Differential protein binding to the c-myc promoter during differentiation of hematopoietic cell lines

Abstract

In vivo footprinting has been used to examine protein binding sites in the c-myc promoter during differentiation of HL60 and K562 cell lines. The c-myc gene is expressed in proliferating cells, but c-myc levels decrease dramatically during differentiation. A number of potential protein binding sites have been identified from in vitro studies of the c-myc promoter, but very little is known about occupancy of these sites in vivo. We have identified in vivo footprints at DNase hypersensitive sites II2 and III1 which disappear during differentiation, while a footprint at site IV is present only in differentiated cells. Footprints at DNase hypersensitive sites I and II1 do not change with differentiation. A protected region near DNase hypersensitive site III2 is present in both undifferentiated and differentiated cells, but it extends further 5' in undifferentiated cells. From the protected sequences we have been able to identify candidate transcription factors likely to be involved in the control of c-myc expression. By electrophoretic mobility shift assay we have demonstrated that a protein binds to the sequence at site IV. We have also examined the 3' end of the first exon and the 5' end of intron I and do not find any evidence for protein binding sites in this region that was thought to be important for the block to transcription elongation during differentiation.