Methods of detecting Charcot-Marie Tooth disease type 2A
Methods are described for screening a subject for risk of Charcot-Marie-Tooth Disease Type 2A or for diagnosing Charcot-Marie-Tooth disease or a predisposition for developing Charcot-Marie-Tooth disease in a subject, by detecting the presence or absence of a mutation in the mitofusin gene in a biological sample collected from the subject. Methods are also described for detecting the presence of a genetic polymorphism associated with Charcot-Marie-Tooth Disease Type 2A in a sample of patient nucleic acid, by amplifying a mitofusin gene sequence in the patient nucleic acid to produce an amplification product; and identifying the presence of a Charcot-Marie-Tooth Disease Type 2A associated polymorphism in the amplification product.
1. A method of screening a human subject for risk of Charcot-Marie-Tooth Disease Type 2A comprising:
assessing a biological sample collected from said subject for the presence of mutations in the mitofusin 2 gene, wherein the mutations comprise a C to T change at position 280 or a G to A change at position 281 in the coding sequence of the mitofusin 2 gene, or a mutation in the coding sequence of the mitofusin 2 gene that results in alteration of the arginine at position 94 in an amino acid sequence of mitofusin 2; and
determining whether the subject is at an increased risk of Charcot-Marie-Tooth Disease Type 2A,
wherein the presence of any one of said mutations in the mitofusin 2 gene is indicative of an increased risk.
2. The method according to claim 1 , wherein said biological sample comprises a chromosomal nucleic acid.
3. The method according to claim 2 , wherein the chromosomal nucleic acid is Chromosome 1 or a fragment thereof.
4. The method according to claim 1 , wherein a mutation results in a change in the amino acid sequence selected from the group consisting of Arg94Gln, and Arg94Trp.
5. The method of claim 1 , wherein said assessing step further comprises detecting whether said subject is homozygous for said mutation.
6. The method of claim 1 , wherein said assessing step further comprises detecting whether said subject is heterozygous for said mutation.
7. A method for detecting the presence of a mutation associated with Charcot-Marie-Tooth Disease Type 2A in a sample of human patient nucleic acid, comprising:
amplifying a mitofusin 2 gene sequence in the patient nucleic acid to produce an amplification product;
assessing the amplification product for the presence of mutations associated with Charcot-Marie-Tooth Disease Type 2A, wherein said mutations comprise a C to T change at position 280 or a G to A change at position 281 in the coding sequence of the mitofusin 2 gene, or a mutation in the coding sequence of the mitofusin 2 gene that results in an alteration of the arginine at position 94 in an amino acid sequence of mitofusin 2, and
identifying the presence of any one of said Charcot-Marie-Tooth Disease Type 2A genetic mutations within the amplification product.
8. The method according to claim 7 , wherein the Charcot-Marie-Tooth Disease Type 2A mutation is identified by sequencing the amplification product.
9. The method according to claim 8 , further comprising digesting the amplification product with a restriction enzyme and identifying the Charcot-Marie-Tooth Disease Type 2A mutation by sequencing a restriction fragment.
10. The method according to claim 7 , wherein said mutation results in a change in the amino acid sequence selected from the group consisting of Arg94Gln, and Arg94Trp.
11. A method of diagnosing Charcot-Marie-Tooth Disease or a genetic predisposition for developing Charcot-Marie-Tooth Disease in a human subject, comprising:
providing a biological sample from the subject wherein said sample comprises a mitofusin 2 gene;
assessing the sample for mutations in the mitofusin 2 gene, wherein the mutations comprise positions a C to T change at position 280 or a G to A change at position 281 in the coding sequence of the mitofusin 2 gene, or a mutation in the coding sequence of the mitofusin 2 gene that results in an alteration of the arginine at position 94 in an amino acid sequence of mitofusin 2; and
determining that the subject has at least one of said mutations in at least one genomic copy of the mitofusin 2 gene, wherein the presence of at least one of said mutations in the mitofusin 2 gene is diagnostic for Charcot-Marie-Tooth Disease or a genetic predisposition for developing Charcot-Marie-Tooth Disease in the subject.
12. The method according to claim 11 , wherein said mutation results in a change in the amino acid sequence selected from the group consisting of Arg94Gln, and Arg94Trp.
13. The method according to claim 11 , wherein said determining step further comprises detecting whether said subject is heterozygous for said mutation.
14. The method according to claim 11 , wherein said determining step further comprises detecting whether said subject is homozygous for said mutation.
15. The method according to claim 1 , further comprising assessing the biological sample for one or more additional mutations in the coding sequence of the mitofusin 2 gene selected from the group consisting of: G2219C, C1252T, G839A, G821A, C751G, C493G, C314T, T227C and G205T, wherein the presence of any one of said mutations is indicative of an increased risk.
16. The method according to claim 1 , further comprising assessing the biological sample for one or more additional mutations in the coding sequence of the mitofusin 2 gene that result in an alteration in the amino acid sequence of mitofusin 2, selected from the group consisting of: Trp740Ser; Arg418Stop, Arg280His, Arg274Gln, Pro251Ala, His165Asp, Thr105Met, Leu76Pro and Val69Phe, wherein the presence of any one of said mutations is indicative of an increased risk.
17. The method according to claim 7 , further comprising assessing the amplification product for one or more additional mutations in the coding sequence of the mitofusin 2 gene selected from the group consisting of: : G2219C, C 1252T, G839A, G821A, C751G, C493G, C314T, T227C and G205T.
18. The method according to claim 7 , further comprising assessing the amplification product for one or more additional mutations in the coding sequence of the mitofusin 2 gene that result in an alteration in the amino acid sequence of mitofusin 2, selected from the group consisting of: Trp740Ser; Arg418Stop, Arg280His, Arg274Gln, Pro251Ala, His165Asp, Thr105Met, Leu76Pro and Val69Phe.
19. The method according to claim 11 , further comprising assessing the biological sample for one or more additional mutations in the coding sequence of the mitofusin 2 gene selected from the group consisting of: G2219C, C1252T, G839A, G821A, C751G, C493G, C314T, T227C and G205T, wherein the presence of any one of said mutations is diagnostic for Charcot-Marie-Tooth Disease or a genetic predisposition for developing Charcot-Marie-Tooth Disease.
20. The method according to claim 11 , further comprising assessing the biological sample for one or more additional mutations in the coding sequence of the mitofusin 2 gene that result in an alteration in the amino acid sequence of mitofusin 2, selected from the group consisting of: Trp740Ser; Arg418Stop, Arg280His, Arg274Gln, Pro251Ala, His165Asp, Thr105Met, Leu76Pro and Val69Phe, wherein the presence of any one of said mutations is diagnostic for Charcot-Marie-Tooth Disease or a genetic predisposition for developing Charcot-Marie-Tooth disease.