- Traffic. 2007 Nov 27 [Epub ahead of print]
Smy2p Participates in COPII Vesicle Formation Through the Interaction with Sec23p/Sec24p Subcomplex.
Molecular Membrane Biology Laboratory, RIKEN Discovery Research Institute, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
The coat protein complex II (COPII) is essential for vesicle formation from the endoplasmic reticulum (ER) and is composed of two heterodimeric subcomplexes, Sec23p/Sec24p and Sec13p/Sec31p, and the small guanosine triphosphatase Sar1p. In an effort to identify novel factors that may participate in COPII vesicle formation, we isolated SMY2, a yeast gene encoding a protein of unknown function, as a multicopy suppressor of the temperature-sensitive sec24-20 mutant. We found that even a low-copy expression of SMY2 was sufficient for the suppression of the sec24-20 phenotypes, and the chromosomal deletion of SMY2 led to a severe growth defect in the sec24-20 background. In addition, SMY2 exhibited genetic interactions with several other genes involved in the ER-to-Golgi transport. Subcellular fractionation analysis showed that Smy2p was a peripheral membrane protein fractionating together with COPII components. However, Smy2p was not loaded onto COPII vesicles generated in vitro. Interestingly, coimmunoprecipitation between Smy2p and the Sec23p/Sec24p subcomplex was specifically observed in sec23-1 and sec24-20 backgrounds, suggesting that this interaction was a prerequisite for the suppression of the sec24-20 phenotypes by overexpression of SMY2. We propose that Smy2p is located on the surface of the ER and facilitates COPII vesicle formation through the interaction with Sec23p/Sec24p subcomplex.
PMID: 17973654 [PubMed - as supplied by publisher]
peripheral membrane protein, 100% in pellet of 100k rcf (no cytosolic pool)
790aa, 87kDa predicted, 100kDa apparent (hi pI of 8.97)
GYF domain (with C-term req'd for suppression), coiled-coil domain
ER - sucrose gradient, IF microscopy (perinuclear ER)
suppressor of ts sec24-20, alsosec16-2, sec22-3, bet1-1, sec34-1, sec35-1
deletion synthetically lethal with sec24-20
co-IPs sec23/24 only in sec24-20, sec23-1, requires coiled coil +carboxy terminal portion
yeast 2-hybrid interaction with Sec23p, but not Sec24p (also Msl5p, Mud2p, mRNA splicing)
Smy2p required to survive sec24-20 defect, scaffold protein?
Background:
sec24p A-site - SNARE Sed5p
B-site - SNAREs Sed5p, Bet1p, Golgi protein Sys1p
C-site - SNARE Sec22p
sec24-20 lacks c-term 30 aa (W897stop) in A site
Sec24p homologues Sfb2p(Iss1p), Sfb3p (Lst1p)
Sfb3p specialized for Pma1p packaging
Sfb2p functionally redundant for sec24p
