position 73 of exon 19 of the human IKBKAP gene, and sequencing the amplified region.