Gfi1 protein levels are high in monocytes compared with granulocytes due to decreased proteasomal degradation. (A) HL60 cells were differentiated with ATRA toward granulocytes for 72 hours. PMA was used to stimulate HL60 cells for monocytic differentiation. Cell lysates were used in an in vitro degradation assay. (B) Lysates of primary monocytes and granulocytes were used in an in vitro degradation assay with ³⁵ S-labeled Gfi1. (C) The Gfi1 turnover in granulocyte lysate in an in vitro degradation assay depended on 26S proteasome activity. (D) Quantitative Gfi1 RT-PCR was performed on RNA from primary monocytes and granulocytes (n = 3). Error bars indicate standard deviation. (E) Cell lysates of isolated monocytes and granulocytes were immunoblotted and stained with α-Gfi1 antibody, and α-actin staining was used to check for equal loading. α-C/EBPα and α-C/EBPϵ protein levels were stained as positive control for the used protein lysates. Cell lysates were normalized using the Bio-Rad protein quantification assay.