Promoter analysis of macrophage- and tick cell-specific differentially expressed Ehrlichia chaffeensis p28-Omp genes

Abstract

BackgroundEhrlichia chaffeensis is a rickettsial agent responsible for an emerging tick-borne illness, human monocytic ehrlichiosis. Recently, we reported that E. chaffeensis protein expression is influenced by macrophage and tick cell environments. We also demonstrated that host response differs considerably for macrophage and tick cell-derived bacteria with delayed clearance of the pathogen originating from tick cells.ResultsIn this study, we mapped differences in the promoter regions of two genes of p28-Omp locus, genes 14 and 19, whose expression is influenced by macrophage and tick cell environments. Primer extension and quantitative RT-PCR analysis were performed to map transcription start sites and to demonstrate that E. chaffeensis regulates transcription in a host cell-specific manner. Promoter regions of genes 14 and 19 were evaluated to map differences in gene expression and to locate RNA polymerase binding sites.ConclusionRNA analysis and promoter deletion analysis aided in identifying differences in transcription, DNA sequences that influenced promoter activity and RNA polymerase binding regions. This is the first description of a transcriptional machinery of E. chaffeensis. In the absence of available genetic manipulation systems, the promoter analysis described in this study can serve as a novel molecular tool for mapping the molecular basis for gene expression differences in E. chaffeensis and other related pathogens belonging to the Anaplasmataceae family.

Human Ehrlichiosis and Anaplasmosis

“…Evaluation of different E. chaffeensis isolates demonstrated a differentially expression of p28/30-Omp proteins in infected macrophages and tick cell cultures. In infected macrophages, the dominant E. chaffeensis expressed proteins are the products of the p28-Omp 19 and 20 genes (40, 41). In cultured tick cells derived from E. chaffeensis vector, Amblyomma americanum and non-vector ( Ixodes scapularis ) ticks, E. chaffeenesis expression consists only of the p28-Omp 14 protein (40, 41).…”

“…In infected macrophages, the dominant E. chaffeensis expressed proteins are the products of the p28-Omp 19 and 20 genes (40, 41). In cultured tick cells derived from E. chaffeensis vector, Amblyomma americanum and non-vector ( Ixodes scapularis ) ticks, E. chaffeenesis expression consists only of the p28-Omp 14 protein (40, 41). It is postulated that this differential expression of proteins within the p28/p30-Omp locus may be critical for the adaptation of Ehrlichia species to their different hosts (mammals and ticks).…”

“…To this end, a recent study by Ganta et al (42) determined that C57BL/6 mice have different responses to infection depending on the source of the inoculum. ISE6 tick cell-derived Ehrlichia inoculated into mice results in a more persistent infection, which includes relapses of increasing bacterial load On the other hand, the A. phagocytophilum genome has three omp-1 , one msp2, two msp2 homologs, one msp4 , and 113 p44 loci belonging to the OMP-1/MSP2/P44 superfamily (37–40). P44 plays a role in the binding of A. phagocytophilum to surfaces of neutrophils and in antigenic variation and evasion of host immune responses.…”

“…P44 plays a role in the binding of A. phagocytophilum to surfaces of neutrophils and in antigenic variation and evasion of host immune responses. The P44s are diverse, include several paralogs (p44-1 to p44-65) expressed in mammals and ticks and confer antigenic environmental adaptation, especially during tick transmission (37–40). …”

Human Ehrlichioses and Rickettsioses in Cameroon

“…They also have small subsets of genes associated with hostpathogen interactions (Ismail et al, 2010). E. chaffeensis have immunodominant outer membrane proteins (OMP-1/MSP2/P28) (Ohashi et al, 1998;Yu et al, 2000;Huang et al, 2008), and in infected macrophages ehrlichiae express the p28-Omp 19 and 20 genes as dominant protein products Peddireddi et al, 2009). Ehrlichiae also express several targets of the humoral immune response including tandem repeat and ankyrin repeat containing proteins (Yu et al, 1997;Sumner et al, 1999;McBride et al, 2003;McBride et al, 2007).…”