Figure 5.

Orientation and rescue efficiency of N-myc- and C-myc-tagged tail constructs. A, Bars schematically illustrate the tail variants myc–α1–iD–TM4 and α1–iD–TM4-myc, showing the location of the myc epitope (hatched box) and TM4 (black box). Transfected COS7 cells were used either unpermeabilized or permeabilized with 0.1% Triton X-100. Bottom pictures represent a live stain experiment. Live staining was performed using a 1 h incubation step at 4°C before fixation. Untransfected cells and GFP-transfected cells were used as controls (data not shown). Scale bars, 20 μm. B, The myc epitope was cut for determination of rescue efficiency by the tail sequence (α1–iD–TM4). Pc–α1–iD–TM4 is modified by harboring the basic stretch RRKRRH attached to the N terminus of the tail construct (gray box). Constructs lacking the myc epitope (α1–iD–TM4) were most efficient in rescue of the appropriate trc construct independent if they were of α1 or spd^ot-origin. C, Bar diagram of the calculated maximal current amplitudes. Note that, in spd^ot-trc + tail coexpressions, the maximal current amplitudes were doubled compared with spd^ot-trc + myc–α1–iD–TM4. *p < 0.01, ***p < 0.0001.