Ehrlichia chaffeensis inclusions are early endosomes which selectively accumulate transferrin receptor

Abstract

Ehrlichia chaffeensis is an obligatory intracellular bacterium which infects macrophages and monocytes. Double immunofluorescence labeling was used to characterize the nature of E. chaffeensis inclusion in the human promyelocytic leukemia cell line THP-1. E. chaffeensis was labeled with dog anti-E. chaffeensis serum and fluorescein isothiocyanate-conjugated anti-dog immunoglobulin G (IgG). Lissamine rhodamine-conjugated anti-mouse IgG was used to label various mouse monoclonal antibodies. Ehrlichial inclusions did not fuse with lysosomes, since they were not labeled with anti-CD63 or anti-LAMP-1. The ehrlichial inclusions were slightly acidic, since they weakly accumulated 3-(2,4-dinitroanilino)-3-amino-N-methyldipropylamine and stained weakly positive for vacuolar type H ؉ ATPase. Some ehrlichial inclusions were labeled positive with antibodies against HLA-DR, HLA-ABC, and ␤ 2 microglobulin, while other inclusions in the same cell were labeled negative. The inclusions were labeled strongly positive for transferrin receptors (TfRs) and negative for the clathrin heavy chain. Time course labeling for TfRs showed that up to 3 h postinfection, most of the ehrlichial inclusions were negative for TfRs. After 6 h postinfection, 100% of the ehrlichial inclusions became TfR positive and the intensity of labeling was increased during the subsequent 3 days. Reverse transcription-PCR showed a gradual increase in the level of TfR mRNA postinfection, which reached a peak at 24 h postinfection. These results suggest that ehrlichial inclusions are early endosomes which selectively accumulate TfRs and that the ehrlichiae up-regulate TfR mRNA expression.

Alteration of epithelial cell transferrin-iron homeostasis by Neisseria meningitidis and Neisseria gonorrhoeae

“…Several other pathogens alter TfR expression and steady-state levels, as well as its cellular distribution in host cells. Ehrlichia chaffeensis induces an increase in TfR mRNA levels in THP-1 cells (Barnewall et al, 1997). Coxiella burnetii induces an increase in TfR steady-state levels in J774A1 human macrophages (Howe and Mallavia, 1999).…”

“…Coxiella burnetii induces an increase in TfR steady-state levels in J774A1 human macrophages (Howe and Mallavia, 1999). E. chaffeensis (THP-1 cells) (Barnewall et al, 1997), Salmonella typhimurium (HeLa cells) (Steele-Mortimer et al, 1999), Chlamydia trachomatis (HeLa cells) (van Ooij et al, 1997) and Leishmania donovani (murine macrophages) (Borges et al, 1998) redirect TfR trafficking towards the phagosomes. These interactions are likely to promote the uptake of Tf-iron by the microbes within these host cells.…”

Ehrlichiasecretes Etf-1 to induce autophagy and capture nutrients for its growth through RAB5 and class III phosphatidylinositol 3-kinase

“…The intracellular membrane compartment (inclusion) that contains E. chaffeensis has early endosome-like characteristics, including the small GTPase RAB5, a RAB5 effector EEA1 (early endosome antigen 1), TF (transferrin), TFRC (transferrin receptor), and vacuolar-type H C -ATPase, but it lacks late endosomal or lysosomal markers or NADPH oxidase components. [8][9][10] Within this compartment, E. chaffeensis acquires all nutrients for its reproduction to form numerous mature infectious forms. E. chaffeensis has a small genome of 1.176 Mb with a limited capacity for biosynthesis and metabolism.…”

“…Although the host cell cytoplasm is rich with these nutrients, it is unlikely that the inclusion membrane is leaky, as ehrlichial inclusions maintain a weakly acidic intralumenal pH. 8 It is also unlikely that varieties of active transporters are synthesized and assembled in the proper orientation on the inclusion membrane to import host nutrients during ehrlichial replication. Considering these limitations, it is possible that a novel mechanism has evolved in this group of obligatory intracellular bacteria to acquire nutrients.…”

The developmental cycle of Ehrlichia chaffeensis in vertebrate cells

“…Ehrlichia chaffeensis is an obligatory intracellular bacterium that resides in early endosomes of mononuclear phagocytes (Barnewall et al, 1997). The life cycle of E. chaffeensis involves a tick vector and a mammalian host.…”