GRANULOCYTE-COLONY-STIMULATING FACTOR UP-REGULATES THE VACUOLAR PROTON ATPASE IN HUMAN NEUTROPHILS

We have previously shown that granulocyte colony-stimulating factor (G -CSF) delays spontaneous neutrophil apoptosis through activation of th e vacuolar proton ATPase (v-ATPase). We have now examined the regulati on of the v-ATPase in neutrophils exposed to G-CSF in vitro. When neut rophils were cultivated in the absence of G-CSF, the 57-kD cytosolic B subunit of the v-ATPase disappeared within 1 to 2 hours, its loss pre ceding the nuclear changes of apoptosis and coinciding with the onset of acidification. By contrast, in neutrophils cultured for 2 hours in the presence of G-CSF, the amount of the 57-kD subunit was similar to that in freshly isolated neutrophils. However, inhibition of protein s ynthesis with cycloheximide and actinomycin D led to loss of the 57-kD subunit even in the presence of G-CSF. These results indicated that o ngoing protein synthesis was required to maintain the v-ATPase, and fu rther suggested that G-CSF acted, at least in part, by maintaining syn thesis of the 57-kD cytosolic subunit. G-CSF also promoted the translo cation of the 57- and 33-kD cytosolic v-ATPase subunits to the membran e. Our findings suggested two coordinate mechanisms by which the activ ity of the v-ATPase could be increased by G-CSF: the synthesis of cyto solic v-ATPase subunits and their translocation to the membrane. (C) 1 997 by The American Society of Hematology.