TY - JOUR
T1 - Genetic Analysis of Yeast Sec24p Mutants Suggests Cargo Binding Is Not Co-operative during ER Export
AU - Buchanan, Roy
AU - Kaufman, Andrew
AU - Kung-Tran, Leslie
AU - Miller, E.A.
PY - 2010/8
Y1 - 2010/8
N2 - Many eukaryotic secretory proteins are selected forexport from the endoplasmic reticulum (ER) through theirinteraction with the Sec24p subunit of the coat protein II(COPII) coat. Three distinct cargo-binding sites on yeastSec24p have been described by biochemical, genetic andstructural studies. Each site recognizes a limited set ofpeptide motifs or a folded structural domain, however,the breadth of cargo recognized by a given site and thedynamics of cargo engagement remain poorly under-stood. We aimed to gain further insight into the broadermolecular function of one of these cargo-binding sitesusing a non-biased genetic approach. We exploited thein vivolethality associated with mutation of the Sec24pB-site to identify genes that suppress this phenotypewhen overexpressed. We identifiedSMY2as a gen-eral suppressor that rescued multiple defects in Sec24p,andSEC22as a specific suppressor of two adjacentcargo-binding sites, raising the possibility of allostericregulation of these domains. We generated a novel setof mutations in Sec24p thatdistinguish these two sitesand examined the ability of Sec22p to rescue these muta-tions. Our findings suggest that co-operativity does notinfluence cargo capture at these sites, and that Sec22prescue occurs via its function as a retrograde SNARE.
AB - Many eukaryotic secretory proteins are selected forexport from the endoplasmic reticulum (ER) through theirinteraction with the Sec24p subunit of the coat protein II(COPII) coat. Three distinct cargo-binding sites on yeastSec24p have been described by biochemical, genetic andstructural studies. Each site recognizes a limited set ofpeptide motifs or a folded structural domain, however,the breadth of cargo recognized by a given site and thedynamics of cargo engagement remain poorly under-stood. We aimed to gain further insight into the broadermolecular function of one of these cargo-binding sitesusing a non-biased genetic approach. We exploited thein vivolethality associated with mutation of the Sec24pB-site to identify genes that suppress this phenotypewhen overexpressed. We identifiedSMY2as a gen-eral suppressor that rescued multiple defects in Sec24p,andSEC22as a specific suppressor of two adjacentcargo-binding sites, raising the possibility of allostericregulation of these domains. We generated a novel setof mutations in Sec24p thatdistinguish these two sitesand examined the ability of Sec22p to rescue these muta-tions. Our findings suggest that co-operativity does notinfluence cargo capture at these sites, and that Sec22prescue occurs via its function as a retrograde SNARE.
KW - cargo selection
KW - COPII vesicles
KW - ER export
KW - intracellular traffic
KW - Sec24
UR - http://www.scopus.com/inward/record.url?eid=2-s2.0-77958128191&partnerID=MN8TOARS
U2 - 10.1111/j.1600-0854.2010.01080.x
DO - 10.1111/j.1600-0854.2010.01080.x
M3 - Article
SN - 1398-9219
VL - 11
SP - 1034
EP - 1043
JO - Traffic
JF - Traffic
IS - 8
ER -