Does CO2 mediated gating of Cx32 occur in Schwann cells?

We wish to transform how we regard CO2: to show that it can be an important messenger rather than merely a universal waste product of metabolism. While this possibility has not received much attention, it is made much more probable by our discovery that certain beta connexins (Cx26, Cx30 and Cx32) are receptors for CO2. Molecular phylogenetic analysis of the Cx26/30/32 clade shows that this motif has been evolutionarily conserved for at least 400 MYs. Single mutations of the K104 or K124 the residues key in Cx32 CO2 gating would result in CO2 insensitive hemichannels. The preservation of these residues suggests a physiological function. Cx32 is expressed in Schwann cells (SCs), the myelin forming cells of the peripheral nervous system, with specific localisation to the paranodal region and Schmidt-Lanterman incisures. Mutations in the gjb1 gene, encoding Cx32, results in a slowly progressing peripheral neuropathy called CMTX. Cx32 null mice display CMTX-like symptoms, such as a reduced grip strength, with rescue of symptoms upon selective re-expression of SC Cx32. CO2 gating of Cx32 is made all the more interesting with our finding that some such CMXT mutants abolish CO2 sensitivity of Cx32 hemichannels, seen via Ca2+ imaging and dye-loading.
Circumstantial evidence suggests that CO2 signalling through Cx32 in myelin may act to maintain myelin integrity by finetuning internodal distance and myelin thickness via purinergic signalling. This project aims to utilise a genetic tool, dnCx32, which assembles with WT to remove CO2 sensitivity in vivo to definitively show CO2 is the physiological gate of Cx32 in myelin. Furthermore, genetically encoded sensors, GCaMP and GRABATP1.0, will be used to study Ca2+ signals and ATP release from peripheral nerve.