Fig. 7. Pex19p functions as a chaperone and transports PMPs in a temporally differentiated manner in peroxisome biogenesis. (A) Flag-tagged PMPs including Pex14p (FL14, a-h), Pex16p (FL16, i-p), and Pex26p (FL26, q-x) were expressed (two left panels each) or co-expressed with HA₂-Pex19p (middle and right panels) in pex19 ZP119 cells. Cells were stained with antibodies to Flag and HA and with MitoTracker, at the times indicated at the top after the transfection. ZP 119 cells co-transfected with Flag-PEX26 and HA₂-PEX19 were also stained with antibodies to Pex14p and HA at 36 hours post transfection (y,z). (B) ZP119 cells were likewise transfected with HA₂-PEX19 and were assessed for endogenous Pex14p by immunostaining with specific antibody, mitochondria by MitoTracker and Pex19p by HA staining. Note that Pex14p and HA₂-Pex19p were detectable in the cytosol at 12 hours post transfection. (C) ZP119 cells expressing Pex3p-EGFP alone (3EGFP, left panels) or with HA₂-Pex19p (right) were verified at 12 hours post transfection for Pex3p by EGFP, mitochondria and HA₂-Pex19p. Note that Pex3p-EGFP and HA₂-Pex19p were detectable in numerous particles, apparently mitochondria. Bar, 20 µm.