caSTAT5a-NLK-NARF–mediated ubiquitination of LEF-1. Interactions between LEF-1, STAT5, NARF, and NLK proteins as well as LEF-1 ubiquitination: (A) lysates of Jurkat cells transduced with caSTAT5A were immunoprecipitated with a polyclonal anti–LEF-1 antibody and the immunoprecipitates were subjected to western blot analysis with anti–LEF-1, anti-STAT5, anti-NARF, anti-NLK, or anti-ubiquitin antibodies. Representative western blot images are shown; (B) lysates of 293T HEK cells transfected with caSTAT5A were immunoprecipitated with anti-NARF antibody and the immunoprecipitates were subjected to western blot analysis with anti-NARF or anti-STAT5 antibodies; representative western blot images are shown. (C-D) Interactions between LEF-1 and STAT5, LEF-1 and NARF, LEF-1 and NLK, LEF-1 and ubiquitin, STAT5 and NLK, and STAT5 and NARF were detected in (C) Jurkat cells and (D) CD34⁺ cells treated with G-CSF using Duolink. DAPI, stained nuclei; Cy3 red dots, protein interactions.