FIG. 4.

Tbμ1 silencing does not affect secretory trafficking in PCF trypanosomes. Control (tet−) and Tbμ1-silenced (tet+) PCF cell lines expressing either the soluble secretory reporter BiPN (A) or the GPI-anchored reporter VSG117 (B) were pulse-chase radiolabeled as for Fig. 3. Reporter polypeptides were specifically immunoprecipitated from cell and medium (med.) fractions at the indicated chase times and analyzed by SDS-PAGE/phosphorimaging (10⁷ cell equivalents per lane). Representative gels from control cells are presented (top). The mobilities of endogenous BiP, recombinant BiPN reporter, and the full-length (F) and truncated (T) forms of the recombinant VSG117 reporter are indicated. Cell-associated (circles) and secreted (squares) reporters for triplicate experiments were quantified by phosphorimaging and are presented as percentages of initial reporters (means ± SEM). Release of truncated VSG is due to cleavage by a resident metalloprotease following arrival at the cell surface (7).