Poster: Organelle biogenesis

Abs # 638: ER is the primary sorting compartment for the post-translational trafficking of AtPex16p to peroxisomes in suspension-cultured cells.

Presenter:	Karnik, Sheetal K, skarnik@asu.edu
Authors	Karnik, Sheetal K (A)   Trelease, Richard N (A)
Affiliations:	(A): Arizona State University

The putative AtPex16 homolog, a 42 kD polypeptide with two predicted internal transmembrane domains, was reported to have a role in ER-dependent protein and oil body biogenesis; however, a role in plant peroxisomal biogenesis could not be determined (Lin et al., 1999, Science 284:328). Our study was aimed at determining whether AtPex16p was involved in ER-related peroxisomal biogenesis. Anti-AtPex16p IgGs recognized in microsomes two polypeptides: 42 kD (expected) and 51 kD (unexpected). Immunofluorescence microscopy revealed the localization of AtPex16p in peroxisomes and also dispersed throughout the ER. To learn whether AtPex16p moves from ER to peroxisomes, cells were held for 5 h at 15º C (slows protein export from ER). AtPex16p accumulated in distinct circular structures reminiscent of ER exit sites and then appeared in peroxisomes and ER upon 5 h recovery at 25º C. Similar cold and BFA treatments of cells transiently expressing mycAtPex16p revealed more compelling ER-to-peroxisome movements. In-vitro cell fractionation experiments confirmed the presence of both the 42 and the 51 kD polypeptides in ER (Mg+2 shifted) and peroxisomes (isopycnic sucrose gradients). Surprisingly, AtPex16p (42 kD) exhibited characteristics of a peripheral membrane protein in ER and peroxisomes. Interestingly, the topological orientations of AtPex16p in ER and peroxisomes differed, i.e., AtPex16p faced the cytosolic side of ER membranes and the matrix side of peroxisomal membranes. Together, the data support our premise that AtPex16p is an authentic peroxin homolog that sorts post-translationally via two non-overlapping internal targeting signals to ER and then to peroxisomes, and likely participates in an ER-dependent biogenesis of peroxisomes. Supported by NSF grant MCB-0091826