Antiapoptotic action of 1,25-dihydroxyvitamin D-3 is associated with increased mitochondrial MCL-1 and RAF-1 proteins and reduced release of cytochrome C

Release of mitochondrial cytochrome c has been recently linked to the activation of the ''executioner'' phase of the cellular programs for death by apoptosis. This release is known to be negatively regulated by Bcl-2 and Bcl-X-L proteins. We show here that treatment of human leukemia cells HL60 with 1,25-dihydroxyvitamin D-3 (1,25D(3)) results in progressive increases in the levels of cellular antiapoptotic protein Mcl-1, a transient increase in AZ protein level, but no increases in Bcl-2 or Bcl-X-L proteins, The increase in Mcl-1 protein levels correlates with a reduced extent of apoptotic cell death induced by etoposide or the calcium ionophore A23187. The Mcl-1 protein is primarily localized in the mitochondria, and etoposide-or A23187-induced cytochrome c release is reduced in cells in which the mitochondria contain the Mcl-1 protein demonstrable by immunoblots. Raf-l protein can also be detected in the mitochondrial fractions that contain Mcl-1 protein but not in the Mcl-1-negative fractions. These findings suggest that in these promyelocytic leukemia cells Mcl-1 has a function analogous to that of Bcl-2 in other cells, i.e., to target Raf-1 to mitochondria and to reduce cell damage-induced release of mitochondrial cytochrome c. Our findings provide a potential mechanism for the antiapoptotic action of 1,25D, and show that differentiation and apoptosis signaling pathways not only interact but involve a proliferation-associated gene, Raf-1. (C) 1997 Academic Press.