Fig. 1. GNL3L interacts with ERR, ERR and ERR. Protein interactions between ERR and nucleostemin family genes (A) and between GNL3L and ERR family genes (B) were examined by in vivo coimmunoprecipitation assays. HEK293 cells were co-transfected with (A) Myc-tagged ERR and HA-tagged nucleostemin, GNL3L or Ngp1 expression plasmids, or (B) HA-tagged GNL3L and Myc-tagged ERR, or expression plasmids. Lysates were immunoprecipitated (IP) with anti-Myc antibody (rows 1 and 2, -Myc) or anti-HA antibody (rows 3 and 4, -HA). The co-purified proteins (rows 1 and 3) and the immunoprecipitates (rows 2 and 4) were immunodetected with the antibodies indicated on the right. Our results show that ERR interacts only with GNL3L, not with nucleostemin or Ngp1, and that GNL3L binds all ERR family proteins. (C) Tissue distribution of GNL3L, ERR and ERR in adult mice is shown by multi-tissue northern blots. GNL3L mRNA is expressed primarily in the neural tissues, including the brain and eye, and at lower levels in the kidney and muscle. The expression levels of GNL3L and ERR match in kidney, muscle and eye, but differ in brain and heart. (D) In U2OS cells, the intensity of GFP-tagged GNL3L (GNL3L-gfp) is higher in the nucleolus than in the nucleoplasm. GFP-tagged ERR (ERR-gfp) is localized exclusively in the nucleoplasm. The nucleolar regions are labeled by anti-fibrillarin (Fib) antibody in the right panels. Dashed lines demarcate nucleo-cytoplasmic boundaries. Bars, 10 µm.