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Sunday, May 13, 2012

Is GDAP1 Causing CMT4?


We are moving on to the 3rd gene to test for CMT4 causing mutations. In the last two posts, we investigated genes FGD4 and FIG4. We found no known CMT4 causing mutations in Stacy's exome. We still have 4 out of 7 genes to check. As we move down the list, the chance that Stacy either was misdiagnosed or has a novel mutation causing CMT4 increases. However, we won't know for sure until we exhaust all known mutations. Here, we check GDAP1 for CMT4 causing mutations in Stacy's exome.


Start IGV, download Stacy's copy of GDAP1, and type in GDAP1 into the address bar at the top as shown in this previous post.

Below is a list of CMT4 causing mutations from the HMDG and the IGN mutation website. We will examine each CMT4 causing mutation in Stacy's GDAP1. As before, I will take a screenshot of anything of interest and discuss potential ramifications.

We have excluded known FGD4 and FIG4 causal mutations.
  1. FGD4  -- 5 mutations cause CMT4
  2. FIG4 -- 20 mutations cause CMT4
  3. GDAP1 -- 34 mutations cause CMT4 Download Stacy's GDAP1
  4. MTMR2 -- 10 mutations cause CMT4
  5. PRX -- 2 mutations cause CMT4
  6. SBF2 -- 5 mutations cause CMT4
  7. SH3TC2 - 19 mutations cause CMT4

Before examining GDAP1, make sure to right-click the box containing the text "RefSeq Genes" on the lower left of the IGV viewer. Then select "Expanded" as opposed to "Collapsed". You should now see two different GDAP1 variants at the bottom of the screen as shown in the image below.

These two variants of the gene are expressed by alternative splicing. Since the exons are different, the codon numbers are different as well. For our purposes, you can safely ignore the bottom GDAP1 and focus solely on the top version of the gene. Below is the list of the known GDAP1 causal mutations for CMT4 that we will analyze.

Mutations causing CMT4 in the GDAP1 gene.

  1. Trp31X Matches Reference Genome
  2. ?57? INSERTION/DELETION Matches Reference Genome
  3. Pro78Leu Matches Reference Genome
  4. Pro111His Matches Reference Genome
  5. Glu114fs FRAMESHIFT Matches Reference Genome
  6. Met116Thr Matches Reference Genome
  7. Met116Arg Matches Reference Genome
  8. Tyr117fs Matches Reference Genome
  9. Arg120Gln Matches Reference Genome
  10. Arg120Trp Matches Reference Genome
  11. Gln122Lys Matches Reference Genome
  12. His123Arg Matches Reference Genome
  13. Arg125X Matches Reference Genome
  14. Thr147fs Matches Reference Genome
  15. Asp149Tyr Matches Reference Genome
  16. Thr157Pro Matches Reference Genome
  17. Arg161His Matches Reference Genome
  18. Ser162fs Matches Reference Genome
  19. Gln163X Matches Reference Genome
  20. Arg191X Matches Reference Genome
  21. Codon 193 5' Splice site Matches Reference Genome
  22. Ser194X Matches Reference Genome
  23. Gln218Glu Matches Reference Genome
  24. Val219Asp Matches Reference Genome
  25. Leu223X Matches Reference Genome
  26. Gln235X Matches Reference Genome
  27. Leu239Phe Matches Reference Genome
  28. Arg257X Matches Reference Genome
  29. ?261? DELETION Matches Reference Genome
  30. Gly262fs Matches Reference Genome
  31. Gly271Arg Matches Reference Genome
  32. Arg282Cys Matches Reference Genome
  33. ?287? INSERTION Matches Reference Genome
  34. Asn297Lys Matches Reference Genome
This gene has had the most mutations to check so far with a total of 34. Mutation number 21 is a splice site mutation. Basically, you go to codon 193 and find that it's the last amino acid in the exon. Adjacent to codon 193, there can be a variant that causes alternative splicing, causing CMT4.


Below is a screenshot of Trp31X, the first mutation in the list above:

The actual location of the 31st codon ins circled in yellow. As you can see, everything is gray, meaning that this region matches the reference genome and this mutation is not causing CMT4 in Stacy. The other mutations are examined in this same way and all of the codons that cause CMT4 are gray in GDAP1. Therefore, all of the CMT4 causing mutations in this gene match the reference genome. We can conclude that no known CMT4 cuasing mutation is in GDAP1 is responsible for Stacy's diagnosis.

In this screen shot above near codon 31 (circled in yellow), you can see that there is a G-->C mutation. It is located on codon 34, which is not known to cause CMT4. Normally, I would not investigate since we are focusing solely on known CMT4 causing mutations, but I couldn't resist in this case since it is so close to a known CMT4 causing mutation. The codon at position 34 codes for is TCG, and therefore codes Ser. The mutation Stacy has is from TCG to TCC. It turns out that TCC also codes for Ser, so this is known as a "silent" mutation. It's unlikely that this mutation is causing problems since silent mutations don't change the protein itself.  However, they are known to interfere with splicing and silent mutations can also cause severe disease (Zich et al., A point mutation within CD45 exon A is the cause of variant CD45RA splicing in humans.  European Journal of Immunology, 1998). We therefore can not exclude this variant, but we will have to come back to it if we exhaust the list of known CMT causing variants before finding a causal mutation.


We have no eliminated 3 genes in the first phase of this project. It's too early to draw conclusions since we are still not halfway through the list of genes. After seeing how many CMT4 causing mutations exist and how frequently they are discovered, it would not surprise me if Stacy does have a novel (previously unknown) CMT4 causing mutation. It would be interesting to gather statistics and figure out how many CMT4 causing mutations are due to a premature stop codon, amino acid substitution, etc. If no known CMT4 causing mutation is identified, I will produce such statistics. I'd like to see if there is a bias towards of one type of mutation versus another in causing CMT4.

In the next post, we will examine MTMR2 for CMT4 causing mutations...

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