INDUCTION OF RELA(P65) AND I-KAPPA-B-ALPHA SUBUNIT EXPRESSION DURING DIFFERENTIATION OF HUMAN PERIPHERAL-BLOOD MONOCYTES TO MACROPHAGES

We evaluated the expression and DNA binding activity of nuclear factor (NF)-kappa B subunits in human peripheral blood monocytes and in mono cyte-derived macrophages (MDMs). Constitutive DNA binding activity con sisting of p50 homodimers was detected in nuclear extracts from both c ell types. An additional complex composed of p50/RelA(p65) heterodimer s appeared only in nuclear extracts from 7-day MDMs. Immunoblot analys is showed that the p50 subunit was constitutively expressed in monocyt es and MDMs. In contrast, the RelA(p65) subunit was barely detectable in monocytes, but its level increased markedly in MDMs. Analysis of Re lA(p65) mRNA revealed that the stability of RelA(p65) mRNA was signifi cantly higher in MDMs, compared with monocytes. In MDMs, an upregulati on of I kappa B alpha synthesis as well as the appearance of a novel M (r) 40,000 form of I kappa B alpha were also observed. These results s uggest that macrophage differentiation results in the expression of ac tive p50/RelA(p65) heterodimers with the capacity to activate target g ene expression. The parallel induction of I kappa B alpha synthesis ma y allow for the continuous presence of a cytoplasmic reservoir of p50/ RelA(p65) complexes that are readily available for inducer-mediated st imulation.