A role for Tlg1p in the transport of proteins within the Golgi apparatus of Saccharomyces cerevisiae

Members of the syntaxin protein family participate in the docking-fusion st ep of several intracellular vesicular transport events. Tlg1p has been iden tified as a nonessential protein required for efficient endocytosis as well as the maintenance of normal levels of trans-Golgi network proteins. In th is study we independently describe Tlg1p as an essential protein required f or cell viability. Depletion of Tlg1p in vivo causes a defect in the transp ort of the vacuolar protein carboxypeptidase Y through the early Golgi. Tem perature-sensitive (ts) mutants of Tlg1p also accumulate the endoplasmic re ticulum/ cis-Golgi form of carboxypeptidase Y at the nonpermissive temperat ure (38 degrees C) and exhibit underglycosylation of secreted invertase. Ov erexpression of Tlg1p complements the growth defect of vti1-11 at the nonpe rmissive temperature, whereas incomplete complementation was observed with vti1-1, further suggesting a role for Tlg1p in the Golgi apparatus. Overexp ression of Sed5p decreases the viability of tlg1 ts mutants compared with w ild-type cells, suggesting that tlg1 ts mutants are more susceptible to ele vated levels of Sed5p. Tlg1p is able to bind His(6)-tagged Sec17p (yeast al pha-SNAP) in a dose-dependent manner and enters into a SNARE complex with V tilp, TIg2p, and Vps45p. Morphological analyses by electron microscopy reve al that cells depleted of Tlg1p or tlg1 ts mutants incubated at the restric tive temperature accumulate 40- to 50-nm vesicles and experience fragmentat ion of the vacuole.