SCD5, A SUPPRESSOR OF CLATHRIN DEFICIENCY, ENCODES A NOVEL PROTEIN WITH A LATE SECRETORY FUNCTION IN YEAST

Clathrin and its associated proteins constitute a major class of coat proteins involved in vesicle budding during membrane transport. An int eresting characteristic of the yeast clathrin heavy chain gene (CHC1) is that in some strains a CHC1 deletion is lethal, while in others it is not. Recently, our laboratory developed a screen that identified fi ve multicopy suppressors that can rescue lethal strains of clathrin he avy chain-deficient yeast (Chc(-) scd1-i) to viability. One of these s uppressors, SCD5, encodes a novel protein of 872 amino acids containin g two regions of repeated motifs of unknown function. Deletion of SCD5 has shown that it is essential for cell growth at 30 degrees C. scd5- Delta strains carrying low copy plasmids encoding C-terminal truncatio ns of Scd5p are temperature sensitive for growth at 37 degrees C. At t he nonpermissive temperature, cells expressing a 338-amino acid deleti on (Scd5p-Delta 338) accumulate an internal pool of fully glycosylated invertase and mature alpha-factor, while processing and sorting of th e vacuolar hydrolase carboxypeptidase Y is normal. The truncation muta nt also accumulates 80- to 100-nm vesicles similar to many late sec mu tants. Moreover, at 34 degrees C, overexpression of Scd5p suppresses t he temperature sensitivity of a sec2 mutant, which is blocked at a pos t-Golgi step of the secretory pathway. Biochemical analyses indicate t hat similar to 50% of Scd5p sediments with a 100,000 X g membrane frac tion and is associated as a peripheral membrane protein. Overall, thes e results indicate that Scd5p is involved in vesicular transport at a late stage of the secretory pathway. Furthermore, this suggests that t he lethality of clathrin-deficient yeast can be rescued by modulation of vesicular transport at this late secretory step.