| Through biochemical and genetic approaches, a conceptual framework for secretion is emerging, although a complete molecular description is still lacking. It is clear that the formation of the SNARE complex lies at the heart of the molecular events that govern transport through the secretory pathway. However, a significant body of evidence suggests that SNARES aren't solely responsible for the selective targeting and docking required to maintain organelle composition, suggesting other factors contribute to the process. Indeed, several proteins that regulate SNARE function have been described, including Sly1p, a t-SNARE-interacting protein that functions in ER to Golgi traffic, as well as a family of so called tethering factors.; We have identified DSL1 through its genetic interaction with SLY1. Like SLY1, conditional dsl1 mutants exhibit a block in ER to Golgi traffic at the restrictive temperature. dsl1 mutants are defective for retrograde Golgi to ER traffic, even under conditions where no anterograde transport block is evident. This suggests that the primary function of Dsl1p may be in retrograde traffic, and is consistent with the concept that retrograde defects can lead to secondary anterograde traffic defects.; Dsl1p is an ER-localized peripheral membrane protein that resides in a multiprotein complex. Immunoisolation of the complex yielded Dsl1p and proteins of ∼80 kD and ∼55 kD. The ∼80 kDa protein is Tip20p, a protein that exists in a tight complex with ∼50 kD Sec20p. Sec20p and Tip20p function in retrograde Golgi to ER traffic, are ER-localized, and bind to the ER t-SNARE Ufe1p. Interestingly, we also found that Dsl1p interacts with the retrograde COPI coat δ-subunit, Ret2p. This novel finding represents the first known interaction between tethering components and COPI subunits. Another component, DSL3, was identified in a two-hybrid protein interaction screen with Dsl1p. Dsl3p is an ∼81 kD ER-localized peripheral membrane protein. Size exclusion chromatography and co-immunoprecipitation experiments show Dsl3p may be associated with the Dsl1p complex. Based on these findings, I present a model in which an ER-localized complex of Dsl1p, Dsl3p, Sec20p and Tip20p functions in retrograde traffic, perhaps upstream of a Sly1p/Ufe1p complex, either in the tethering or uncoating of COPI vesicles. |
