DIFFERENTIATION-STIMULATED ACTIVITY BINDS AN ETS-LIKE, ESSENTIAL REGULATORY ELEMENT IN THE HUMAN PROMYELOCYTIC DEFENSIN-1 PROMOTER

The human HNP-defensin-1 gene encodes a peptide antibiotic found exclu sively in neutrophils and is key to elimination of microbes. Expressio n is a marker for the granulocytic lineage and for certain stages of d ifferentiation and is not known to be inducible in mature cells under physiological conditions. Low level of transcription also occurs in HL -60 promyelocytic leukemia cells and is greatly activated upon drug-in duced granulocytic maturation and by low doses of retinoic acid, in a strictly cell-specific manner (Herwig, S., Su, Q., Ma, Y., and Tempst, P. (1996) Blood 87, 350-364). We have analyzed a 10-kilobase pair reg ion, upstream of the defensin-1 cap site, for the presence of control elements, and we describe a minimal promoter (position -83 to +82) req uired to drive transcription in HL-60 cells in a quasi cell-specific m anner. Our data also suggest the presence of negative regulatory eleme nts in the -416/-191 region that may further contribute to cell specif icity in a chromosomal context. The basal promoter contains two functi onally essential, ETS-like (GGAA core sequence) elements. The proximal site (-22/-19) constitutively binds the PU.1 transcription factor in vitro and could function, together perhaps with an adjacent TA-rich se quence (-32/-25), in assembly of a myeloid-restricted, basal transcrip tion factor complex. The distal site (-62/-59) interacts in vitro with an unidentified activity, distinct from PU.1, ETS-1, PEA3, and ELK-1 (factors with definite binding site similarities), and is greatly stim ulated by phosphorylation during granulocytic differentiation of HL-60 cells. Identification of this protein will be important to resolve th e molecular mechanisms controlling temporal, granulocytic restricted g ene expression.