Abstract

HL-60 human promyelocytic leukocytes express G protein-coupled P_2U-purinergic nucleotide receptors (P_2UR or P2Y₂R) that activate inositol phospholipid hydrolysis and Ca²⁺ mobilization in response to ATP or UTP. We examined the expression of functional P_2UR and P_2UR mRNA levels during in vitro differentiation of HL-60 cells by dibutyryl-cAMP (Bt₂cAMP), which induces a granulocyte/neutrophil phenotype, or by phorbol-12-myristate-13-acetate (PMA), which induces a monocyte/macrophage phenotype. Both P_2UR function and P_2UR mRNA levels were only modestly attenuated during granulocytic differentiation by Bt₂cAMP. In contrast, P_2UR function, as assayed by either Ca²⁺ mobilization or inositol trisphosphate generation, was greatly reduced in PMA-differentiated cells. This inhibition of P_2UR function was strongly correlated with PMA-induced decreases in P_2UR mRNA levels, as assayed by Northern blot analysis or reverse transcription-polymerase chain reaction-based quantification. Although PMA induced an early, transient up-regulation of P_2UR mRNA, this was rapidly followed by a sustained decrease in P_2UR mRNA to a level 5–10-fold lower than that in undifferentiated HL-60 cells. The half-life of the P_2UR transcript in HL-60 cells was ∼60 min, and this was not affected by acute exposure (≤4 hr) to Bt₂cAMP or PMA. PMA down-regulated P_2UR mRNA in THP-1 monocytes and HL-60 granulocytes but not in A431 human epithelial cells or human keratinocytes. P_2UR mRNA was also down-regulated in THP-1 monocytes differentiated into inflammatory macrophages by γ-interferon and endotoxin. These data indicate that myeloid leukocytes possess tissue-specific mechanisms for the rapid modulation of P_2UR expression and function during differentiation and inflammatory activation.