dcyphr | COPII: A Membrane Coat Formed by Sec Proteins That Drive Vesicle Budding from the Endoplasmic Reticulum.


The three proteins, Sar1p, Sec13p complex, and Sec23p complex, contribute to the synthesis of transport vesicles. The vesicles derive from the endoplasmic reticulum. Vesicle formation requires GTP. But, GMP-PNP, a nonhydrolyzable analog of GTP, can cause vesicle formation as well. But, GMP-PNP vesicles cannot target and fuse with the Golgi complex. All three proteins remain on the vesicles. Yet, GTP vesicles are functional. Sar1p dissociates from GTP vesicles, allowing for fusion. Thin section electron microscopy reveal a thick coat around the vesicles. The subunits of the novel coat are different from clathrin, another coat protein. Two proteins drive the budding cycle in similar ways. So, Barlowe et al. proposes to call these two coat structures COPI and COPII.


In intercompartmental transport in eukaryotes, vesicles uncoat before fusion with the acceptor compartment. For example, clathrin mediates transport involving the trans-Golgi and plasma membrane. COP-coated Golgi-derived vesicles mediate intra-Golgi transport.

The three different proteins have different functions. Sar1p is a GTPase functioning on the cytosolic face of the ER. Sec12p, an integral membrane glycoprotein, activates Sar1p by a GDP/GTP exchange. The Sec23p complex consists of two proteins: Sec23p and Sec24p. Sec23p is a Sar1p-specific GTPase activating protein. Sec24p aids secretion of vesicles. The Sec13p complex consists of  Sec13p and another 150kDa polypeptide. Vesicle formation requires both proteins in vitro. These three proteins are components of the vesicle coat and aid in vesicle synthesis. A distinct coat structure appears to be mediating each intracellular vesicle budding. These vesicles contain membrane-bound proteins that are characteristic of uncoated ER-derived transport vesicles.


Barlowe et al. tries to understand the roles COPII and COPI play in vesicle budding.


Researchers produced a recombinant Sar1p with restriction sites and a plasmid. They used yeast and E. coli. To study budding and vesicle formation, they did an analytical budding assay. They measured the amount of protease protected to determine vesicle release. Then, they did a vesicle chase to track vesicle transport. Proteins and vesicles were viewed with immunoblots and electron microscopy, respectively.

Results / Conclusion

The budding process if as follows (Figure 9). Sar1p goes to the ER. Sec12p converts Sar1p-GDP to the activated GTP-bound form. This causes Sar1p to be membrane-bound. GTP is the best nucleotide for budding. Then, Sar1p-GTP recruits Sec23p and Sec13p complexes to form COPII. COPII executes the budding event. Sar1p hydrolyzes GTP, stimulated by the Sec23p subunit of the Sec23p complex. GTP hydrolysis leads to the loss of Sar1p from vesicles, causing coat instability. The Sar1p loss exposes targeting proteins on the vesicle, such as Sec22p and Bos1p. These proteins provide binding selectivity to the target proteins on the Golgi membrane. If vesicles form with a nonhydrolyzable analog like GMP-PNP, Sar1p and the COPII proteins remain on the vesicle (Figure 8). The remaining Sar1p and COPII proteins hinder access of the vesicle to the Golgi. The vesicle will not be able to fuse with the acceptor membrane.

COPII formation resembles the assembly of COPI. But, COPI and COPII are distinct. Unlike COPI, COPII contains a GTPase activating protein (GAP) subunit. For COPII, retention of the Sec23p and Sec13p complexes do not depend on the presence of Sar1p. In contrast, COPI-coated vesicles keep ADP ribosylation factor (ARF) when the vesicles form with GTP or with GTPγS. ARF-GAP may stimulate GTP hydrolysis and COPI disassembly. Palmityl coenzyme A aids COPI assembly in incubation. Formation of COPII-coated vesicles do not need acyl coenzyme A.

COPII vesicles have specific protein packages. The proteins selected have more affinity for COPII. Proteins that function in the ER are not included in these vesicles. For example, Sec12p is not on vesicles. Sec12p has no affinity for COPII, so it will remain in the ER. Sec12p helps later rounds of budding. GTP hydrolysis is not required for protein sorting. Rather, GTP hydrolysis is important for the fusion of the vesicle. The depletion of coatomer does not affect ER budding.

Vesicles formed with crude cytosolic proteins contain Bos1p, Sec22p, and Ypt1p. The researchers find Sec22p in vesicles formed with pure cytosolic proteins. But, they cannot detect Ypt1p in these vesicles. Ypt1p may be normally recruited from the cytosol to form a bud. Vesicles that do not have Ypt1p may get it on the way to targeting the Golgi complex. Vesicles formed with pure proteins need Ypt1p to target and fuse with the Golgi. 

Vesicle targeting requires Bet1p. But, Bet1p is not recovered in vesicles formed with crude cytosol. Rather, Bet1p is packaged along with α-factor precursor and  Sec22p in these vesicles. This may be due to the presence or absence of Ypt1p. Ypt1p may regulate vesicle packaging or retention of certain targeting or cargo molecules.

COPI vesicles seem to exclude Golgi membrane proteins while incorporating relevant cargo molecules. COPII- and COPI-mediated budding events likely differ in cargo enrichment during transport. Viral glycoproteins become concentrated about 10-fold after exiting the ER. But, they are not further concentrated once they are in the Golgi. The COPII budding process likely includes a mechanism for cargo concentration.

COPI is essential for protein transport from the ER and within the Golgi complex. SEC21 codes for the γ subunit of yeast coatomer. The sec21 mutant blocks transport between the ER and Golgi. Polyclonal antibodies and Fab fragments against mammalian β-COP inhibit transport of vesicular stomatitis virus G protein. Addition of COPI and ARF have no effects on vesicle formation mediated by COPII.

Various transport reactions need COPI to differing extents. Sometimes, COPII may be sufficient for the complete budding process. Other times, COPI and COPII may cohabit the same vesicle or create different vesicles. COPI may replace COPII on ER-derived vesicles. In this case, COPI may serve as a scaffold for the attachment of Ypt1p. COPI  may regulate the targeting of a vesicle to the correct compartment. COPI may also affect the budding of a vesicle.