ABSTRACTCoevolution of intracellular bacterial pathogens and their host cells resulted in the appearance of effector molecules that when translocated into the host cell modulate its function, facilitating bacterial survival within the hostile host environment. Some of these effectors interact with host chromatin and other nuclear components. In this report, we show that the AnkA protein ofAnaplasma phagocytophilum, which is translocated into the host cell nucleus, interacts with gene regulatory regions of host chromatin and is involved in downregulating expression ofCYBB(gp91phox) and other key host defense genes. AnkA effector protein rapidly accumulated in nuclei of infected cells coincident with changes inCYBBtranscription. AnkA interacted with transcriptional regulatory regions of theCYBBlocus at sites where transcriptional regulators bind. AnkA binding to DNA occurred at regions with high AT contents. Mutation of AT stretches at these sites abrogated AnkA binding. Histone H3 acetylation decreased dramatically at theCYBBlocus duringA. phagocytophiluminfection, particularly around AnkA binding sites. Transcription ofCYBBand other defense genes was significantly decreased in AnkA-transfected HL-60 cells. These data suggest a mechanism by which intracellular pathogens directly regulate host cell gene expression mediated by nuclear effectors and changes in host chromatin structure.