RESTRICTION OF INTERFERON-GAMMA RESPONSIVENESS AND BASAL EXPRESSION OF THE MYELOID HUMAN FC-GAMMA-R1B GENE IS MEDIATED BY A FUNCTIONAL PU.1SITE AND A TRANSCRIPTION INITIATOR CONSENSUS

The restricted expression of the human Fc gamma R1b gene to myeloid ce lls is likely to be regulated by a combination of transcription factor s that may not be solely expressed in myeloid cells, but act together to restrict the expression of the gene to myeloid cells. Low basal exp ression of the human Fc gamma R1b gene is specifically upregulated by interferon gamma (IFN-gamma). A 181-bp region of 5' flanking sequence contains several key regulatory motifs that include the extended gamma response region (XGRR) and the PIE region. The XGRR contains the 39-b p gamma response region originally defined in the highly homologous Fc gamma R1a gene. The XGRR is in close proximity to the 21-bp PIE motif that is conserved in the promoters of some other myeloid genes. The P IE motif contains a consensus site for the macrophage and B cell trans cription factor, PU.1, and is adjacent to the cluster of transcription start sites. An active transcription initiator, Inr, consensus spans the start sites and appears to direct transcription initiation of this TATA-less gene. In this study, we demonstrate that the PIE region con tains a functional PU.1 site that binds a human PU.1-like protein and that associated factors present in myeloid extracts also bind in this PIE region. Mutational analysis reveals an absolute requirement for an intact PU.1 box for both basal and IFN-gamma inducible expression of this gene. In addition, mutations in the Inr greatly reduce basal and inducible transcription. Insertion of a strong TATA box downstream fro m the Inr or at -30 bp from the transcription start sites restores bas al and inducible activity in the presence of a mutated PU.1 site. We a lso demonstrate that indeed, when the XGRR is positioned in the contex t of a heterologous TATA containing promoter, it is able to respond eq uivalently to either IFN-alpha or IFN-gamma. However, IFN-alpha respon siveness does not occur in the context of the physiological Fc gamma R 1b TATA-less basal promoter. Our results suggest that a human PU.1-lik e factor acts as a ''bridging factor'' between the upstream IFN-gamma enhancer and the Inr dependent preinitiation complex. These findings i ndicate that the structure of the basal promoter in combination with r estricted activators like PU.1 are important in regulating the express ion of this gene.