Fig. 6.

The Vps51-containg VFT complex interacts with Tlg1. A, Tlg1 interacts with the Vps51-containing VFT complex. Spheroplasts from wild type (PSY118) cells expressing Vps52-HA and transformed with either the plasmid expressing PA-Vps51 (pCuPAYKR020(416)) or PA alone (pRS416-CuProtA) were treated with 1.5 mm of the cross-linker (dithiobis(succinimidyl propionate) (DSP), detergent solubilized, and the PA fusions were affinity purified on IgG-Sepharose as described in the legend to Fig. 5A. Samples from the extracts (Ext) or purified fractions (IP) were analyzed by Western blot using anti-HA, Tlg1, Sed5, and Pep12 antibodies or antiserum. A longer film exposure is also shown to demonstrate the total absence of cross-linking between the VFT complex and the control tSNAREs. B, the tlg2Δ strain accumulates prApe1 in YPD medium and this defect is bypassed in nitrogen starvation conditions. Wild type (WT) and tlg2Δ cells in the BY4742 background were essentially treated as in Fig. 1A. C, the tlg2Δ strain has a reversible accumulation of GFP-prApe1 in a cytosolic punctate structure. The same strains used in panel B were transformed with the plasmid carrying the GFP-prApe1 construct (pTS466) and analyzed as described in the legend to Fig. 1C. DIC, differential interference contrast.