Genetic and physical interactions between Microphthalmia transcription factor and PU.1 are necessary for osteoclast gene expression and differentiation

The microphthalmia transcription factor (MITF), a basic-helix-loop-helix zi pper factor, regulates distinct target genes in several cell types. We hypo thesized that interaction with the Ets family factor PU.1, whose expression is limited to hematopoietic cells, might be necessary for activation of ta rget genes like tartrate-resistant acid phosphatase (TRAP) in osteoclasts. Several lines of evidence were consistent with this model. The combination of MITF and PU.1 synergistically activated the TRAP promoter in transient a ssays. This activation was dependent on intact binding sites for both facto rs in the TRAP promoter. MITF and PU.1 physically interacted when coexpress ed in COS cells or in vitro when purified recombinant proteins were studied . The minimal regions of MITF and PU.1 required for the interaction were th e basic-helix-loop-helix zipper domain and the Ets DNA binding domain, resp ectively. Significantly, mice heterozygous for both the mutant mi allele an d a PU.1 null allele developed osteopetrosis early in life which resolved w ith age. The size and number of osteoclasts were not altered in the double heterozygous mutant mice, indicating that the defect lies in mature osteocl ast function. Taken in total, the results afford an example of how lineage- specific gene regulation can be achieved by the combinatorial action of two broadly expressed transcription factors.