HEAT repeats mediate plasma membrane localization of Tor2p in yeast.
The subcellular distribution of Tor1p and Tor2p, two phosphatidylinositol kinase homologs and targets of the immunosuppressive drug rapamycin in Saccharomyces cerevisiae, was analyzed. We found that Tor protein is peripherally associated with membranes. Subcellular fractionation and immunofluorescence studies showed that Tor1p and Tor2p associate with the plasma membrane and a second fraction that is distinct from Golgi, vacuoles, mitochondria, and nucleus and may represent vesicular structures. Pulse-chase experiments showed that association of Tor protein with plasma membrane and the second compartment is fast, does not appear to involve components of endocytic, secretory, or Golgi to vacuole transport pathways, and is not affected by the immunosuppressive drug rapamycin. Deletion analysis reveals that two domains within Tor2p independently mediate localization to both compartments. These domains are composed of HEAT repeats that are thought to act as protein-protein interaction surfaces. Our studies therefore place Tor proteins at the site of action of their known downstream effectors and suggest that they may be part of a multiprotein complex.
Kunz
J
J
Department of Biochemistry, Biozentrum, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
Schneider
U
U
Howald
I
I
Schmidt
A
A
Hall
M N
MN
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D018797:Cell Cycle Proteins; D002462:Cell Membrane / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D011485:Protein Binding; D011817:Rabbits; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D020123:Sirolimus / Q000378:metabolism; D015003:Yeasts
D018797:Cell Cycle Proteins; D005656:Fungal Proteins; D008565:Membrane Proteins; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D029701:Saccharomyces cerevisiae Proteins; D020123:Sirolimus; C083324:TOR1 protein, S cerevisiae; C081135:TOR2 protein, S cerevisiae