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

E. chaffeensis induces autophagy independently of MTOR, ULK1, PRKA/AMPK, and ubiquitination pathways Macroautophagy induction by amino acid starvation is initiated by the activation of ULK, which is mainly regulated by MTOR kinase complex or by PRKA/AMPK (protein kinase, AMP-activated) independently from MTOR activity. 73, 74 MTOR inhibition leads to dephosphorylation of ULK1 at Ser757, 74 and of RPS6 (ribosomal protein S6) at Ser240/244. 75 PRKA is a heterotrimeric protein composed of a (PRKAA), b, and g subunits. The g subunit detects shifts in the AMP-to-ATP ratio and activates PRKA through phosphorylation of the a subunit at Thr172. 76 We therefore investigated whether E. chaffeensis induces host autophagy through these well-known signaling pathways using phosphorylation-specific antibodies that demonstrate the activation status of ULK1, PRKA, and RPS6. Western blotting results showed that tricyclic benzonaphthyridinone (Torin-1; a potent inhibitor of MTOR complex 1 [MTORC1] or MTORC2) 77 or rapamycin treatment activated ULK1 by dephosphorylation at Ser757 and RPS6 at Ser240/ 244 . , but not PRKAA at Thr172 (Fig. S6B) . However, infection of E. chaffeensis in THP-1 cells for 1 d did not alter the activation status of ULK1, RPS6, or PRKA (Fig. S6B) .