Fig. 4.

SEA complex is involved in the regulation of the TORC1 pathway. A, the yeast SEA complex physically interacts with the TORC1 complex. Sea2-PrA was immunoprecipitated as described under “Experimental Procedures.” Co-precipitating proteins were resolved via SDS-PAGE, visualized with Coomassie Blue stain, and identified via mass spectrometry (supplemental Tables S2 and S3). SEA complex members are marked in blue, TORC1 members in orange, mitochondria proteins in green, proteins involved in ribosome biogenesis and translation in blue, contaminants in gray, and others in pink. B, deletions of Sea1, Npr2, and Npr3 provoked Tor1 relocalization to the cytoplasm during nitrogen starvation. The localization of Tor1-GFP was followed by light fluorescence microscopy in wild-type and deletion strains of indicated SEA complex members, either in synthetic complete (SC) medium or in synthetic media lacking nitrogen SD-N, (nitrogen starvation). Observations were made for the strains grown in YPD or subjected to nitrogen starvation. C, the “vacuole-to-cytoplasm” GFP signal ratio was calculated for 25 cells in each strain shown in B.