Fig. 2.

Coexpression of nephrin with Slo1_VEDEC increases steady-state expression on the surface of HEK-293T cells. A: confocal immunofluorescence using antibodies against the Myc tags of transiently expressed Slo1_VEDEC channels. Cell surface channels in cells expressing Myc-tagged Slo1_VEDEC were labeled with an FITC-conjugated goat anti-Myc applied to intact cells (green). After fixation and permeabilization, intracellular channels were stained using a mouse anti-Myc revealed using Alexa568-conjugated anti-mouse IgG (red). Identical laser excitation intensities and detection sensitivities were used for image collection from all of these samples. B: representative cell surface biotinylation assay shows that nephrin coexpression caused marked increase in surface expression of Slo1_VEDEC but had no effect on Slo1_QEERL or Slo1_EMVYR, which had high levels of constitutive surface expression even in the absence of nephrin. C: densitometric quantification of 3 repetitions of the experiment shown in B. Data are means ± SE of relative Slo1 expression in the absence (C) or presence (N) of nephrin.