Stable and tunable expression of human peripheral myelin protein 22 in Rat Schwann cells

Duplications and deletions of the gene encoding peripheral myelin protein 22 (PMP22) cause the most common forms of Charcot-Marie-Tooth (CMT) disease: type 1A (CMT1A) and hereditary neuropathy with liability to pressure palsies (HNPP), respectively. The resulting over- or under-expression of PMP22 in Schwann cells (SCs) causes myelin defects in the peripheral nervous system (PNS). Although the roles of PMP22 in myelin structure and maintenance have been studied extensively, the mechanisms by which perturbed PMP22 expression levels cause dysmyelination are not yet fully understood. We therefore developed a clonal rat Schwann cell (RSC) line that can express myc-tagged human PMP22 from a doxycycline-inducible genomic locus. MycPMP22 expression levels in this cell line can be tuned by adjusting the doxycycline (DOX) concentration in culture media, achieving maximal expression between 800 to 1600 ng/ml. The percentage of the total protein that reached the plasma membrane was found to inversely correlate with expression level. At high expression levels, intracellularly retained mycPMP22 localizes to puncta that can be ameliorated by serum starvation. These observations support the hypothesis that overexpression of PMP22 in CMT1A causes increased misfolding accompanied by formation of intracellular inclusions. Moreover, this work enables the precise control of mycPMP22 levels for studying CMT-related phenomena in SCs.