Sec16 plays a key role in the formation of coat protein II vesicles, which mediate protein transport from the endoplasmic reticulum (ER) to the Golgi apparatus. Mammals have two Sec16 isoforms: Sec16A, which is a longer primary ortholog of yeast Sec16, and Sec16B, which is a shorter distant ortholog. Previous studies have shown that Sec16B, as well as Sec16A, defines ER exit sites, where coat protein II vesicles are formed in mammalian cells. Here, we reveal an unexpected role of Sec16B in the biogenesis of mammalian peroxisomes. When overexpressed, Sec16B was targeted to the entire ER, whereas Sec16A was mostly cytosolic. Concomitant with the overexpression of Sec16B, peroxisomal membrane biogenesis factors peroxin 3 (Pex3) and Pex16 were redistributed from peroxisomes to Sec16B-positive ER membranes. Knockdown of Sec16B but not Sec16A by RNAi affected the morphology of peroxisomes, inhibited the transport of Pex16 from the ER to peroxisomes, and suppressed expression of Pex3. These phenotypes were significantly reversed by the expression of RNAi-resistant Sec16B. Together, our results support the view that peroxisomes are formed, at least partly, from the ER and identify a factor responsible for this process.

The origin of peroxisomes has long been disputed. However, recent evidence suggests that peroxisomes can be formed de novo from the endoplasmic reticulum (ER) in yeast and higher eukaryotes. Sec16A and Sec16B, mammalian orthologs of yeast Sec16, are scaffold proteins that organize ER exit sites by interacting with COPII components. We recently demonstrated that Sec16B, but not Sec16A, regulates the transport of peroxisomal biogenesis factors from the ER to peroxisomes in mammalian cells. The C-terminal region of Sec16B, which is not conserved in Sec16A, is required for this function. The data suggest that Sec16B in ER areas other than ER exit sites plays this role. Our findings provide an unexpected connection between at least part of the COPII machinery and the formation of preperoxisomal vesicles at the ER, and offer an explanation of how secretory and peroxisomal trafficking from the ER are distinguished.