Figure 1. Glycolytic enzymes form G bodies in response to hypoxia.

(A) Glycolytic enzymes form granules upon hypoxia treatment. Yeast cells with the indicated GFP-tagged proteins were cultured with oxygen (+) or in a hypoxic chamber (−) for 24 h. (B) Pfk2p does not form granules in glucose (-Glu) or nitrogen (-N) starved yeast or in yeast grown on a non-fermentable carbon source (EtOH+Gly). (C) Pfk2p colocalizes with Pfk1p, Fba1p, and Cdc19p in yeast grown in hypoxia for 24 h. GFP, green; Azurite, magenta. (D) Pfk2p-GFP forms non-membrane-bound structures after 24 h hypoxic treatment as measured by immunogold (anti-GFP) staining and TEM. (E) Yeast strains co-expressing GFP-tagged Pab1p or Edc3p with Pfk2p-Azurite were subjected to glucose or oxygen deprivation, respectively. Cells were cultured in SMD (5%) to the mid-log phase, and shifted to SM medium for 30 min to induce stress granule formation, or cultured in the hypoxic chamber for 24 h to induce Pfk2 granule formation. Images are representative fluorescence images taken with FITC (GFP) and DAPI (Azurite) channels. (F) Pfk2p granule formation adjacent to a P-body (Edc3p-GFP) was observed occasionally in -O₂ treated cells in (E). Scale bar 5 μm (A–E). [See also Figures S1–S2.]