Host cell RhoA and Rac1 were activated after T. gondii invasion. The decisive domains for the RhoA accumulation on the PVM were identified as the GTP/Mg2+ binding site, the mDia effector interaction site, the G1 box, the G2 box and the G5 box, respectively, which were related to the binding of GTP for enzymatic activity and to mDia for the regulation of microtubules. The reorganization of host cell cytoskeleton facilitates the PV formation and enlargement in the host cell. The recruited RhoA on the PVM could not be activated by epithelial growth factor (EGF) and no translocation was
observed, which indicated that the recruited RhoA or Rac1 on the PVM might be in GTP-bound active form. Wild-type RhoA or Rac1 overexpressed cells
had almost the same infection Eltanexor rates by T. gondii as the mock-treated cells, while RhoA-N19 or Rac1-N17 transfected cells and RhoA or Rac1 siRNA- and RhoA + Rac1 siRNA-treated cells showed significantly diminished infection rates than mock cells, which indicated that the normal activity of RhoA and Rac1 GTPases are indispensable to the internalization of the tachyzoite. The accumulation of the RhoA and Rac1 on the PVM and the requisite of their normal GTPase activities for efficient invasion implied their involvement and function in T. gondii invasion. The summary of the host cell RhoA and Rac1 cell signaling involved in the T. gondii invasion is show in Figure 8. Acknowledgement AZD7762 manufacturer This work was supported by National Natural Science Foundation of China (No. 81071377, 81271866), the Research Fund for the Doctoral Program of Higher Education of China (20104433120014), Guangdong provincial Masitinib (AB1010) key scientific and technological project to HJP (2011B010500003), Guangdong Province talent introduction of special funds (2011–26), the Guangdong Province College Students Renovation
Experimental Program (1212111020) and the Grant from the School of Public Health and Tropical Medicine of Southern Medical University (GW201110) to HJ Peng; Province Universities and Colleges Pearl River Scholar Funded Scheme (2009) and National Natural Science Foundation of China (Key program:31030066) to XG Chen. Electronic supplementary material Additional file 1: Data S1. The florescence images of the real-time observation of the cell invasion by T. gondii. The invasion position was indicated with a purple arrowhead. The green florescence pictures showed the accumulation of the CFP-tagged RhoA to the PVM (purple arrowhead) at the time points of -10 min (5 min post infection), -5 min (10 min post infection), 0 min (15 min post infection), 5 min (20 min post infection), 10 min (25 min post infection) and 15 min (30 min post infection). The focal point of RhoA at the immediate point of invasion on the host cell membrane is not visible. (JPG 412 KB) Additional file 2: Data S2. The DIC images of the real-time observation of the cell invasion by T. gondii.