Down-regulation of P-2U-purinergic nucleotide receptor messenger RNA expression during in vitro differentiation of human myeloid leukocytes by phorbol esters or inflammatory activators

HL-60 human promyelocytic leukocytes express G protein-coupled P-2U-purinergic nucleotide receptors (P(2U)R or P2Y(2)R) that activate inositol phospholipid hydrolysis and Ca2+ mobilization in response to ATP or UTP. We examined the expression of functional P(2U)R and P(2U)R mRNA levels during in vitro differentiation of HL-60 cells by dibutyryl-cAMP (Bt(2)cAMP), which induces a granulocyte/neutrophil phenotype, or by phorbol-12-myristate-13-acetate (PMA), which induces a monocyte/macrophage phenotype. Both P(2U)R function and P(2U)R mRNA levels were only modestly attenuated during granulocytic differentiation by Bt(2)cAMP. In contrast, P(2U)R function, as assayed by either Ca2+ mobilization or inositol trisphosphate generation, was greatly reduced in PMA-differentiated cells. This inhibition of P(2U)R function was strongly correlated with PMA-induced decreases in P(2U)R 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(2U)R mRNA, this was rapidly followed by a sustained decrease in P(2U)R mRNA to a level 5-10-fold lower than that in undifferentiated HL-60 cells. The half-life of the P(2U)R transcript in HL-60 cells was similar to 60 min, and this was not affected by acute exposure (less than or equal to 4 hr) to Bt(2)cAMP or PMA. PMA down-regulated P(2U)R mRNA in THP-1 monocytes and HL-60 granulocytes but not in A431 human epithelial cells or human keratinocytes. P(2U)R mRNA was also down-regulated in THP-1 monocytes differentiated into inflammatory macrophages by gamma-interferon and endotoxin. These data indicate that myeloid leukocytes possess tissue-specific mechanisms for the rapid modulation of P(2U)R expression and function during differentiation and inflammatory activation.