INTERACTION OF EHRLICHIA CHAFFEENSIS ANKYRIN REPEAT PROTEIN WITH REPETITIVE HOST GENOME ALU ELEMENTS

Bing Zhu, Ph.D, K.A. Nethery, M.S., J. Kuriakose, B.S., A, Wakeel, Ph.D, X.F. Zhang, M.S., J.W. McBride, Ph.D

Department of Pathology, UTMB

Background: Ehrlichiae are obligately intracellular bacteria that survive and persist in innate immune cells by modulating host cell gene transcription. Numerous ankyrin and tandem repeat-containing proteins have been identified in Ehrlichia spp., but their role in pathobiology is unknown. Objective: In the current study, we investigated the nuclear translocation of an immunoreactive ankyrin repeat-containing protein, p200, and its interaction with a small subset of p200 target genes. Methods: The nuclear translocation of p200 was demonstrated by immunofluorescent confocal microscopy and Western immunoblot using anti-p200 antibody. p200 bound DNA from nuclear extracts was obtained by chromatin immunoprecipitation (ChIP) and DNA sequencing (ChIP-Seq). Gene targets were determined using a whole genome promoter (5 kb) microarray and p200 interaction with specific DNA motifs was determined using electrophoretic mobility shift assays and NoShift assays. Results: ChIP and ChIP-Seq revealed an E. chaffeensis p200 interaction with an adenine-rich (mid A-stretch) motif within host promoter and intronic Alu elements, the most abundant repetitive elements in the human genome. Whole genome analysis with ChIP-chip determined that genes (n=456) with promoter Alu elements primarily related to transcription, apoptosis, ATPase activity were the primary targets of p200. Several p200 target genes associated with ehrlichial pathobiology were strongly upregulated during infection as determined by quantitative PCR. Conclusions: The p200 interaction with the host cell chromatin reveals a novel gene expression regulation strategy utilized by ehrlichiae to survive and persist in phagocytes. Supported by the grants from National Institutes of Allergy and Infectious Diseases grant, and additional support was provided by the Clayton Foundation for Research.