Three putative T4S effectors, including ECH0825, are expressed by E. chaffeensis in THP-1 cells.
A. ECH0261, ECH0767, and ECH0825 are transcribed by E. chaffeensis in THP-1 cells. M, molecular size marker; + and − indicate the presence or absence of reverse transcriptase, respectively; D, positive control (chromosomal DNA used as template) for PCR. Corresponding genes and sizes (in base pairs) of amplified products are indicated. No amplicon was detected without reverse transcriptase, indicating that the RNA preparation was not contaminated with genomic DNA.
B. rECH0825 (C-terminal 250 residues of ECH0825) expressed in E. coli was purified by immobilized Ni2+ affinity chromatography and subjected to SDS-PAGE followed by GelCode blue staining. M, protein molecular mass marker.
C. Western blot analysis of rECH0825, uninfected THP-1 cells, E. chaffeensis (ECH)-infected THP-1 cells, and purified ECH organisms using rabbit anti-ECH0825 antiserum. Molecular mass markers are indicated at the left. Arrows indicate rECH0825 and native ECH0825.