Poster Presentation Hunter Cell Biology Meeting 2022

Investigating the pathomechanism of CMTX3 neuropathy; how does intergenic structural variation alter local gene expression?  (#74)

Alexandra Boyling 1 2 , Anthony N Cutrupi 1 2 , Garth A Nicholson 1 2 3 , Gonzalo Perez-Siles 1 2 , Marina L Kennerson 1 2 3
  1. Northcott Neuroscience Laboratory, ANZAC Research Institute, Concord, NSW, Australia
  2. Sydney Medical School, University of Sydney, Sydney, NSW , Australia
  3. Molecular Medicine Laboratory, Concord Repatriation General Hospital, Sydney, NSW, Australia

Charcot-Marie-Tooth neuropathy (CMT) is an inherited disease characterised by the length-dependent degeneration of peripheral nerves leading to lifelong chronic disability. Currently there is no cure for this condition. CMTX3 is a rare, X-linked form of CMT. Our group identified the genetic cause of CMTX3 as a large structural variation mutation located within a non-coding region of the CMTX3 locus at chromosome Xq26.3-Xq27.3(1). Specifically, a 78 kb of region of DNA from chromosome 8q24.3 has been duplicated and inserted into a gene desert at chromosome Xq27.1. The pathomechanism of this complex DNA re-arrangement remains unsolved. We hypothesise that the CMTX3 insertion may cause neuropathy by dysregulating the expression of nearby genes. In line with this, our group previously showed that patient lymphoblasts displayed increased expression of FGF13; a gene located ~1.2 Mb away from the pathogenic insertion site(1). However, given the spatiotemporal specificity of transcriptional regulation, assessing gene expression within disease-relevant tissue is essential. Therefore, we reprogrammed CMTX3 patient fibroblasts to induced pluripotent stem cells (iPSC), which were then differentiated into spinal motor neurons. A customised NanoString nCounter® panel was designed to assess the expression levels of candidate genes most likely to be impacted by the CMTX3 insertion, and included genes contained within the insertion sequence as well as genes located within 3 Mb on either side of the insertion breakpoint. Candidate gene expression was assessed in mature motor neurons and neuroepithelial cells. Unlike the dysregulation previously observed in CMTX3 lymphoblasts, FGF13 expression was comparable between patient and control samples. However, preliminary results show dysregulated expression of SOX3 in CMTX3 neuroepithelial cells but not mature motor neurons. Interestingly, SOX3 encodes a transcription factor implicated in neuronal development, and is the closest gene to the insertion breakpoint. This gene is therefore a high-priority candidate, and further investigations will be performed to determine if SOX3 dysregulation underlies CMTX3 neuropathy. CMTX3 is an excellent disease paradigm that can be utilised to enhance our understanding of the pathogenic consequences caused by intergenic structural variation, and more broadly, the complex process of long-range gene regulation.

  1. Brewer, M.H., Chaudhry, R., Qi, J., Kidambi, A., Drew, A.P., Menezes, M.P., Ryan, M.M., Farrar, M.A., Mowat, D., Subramanian, G.M. and Young, H.K., 2016. Whole genome sequencing identifies a 78 kb insertion from chromosome 8 as the cause of Charcot-Marie-Tooth neuropathy CMTX3. PLoS genetics, 12(7), p.e1006177.