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

Lin, Mingqun; Liu, Hongyan; Xiong, Qingming; Niu, Hua; Cheng, Zhihui; Yamamoto, Akitsugu; Rikihisa, Yasuko

To determine whether host preincorporated amino acids are taken up by E. chaffeensis via autophagy, uninfected THP-1 cells were prelabeled with [ 3 H]-glutamine for 1 d and then infected with E. chaffeensis for 2 d in the absence of [ 3 H]-glutamine in the fresh culture medium. The cells were then incubated with or without 3-MA for an additional 6 h prior to harvesting at 2 d p.i. Host preincorporated L-glutamine was indeed taken up by E. chaffeensis, and 3-MA significantly blocked this uptake (Fig. 5A) . The results suggest that class III PtdIns3K activity is critical for E. chaffeensis to have access to host-derived amino acids. Next, we examined whether the simultaneous addition of essential amino acids and 3-MA to infected cells would abrogate the 3-MA-induced inhibition of E. chaffeensis replication. Our result showed that E. chaffeensis growth inhibition by 3-MA was partially rescued ( Fig. 5B and C). Taken together, E. chaffeensis dysregulates the homeostasis of cellular free amino acid levels and raises cellular free glutamine and glutamate levels during exponential growth, and E. chaffeensis within membrane-bound inclusions has the ability to take up free amino acids from host cells in a class III PtdIns3K activity-dependent manner.