FIGURE 3.

PDI is required for catalytically active collagen-tailed AChE form expression. A, primary QMCs were transfected with pSilencer qPDI shRNA (open diamonds) or qColQ shRNA (open circles). The AChE forms were fractionated by velocity sedimentation in 5–20% sucrose gradients. G1, G2, and G4 refer to AChE monomers, dimers, and tetramers, and A4, A8, and A12 refer to the various collagen-tailed forms consisting of one, two, or three tetramers attached to the ColQ subunit, respectively. EGFP shRNA (solid circles) was used as negative control. Expression of the synaptic ColQ-AChE and the collagen-tailed A4 forms was decreased in muscle cells when either PDI or ColQ was knocked down specifically by the shRNAs. Panel B, relative PDI content was determined by a Western blot of triplicate QMC crude extracts from cells transfected with either EGFP shRNA (control) or PDI shRNA. Overexpression of PDI shRNA knocks down PDI in QMCs.