Figure S6.

The Ssh4 transmembrane domain is important in Wsc1*-GFP degradation. Related to Fig. 4. (A) Flow cytometry heat map showing the degradation defect on Wsc1-EQSPLL-GFP (hereafter, Wsc1*-GFP) imparted by Ssh4 transmembrane domain mutants (cutoff = 50%). Degradation-blocking mutants are noted as "Hits." Ssh4 mutants that had low expression (low exp) levels or were lumenal are also noted. (B) Left: Western blot showing degradation of Wsc1*-GFP in the presence of single-residue and double-residue Ssh4 mutants. Right: Quantification (±SD, n = 3). (C and D) Protein levels of NeonGreen-3HA-tagged single-residue and double-residue Ssh4 mutants. (E and F) Subcellular localization of NeonGreen-3HA-tagged single-residue and double-residue Ssh4 mutants. Scale bar, 2 µm. (G) Helical wheel showing the position of residues conferring partial degradation block when mutated to Trp. (H) Summary of Ssh4 TM residues that reduce/block Wsc1*-GFP or Ypq1-GFP degradation when mutated to Trp.