The calcium-binding proteins MRP8 and MRP14 present in mature monomyelocytic cells are induced during differentiation and may mediate the growth arrest in differentiating HL-60 cells. We determined the levels of a protein complex (PC) containing MRP8 and MRP14 and investigated the mechanism by which the genes encoding these proteins are regulated in HL-60 cells treated with the differentiation-inducing agent mycophenolic acid (MPA). While the PC was barely detectable in untreated cells, MPA treatment resulted in elevated levels of the PC which were maximal at 3-4 days and were found to directly parallel gains in the steady-state levels of MRP8 and MRP14 mRNA. Transcription studies with the use of nuclear run-on experiments revealed increased transcription initiation at the MRP8 and MRP14 promoters after MPA treatment. 1 alpha,25-Dihydroxyvitamin D3, which induces HL-60 cell differentiation by another mechanism, was also found to increase transcription initiation at the MRP8 and MRP14 promoters. Our results suggest that this initiation is the major control of maturation agent-mediated increases in MRP8 and MRP14 gene expression, and supports a role for the PC in terminal differentiation of human monomyelocytic cells.